Does electricity cause damage on a cellular level?

Does electricity cause damage on a cellular level?

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As I understand it the mechanism of death when a mammal is electrocuted is that the current disrupts the SAN/AVN in the heart causing it to fibrilate or arrest. That's on a macro scale, however. What damage, if any, does electricity cause on a cellular level? I've noticed some sort of moss or lichen growing on the third rail of the train lines so I'm aware that they must be able to cope with the current, do they have adaptations ot allow this or is it just that they are not connected to the earth?

Regarding the moss or lichen on the third rail on British train lines: there is no current passing through the moss/lichen as they are not completing a circuit. They are just sitting on one connector. If another connector passed over them, completing the circuit across them, then a current might pass across them for a very short time. But I doubt they grow on the top of the rail (which is the contact part in the UK system) as that is constantly being polished smooth by the contacts from trains sliding across it.

Regarding cell damage: Apart from fibrillation in animals, and burns caused by Joule heating, electricity does cause cellular damage.

At low frequencies (<10kHz), electricity disrupts cell membranes and makes them much more permeable (we actually harness this when we use electroporation to transform bacteria). All organisms rely on electrochemical potential differences across membranes for their metabolism (Berry, 2002). The exact effect is dependent on the previous electrical state of the cells, for example the electrical potential difference across the membrane, and on the surface area to volume ratio of the cell (greater volume relative to surface area leads to more disruption). In any case, extreme electroporation can cause solutes to flow in or out of a cell, and generally disrupt the balance of solute concentrations and cause organelles and other bodies to move out of a cell. When the electrical stimulation stops, the contents are then fixed in a disrupted state. The cell then has to expend enormous amounts of ATP using ion channels and transport proteins in an attempt to reinstate the necessary chemiosmotic potentials, and in doing so exhausts the entire ATP supply and goes into biochemical arrest (no metabolism occurs), which is when a cell is dead. Dead cells break apart because there are is no maintenance occuring.

At higher frequencies (10-100kHz) proteins become permanently denatured. Many proteins carry charges which give them an overall polarity. When placed in an electric field, the proteins reorient themselves and will undergo conformational change to achieve the optimum dipole moment in the direction of the field. Ion channels and pumps are particularly sensitive to these disruptions (since their charges are crucial to their function).

Rather than provide lots of references, there is one excellent review from which I drew all this information, and which you should read for more of the physical detail (Lee et al., 2000).


  • Berry, S. (2002) The Chemical Basis of Membrane Bioenergetics. Journal of Molecular Evolution. [Online] 54 (5), 595-613. Available from: doi:10.1007/s00239-001-0056-3 [Accessed: 9 February 2012].
  • Lee, R.C., Zhang, D. & Hannig, J. (2000) Biophysical injury mechanisms in electrical shock trauma. Annual Review of Biomedical Engineering. [Online] 2 (1), 477-509. Available from: doi:10.1146/annurev.bioeng.2.1.477 [Accessed: 9 February 2012].

Cell damage

Cell damage (also known as cell injury) is a variety of changes of stress that a cell suffers due to external as well as internal environmental changes. Amongst other causes, this can be due to physical, chemical, infectious, biological, nutritional or immunological factors. Cell damage can be reversible or irreversible. Depending on the extent of injury, the cellular response may be adaptive and where possible, homeostasis is restored. [1] Cell death occurs when the severity of the injury exceeds the cell's ability to repair itself. [2] Cell death is relative to both the length of exposure to a harmful stimulus and the severity of the damage caused. [1] Cell death may occur by necrosis or apoptosis..

Most bacteria are either beneficial or harmless to humans – those that cause disease are pathogens:

- The symptoms of the disease are usually caused by waste products of the pathogens

- An infection is when the effects are noticeable on the body

- Transmission is when an infection is passed on to somebody else

Diseases such as typhoid and cholera are transmitted through water, and can cause diarrhoea

To avoid water contamination, water-treatment processes take place

Food-borne infections including Salmonella are spread in two ways:

- By not cooking food thoroughly (e.g. raw eggs: newly laid eggs may be contaminated with poultry faeces)

- By contaminating cooked meat from handling raw meat first e.g. chicken

Air-borne infections are spread when an infected person coughs, sneezes, talks or breathes, as the pathogens are passed into the air in small droplets saliva, mucus and water

Infections that can be transmitted by direct contact are said to be contagious

Insect bites can transmit pathogens through the saliva of the insect

Pathogenicity is the ability of a bacterium to cause disease.

- The way in which the bacterium attaches and gains entry to host cells

- The types of toxin produced by the bacterium

- The infectivity of the bacterium (the number needed to cause an infection)

- The invasiveness of the bacterium (its ability to spread within the host)

After infection, a pathogen must do three things in order to produce a disease:

Disturbance of the immune system by electromagnetic fields—A potentially underlying cause for cellular damage and tissue repair reduction which could lead to disease and impairment

A number of papers dealing with the effects of modern, man-made electromagnetic fields (EMFs) on the immune system are summarized in the present review. EMFs disturb immune function through stimulation of various allergic and inflammatory responses, as well as effects on tissue repair processes. Such disturbances increase the risks for various diseases, including cancer. These and the EMF effects on other biological processes (e.g. DNA damage, neurological effects, etc.) are now widely reported to occur at exposure levels significantly below most current national and international safety limits. Obviously, biologically based exposure standards are needed to prevent disruption of normal body processes and potential adverse health effects of chronic exposure.

Based on this review, as well as the reviews in the recent Bioinitiative Report [] [C.F. Blackman, M. Blank, M. Kundi, C. Sage, D.O. Carpenter, Z. Davanipour, D. Gee, L. Hardell, O. Johansson, H. Lai, K.H. Mild, A. Sage, E.L. Sobel, Z. Xu, G. Chen, The Bioinitiative Report—A Rationale for a Biologically-based Public Exposure Standard for Electromagnetic Fields (ELF and RF), 2007)], it must be concluded that the existing public safety limits are inadequate to protect public health, and that new public safety limits, as well as limits on further deployment of untested technologies, are warranted.


Developmental bioelectricity is a sub-discipline of biology, related to, but distinct from, neurophysiology and bioelectromagnetics. Developmental bioelectricity refers to the endogenous ion fluxes, transmembrane and transepithelial voltage gradients, and electric currents and fields produced and sustained in living cells and tissues. [2] [3] This electrical activity is often used during embryogenesis, regeneration, and cancer - it is one layer of the complex field of signals that impinge upon all cells in vivo and regulate their interactions during pattern formation and maintenance (Figure 1). This is distinct from neural bioelectricity (classically termed electrophysiology), which refers to the rapid and transient spiking in well-recognized excitable cells like neurons and myocytes [4] and from bioelectromagnetics, which refers to the effects of applied electromagnetic radiation, and endogenous electromagnetics such as biophoton emission and magnetite. [5] [6]

The inside/outside discontinuity at the cell surface enabled by a lipid bilayer membrane (capacitor) is at the core of bioelectricity. The plasma membrane was an indispensable structure for the origin and evolution of life itself. It provided compartmentalization permitting the setting of a differential voltage/potential gradient (battery or voltage source) across the membrane, probably allowing early and rudimentary bioenergetics that fueled cell mechanisms. [9] [10] During evolution, the initially purely passive diffusion of ions (charge carriers), become gradually controlled by the acquisition of ion channels, pumps, exchangers, and transporters. These energetically free (resistors or conductors, passive transport) or expensive (current sources, active transport) translocators set and fine tune voltage gradients – resting potentials – that are ubiquitous and essential to life's physiology, ranging from bioenergetics, motion, sensing, nutrient transport, toxins clearance, and signaling in homeostatic and disease/injury conditions. Upon stimuli or barrier breaking (short-circuit) of the membrane, ions powered by the voltage gradient (electromotive force) diffuse or leak, respectively, through the cytoplasm and interstitial fluids (conductors), generating measurable electric currents – net ion fluxes – and fields. Some ions (such as calcium) and molecules (such as hydrogen peroxide) modulate targeted translocators to produce a current or to enhance, mitigate or even reverse an initial current, being switchers. [11] [12]

Endogenous bioelectric signals are produced in cells by the cumulative action of ion channels, pumps, and transporters. In non-excitable cells, the resting potential across the plasma membrane (Vmem) of individual cells propagate across distances via electrical synapses known as gap junctions (conductors), which allow cells to share their resting potential with neighbors. Aligned and stacked cells (such as in epithelia) generate transepithelial potentials (battery in series) and electric fields (Figures 2 and 3), which likewise propagate across tissues. [13] Tight junctions (resistors) efficiently mitigate the paracellular ion diffusion and leakage, precluding the voltage short circuit. Together, these voltages and electric fields form rich and dynamic and patterns (Figure 5) inside living bodies that demarcate anatomical features, thus acting like blueprints for gene expression and morphogenesis in some instances. More than correlations, these bioelectrical distributions are dynamic, evolving with time and with the microenvironment and even long-distant conditions to serve as instructive influences over cell behavior and large-scale patterning during embryogenesis, regeneration, and cancer suppression. [3] [14] [8] [15] [16] Bioelectric control mechanisms are an important emerging target for advances in regenerative medicine, birth defects, cancer, and synthetic bioengineering. [17] [18]

The modern roots of developmental bioelectricity can be traced back to the entire 18th century. Several seminal works stimulating muscle contractions using Leyden jars culminated with the publication of classical studies by Luigi Galvani in 1791 (De viribus electricitatis in motu musculari) and 1794. In these, Galvani thought to have uncovered intrinsic electric-producing ability in living tissues or “animal electricity”. Alessandro Volta showed that the frog's leg muscle twitching was due to a static electricity generator and from dissimilar metals contact. Galvani showed, in a 1794 study, twitching without metal electricity by touching the leg muscle with a deviating cut sciatic nerve, definitively showing “animal electricity”. [19] [20] [21] Unknowingly, Galvani with this and related experiments discovered the injury current (ion leakage driven by the intact membrane/epithelial potential) and injury potential (potential difference between injured and intact membrane/epithelium). The injury potential was, in fact, the electrical source behind the leg contraction, as realized in the next century. [22] [23] Subsequent work ultimately extended this field broadly beyond nerve and muscle to all cells, from bacteria to non-excitable mammalian cells.

Building on earlier studies, further glimpses of developmental bioelectricity occurred with the discovery of wound-related electric currents and fields in the 1840s, when one of the founding fathers of modern electrophysiology – Emil du Bois-Reymond – reported macroscopic level electrical activities in frog, fish and human bodies. He recorded minute electric currents in live tissues and organisms with a then state-of-the-art galvanometer made of insulated copper wire coils. He unveiled the fast-changing electricity associated with muscle contraction and nerve excitation – the action potentials. [24] [25] [26] At the same time, du Bois-Reymond also reported in detail less fluctuating electricity at wounds – injury current and potential – he made to himself. [27] [28]

Bioelectricity work began in earnest at the beginning of the 20th century. [30] [31] [32] [33] [34] [35] Since then, several waves of research produced important functional data showing the role that bioelectricity plays in the control of growth and form. In the 1920s and 1930s, E. J. Lund [36] and H. S. Burr [37] were some of the most prolific authors in this field. [29] Lund measured currents in a large number of living model systems, correlating them to changes in patterning. In contrast, Burr used a voltmeter to measure voltage gradients, examining developing embryonic tissues and tumors, in a range of animals and plants. Applied electric fields were demonstrated to alter the regeneration of planaria by Marsh and Beams in the 1940s and 1950s, [38] [39] inducing the formation of heads or tails at cut sites, reversing the primary body polarity. The introduction and development of the vibrating probe, the first device for quantitative non-invasive characterization of the extracellular minute ion currents, by Lionel Jaffe and Richard Nuccittelli, [40] revitalized the field in the 1970s. They were followed by researchers such as Joseph Vanable, Richard Borgens, Ken Robinson, and Colin McCaig, among many others, who showed roles of endogenous bioelectric signaling in limb development and regeneration, embryogenesis, organ polarity, and wound healing. [41] [42] [43] [44] [45] [46] [23] [47] C.D. Cone studied the role of resting potential in regulating cell differentiation and proliferation [48] [49] and subsequent work [50] has identified specific regions of the resting potential spectrum that correspond to distinct cell states such as quiescent, stem, cancer, and terminally differentiated (Figure 5).

Although this body of work generated a significant amount of high-quality physiological data, this large-scale biophysics approach has historically been in the shadow of the limelight of biochemical gradients and genetic networks in biology education, funding, and overall popularity among biologists. A key factor that contributed to this field lagging behind molecular genetics and biochemistry is that bioelectricity is inherently a living phenomenon – it cannot be studied in fixed specimens. Working with bioelectricity is more complex than traditional approaches to developmental biology, both methodologically and conceptually, as it typically requires a highly interdisciplinary approach. [15]

The gold standard techniques to quantitatively extract electric dimensions from living specimens, ranging from cell to organism levels, are the glass microelectrode (or micropipette), the vibrating (or self-referencing) voltage probe, and the vibrating ion-selective microelectrode. The former is inherently invasive and the two latter are non-invasive, but all are ultra-sensitive [51] and fast-responsive sensors extensively used in a plethora of physiological conditions in widespread biological models. [52] [53] [11] [54] [23]

The glass microelectrode was developed in the 1940s to study the action potential of excitable cells, deriving from the seminal work by Hodgkin and Huxley in the giant axon squid. [55] [56] It is simply a liquid salt bridge connecting the biological specimen with the electrode, protecting tissues from leachable toxins and redox reactions of the bare electrode. Owing to its low impedance, low junction potential and weak polarization, silver electrodes are standard transducers of the ionic into electric current that occurs through a reversible redox reaction at the electrode surface. [57]

The vibrating probe was introduced in biological studies in the 1970s. [58] [59] [40] The voltage-sensitive probe is electroplated with platinum to form a capacitive black tip ball with large surface area. When vibrating in an artificial or natural DC voltage gradient, the capacitive ball oscillates in a sinusoidal AC output. The amplitude of the wave is proportional to the measuring potential difference at the frequency of the vibration, efficiently filtered by a lock-in amplifier that boosts probe's sensitivity. [40] [60] [61]

The vibrating ion-selective microelectrode was first used in 1990 to measure calcium fluxes in various cells and tissues. [62] The ion-selective microelectrode is an adaptation of the glass microelectrode, where an ion-specific liquid ion exchanger (ionophore) is tip-filled into a previously silanized (to prevent leakage) microelectrode. Also, the microelectrode vibrates at low frequencies to operate in the accurate self-referencing mode. Only the specific ion permeates the ionophore, therefore the voltage readout is proportional to the ion concentration in the measuring condition. Then, flux is calculated using the Fick's first law. [60] [63]

Emerging optic-based techniques, [64] for example, the pH optrode (or optode), which can be integrated into a self-referencing system may become an alternative or additional technique in bioelectricity laboratories. The optrode does not require referencing and is insensitive to electromagnetism [65] simplifying system setting up and making it a suitable option for recordings where electric stimulation is simultaneously applied.

Much work to functionally study bioelectric signaling has made use of applied (exogenous) electric currents and fields via DC and AC voltage-delivering apparatus integrated with agarose salt bridges. [66] These devices can generate countless combinations of voltage magnitude and direction, pulses, and frequencies. Currently, lab-on-a-chip mediated application of electric fields is gaining ground in the field with the possibility to allow high-throughput screening assays of the large combinatory outputs. [67]

The remarkable progress in molecular biology over the last six decades has produced powerful tools that facilitate the dissection of biochemical and genetic signals yet, they tend to not be well-suited for bioelectric studies in vivo. Prior work relied extensively on current applied directly by electrodes, reinvigorated by significant recent advances in materials science [69] [70] [71] [72] [73] [74] and extracellular current measurements, facilitated by sophisticated self-referencing electrode systems. [75] [76] While electrode applications for manipulating neutrally-controlled body processes have recently attracted much attention, [77] [78] the nervous system is just the tip of the iceberg [ peacock term ] when it comes to the opportunities for controlling somatic processes, as most cell types are electrically active and respond to ionic signals from themselves and their neighbors (Figure 6).

In the last 15 years, a number of new molecular techniques [79] have been developed that allowed bioelectric pathways to be investigated with a high degree of mechanistic resolution, and to be linked to canonical molecular cascades. These include (1) pharmacological screens to identify endogenous channels and pumps responsible for specific patterning events [80] [81] [82] (2) voltage-sensitive fluorescent reporter dyes and genetically-encoded fluorescent voltage indicators for the characterization of the bioelectric state in vivo [83] [84] [85] [86] [87] (3) panels of well-characterized dominant ion channels that can be misexpressed in cells of interest to alter the bioelectric state in desired ways [82] [88] [89] and (4) computational platforms that are coming on-line [90] [91] to assist in building predictive models of bioelectric dynamics in tissues. [92] [93] [94]

Compared with the electrode-based techniques, the molecular probes provide a wider spatial resolution and facilitated dynamic analysis over time. Although calibration or titration can be possible, molecular probes are typically semi-quantitative, whereas electrodes provide absolute bioelectric values. Another advantage of fluorescence and other probes is their less-invasive nature and spatial multiplexing, enabling the simultaneous monitoring of large areas of embryonic or other tissues in vivo during normal or pathological pattering processes. [95]

Work in model systems such as Xenopus laevis and zebrafish has revealed a role for bioelectric signaling in the development of heart, [96] [97] face, [98] [99] eye, [88] brain, [100] [101] and other organs. Screens have identified roles for ion channels in size control of structures such as the zebrafish fin, [102] while focused gain-of-function studies have shown for example that bodyparts can be re-specified at the organ level – for example creating entire eyes in gut endoderm. [88] As in the brain, developmental bioelectrics can integrate information across significant distance in the embryo, for example such as the control of brain size by bioelectric states of ventral tissue. [101] and the control of tumorigenesis at the site of oncogene expression by bioelectric state of remote cells. [103] [104]

Human disorders, as well as numerous mouse mutants show that bioelectric signaling is important for human development (Tables 1 and 2). Those effects are pervasively linked to channelopathies, which are human disorders that result from mutations that disrupt ion channels.

Several channelopathies result in morphological abnormalities or congenital birth defects in addition to symptoms that affect muscle and or neurons. For example, mutations that disrupt an inwardly rectifying potassium channel Kir2.1 cause dominantly inherited Andersen-Tawil Syndrome (ATS). ATS patients experience periodic paralysis, cardiac arrhythmias, and multiple morphological abnormalities that can include cleft or high arched palate, cleft or thin upper lip, flattened philtrum, micrognathia, dental oligodontia, enamel hypoplasia, delayed dentition eruption, malocclusion, broad forehead, wide set eyes, low set ears, syndactyly, clinodactyly, brachydactyly, and dysplastic kidneys. [105] [106] Mutations that disrupt another inwardly rectifying K+ channel Girk2 encoded by KCNJ6 cause Keppen-Lubinsky syndrome which includes microcephaly, a narrow nasal bridge, a high arched palate, and severe generalized lipodystrophy (failure to generate adipose tissue). [107] KCNJ6 is in the Down syndrome critical region such that duplications that include this region lead to craniofacial and limb abnormalities and duplications that do not include this region do not lead to morphological symptoms of Down syndrome. [108] [109] [110] [111] Mutations in KCNH1, a voltage gated potassium channel lead to Temple-Baraitser (also known as Zimmermann- Laband) syndrome. Common features of Temple-Baraitser syndrome include absent or hypoplastic of finger and toe nails and phalanges and joint instability. Craniofacial defects associated with mutations in KCNH1 include cleft or high arched palate, hypertelorism, dysmorphic ears, dysmorphic nose, gingival hypertrophy, and abnormal number of teeth. [112] [113] [114] [115] [116] [117] [118]

Mutations in CaV1.2, a voltage gated Ca2+ channel, lead to Timothy syndrome which causes severe cardiac arrhythmia (long-QT) along with syndactyly and similar craniofacial defects to Andersen-Tawil syndrome including cleft or high-arched palate, micrognathia, low set ears, syndactyly and brachydactyly. [119] [120] While these channelopathies are rare, they show that functional ion channels are important for development. Furthermore, in utero exposure to anti-epileptic medications that target some ion channels also cause increased incidence of birth defects such as oral clefting. [121] [122] [123] [124] [125] The effects of both genetic and exogenous disruption of ion channels lend insight into the importance of bioelectric signaling in development.

One of the best-understood roles for bioelectric gradients is at the tissue-level endogenous electric fields utilized during wound healing. It is challenging to study wound-associated electric fields, because these fields are weak, less fluctuating, and do not have immediate biological responses when compared to nerve pulses and muscle contraction. The development of the vibrating and glass microelectrodes, demonstrated that wounds indeed produced and, importantly, sustained measurable electric currents and electric fields. [40] [126] [59] [127] [128] [129] These techniques allow further characterization of the wound electric fields/currents at cornea and skin wounds, which show active spatial and temporal features, suggesting active regulation of these electrical phenomena. For example, the wound electric currents are always the strongest at the wound edge, which gradually increased to reach a peak about 1 hour after injury. [130] [131] [61] At wounds in diabetic animals, the wound electric fields are significantly compromised. [132] Understanding the mechanisms of generation and regulation of the wound electric currents/fields is expected to reveal new approaches to manipulate the electrical aspect for better wound healing.

How are the electric fields at a wound produced? Epithelia actively pump and differentially segregate ions. In the cornea epithelium, for example, Na+ and K+ are transported inwards from tear fluid to extracellular fluid, and Cl− is transported out of the extracellular fluid into the tear fluid. The epithelial cells are connected by tight junctions, forming the major electrical resistive barrier, and thus establishing an electrical gradient across the epithelium – the transepithelial potential (TEP). [133] [134] Breaking the epithelial barrier, as occurs in any wounds, creates a hole that breaches the high electrical resistance established by the tight junctions in the epithelial sheet, short-circuiting the epithelium locally. The TEP therefore drops to zero at the wound. However, normal ion transport continues in unwounded epithelial cells beyond the wound edge (typically <1 mm away), driving positive charge flow out of the wound and establishing a steady, laterally-oriented electric field (EF) with the cathode at the wound. Skin also generates a TEP, and when a skin wound is made, similar wound electric currents and fields arise, until the epithelial barrier function recovers to terminate the short-circuit at the wound. When wound electric fields are manipulated with pharmacological agents that either stimulate or inhibit transport of ions, the wound electric fields also increase or decrease, respectively. Wound healing can be speed up or slowed down accordingly in cornea wounds. [130] [131] [135]

How do electric fields affect wound healing? To heal wounds, cells surrounding the wound must migrate and grow directionally into the wound to cover the defect and restore the barrier. Cells important to heal wounds respond remarkably well to applied electric fields of the same strength that are measured at wounds. The whole gamut of cell types and their responses following injury are affected by physiological electric fields. Those include migration and division of epithelial cells, sprouting and extension of nerves, and migration of leukocytes and endothelial cells. [136] [137] [138] [139] The most well studied cellular behavior is directional migration of epithelial cells in electric fields – electrotaxis. The epithelial cells migrate directionally to the negative pole (cathode), which at a wound is the field polarity of the endogenous vectorial electric fields in the epithelium, pointing (positive to negative) to the wound center. Epithelial cells of the cornea, keratinocytes from the skin, and many other types of cells show directional migration at electric field strengths as low as a few mV mm−1. [140] [141] [142] [143] Large sheets of monolayer epithelial cells, and sheets of stratified multilayered epithelial cells also migrate directionally. [131] [144] Such collective movement closely resembles what happens during wound healing in vivo, where cell sheets move collectively into the wound bed to cover the wound and restore the barrier function of the skin or cornea.

How cells sense such minute extracellular electric fields remains largely elusive. Recent research has started to identify some genetic, signaling and structural elements underlying how cells sense and respond to small physiological electric fields. These include ion channels, intracellular signaling pathways, membrane lipid rafts, and electrophoresis of cellular membrane components. [145] [146] [147] [148] [149] [150] [151]

In the early 20th century, Albert Mathews seminally correlated regeneration of a cnidarian polyp with the potential difference between polyp and stolon surfaces, and affected regeneration by imposing countercurrents. Amedeo Herlitzka, following on the wound electric currents footsteps of his mentor, du Bois-Raymond, theorized about electric currents playing an early role in regeneration, maybe initiating cell proliferation. [152] Using electric fields overriding endogenous ones, Marsh and Beams astoundingly generated double-headed planarians and even reversed the primary body polarity entirely, with tails growing where a head previously existed. [153] After these seed studies, variations of the idea that bioelectricity could sense injury and trigger or at least be a major player in regeneration have spurred over the decades until the present day. A potential explanation lies on resting potentials (primarily Vmem and TEP), which can be, at least in part, dormant sensors (alarms) ready to detect and effectors (triggers) ready to react to local damage. [126] [154] [155] [12]

Following up on the relative success of electric stimulation on non-permissive frog leg regeneration using an implanted bimetallic rod in the late 1960s, [156] the bioelectric extracellular aspect of amphibian limb regeneration was extensively dissected in the next decades. Definitive descriptive and functional physiological data was made possible owing to the development of the ultra-sensitive vibrating probe and improved application devices. [40] [157] Amputation invariably leads to a skin-driven outward current and a consequent lateral electric field setting the cathode at the wound site. Although initially pure ion leakage, an active component eventually takes place and blocking ion translocators typically impairs regeneration. Using biomimetic exogenous electric currents and fields, partial regeneration was achieved, which typically included tissue growth and increased neuronal tissue. Conversely, precluding or reverting endogenous electric current and fields impairs regeneration. [59] [158] [157] [159] These studies in amphibian limb regeneration and related studies in lampreys and mammals [160] combined with those of bone fracture healing [161] [162] and in vitro studies, [131] led to the general rule that migrating (such as keratinocytes, leucocytes and endothelial cells) and outgrowing (such as axons) cells contributing to regeneration undergo electrotaxis towards the cathode (injury original site). Congruently, an anode is associated with tissue resorption or degeneration, as occurs in impaired regeneration and osteoclastic resorption in bone. [161] [159] [163] Despite these efforts, the promise for a significant epimorphic regeneration in mammals remains a major frontier for future efforts, which includes the use of wearable bioreactors to provide an environment within which pro-regenerative bioelectric states can be driven [164] [165] and continued efforts at electrical stimulation. [166]

Recent molecular work has identified proton and sodium flux as being important for tail regeneration in Xenopus tadpoles, [12] [167] [168] and shown that regeneration of the entire tail (with spinal cord, muscle, etc.) could be triggered in a range of normally non-regenerative conditions by either molecular-genetic, [169] pharmacological, [170] or optogenetc [171] methods. In planaria, work on bioelectric mechanism has revealed control of stem cell behavior, [172] size control during remodeling, [173] anterior-posterior polarity, [174] and head shape. [68] [175] Gap junction-mediated alteration of physiological signaling produces 2-headed worms in Dugesia japonica remarkably, these animals continue to regenerate as 2-headed in future rounds of regeneration months after the gap junction-blocking reagent has left the tissue. [176] [177] [178] This stable, long-term alteration of the anatomical layout to which animals regenerate, without genomic editing, is an example of epigenetic inheritance of body pattern, and is also the only available “strain” of planarian species exhibiting an inherited anatomical change that is different from the wild-type. [179]

Defection of cells from the normally tight coordination of activity towards an anatomical structure results in cancer it is thus no surprise that bioelectricity – a key mechanism for coordinating cell growth and patterning – is a target often implicated in cancer and metastasis. [180] [181] Indeed, it has long been known that gap junctions have a key role in carcinogenesis and progression. [182] [183] [184] Channels can behave as oncogenes and are thus suitable as novel drug targets. [3] [92] [182] [185] [186] [187] [188] [189] [190] [191] Recent work in amphibian models has shown that depolarization of resting potential can trigger metastatic behavior in normal cells, [192] [193] while hyperpolarization (induced by ion channel misexpression, drugs, or light) can suppress tumorigenesis induced by expression of human oncogenes. [194] Depolarization of resting potential appears to be a bioelectric signature by which incipient tumor sites can be detected non-invasively. [195] Refinement of the bioelectric signature of cancer in biomedical contexts, as a diagnostic modality, is one of the possible applications of this field. [180] Excitingly, the ambivalence of polarity – depolarization as marker and hyperpolarization as treatment – make it conceptually possible to derive theragnostic (portmanteau of therapeutics with diagnostics) approaches, designed to simultaneously detect and treat early tumors, in this case based on the normalization of the membrane polarization. [194]

Recent experiments using ion channel opener/blocker drugs, as well as dominant ion channel misexpression, in a range of model species, has shown that bioelectricity, specifically, voltage gradients instruct not only stem cell behavior [196] [197] [198] [199] [200] [201] but also large-scale patterning. [29] [202] [203] Patterning cues are often mediated by spatial gradients of cell resting potentials, or Vmem, which can be transduced into second messenger cascades and transcriptional changes by a handful of known mechanisms (Figure 7). These potentials are set by the function of ion channels and pumps, and shaped by gap junctional connections which establish developmental compartments (isopotential cell fields). [204] Because both gap junctions and ion channels are themselves voltage-sensitive, cell groups implement electric circuits with rich feedback capabilities (Figure 8). The outputs of developmental bioelectric dynamics in vivo represent large-scale patterning decisions such as the number of heads in planaria, [178] the shape of the face in frog development, [98] and the size of tails in zebrafish. [102] Experimental modulation of endogenous bioelectric prepatterns have enabled converting body regions (such as the gut) to a complete eye [88] (Figure 9), inducing regeneration of appendages such as tadpole tails at non-regenerative contexts, [171] [170] [169] and conversion of flatworm head shapes and contents to patterns appropriate to other species of flatworms, despite a normal genome. [175] Recent work has shown the use of physiological modeling environments for identifying predictive interventions to target bioelectric states for repair of embryonic brain defects under a range of genetic and pharmacologically-induced teratologies. [89] [100]

Life is ultimately an electrochemical enterprise research in this field is progressing along several frontiers. First is the reductive program of understanding how bioelectric signals are produced, how voltage changes in the cell membrane are able to regulate cell behavior, and what are the genetic and epigenetic downstream targets of bioelectric signals. A few mechanisms that transduce bioelectric change into alterations of gene expression are already known, including the bioelectric control of movement of small second-messenger molecules through cells, including serotonin and butyrate, voltage sensitive phosphatases, among others. [205] [206] Also known are numerous gene targets of voltage signaling, such as Notch, BMP, FGF, and HIF-1α. [127] Thus, the proximal mechanisms of bioelectric signaling within single cells are becoming well-understood, and advances in optogenetics [79] [171] [4] [207] [208] and magnetogenetics [209] continue to facilitate this research program. More challenging however is the integrative program of understanding how specific patterns of bioelectric dynamics help control the algorithms that accomplish large-scale pattern regulation (regeneration and development of complex anatomy). The incorporation of bioelectrics with chemical signaling in the emerging field of probing cell sensory perception and decision-making [210] [211] [212] [213] [214] [215] is an important frontier for future work.

Bioelectric modulation has shown control over complex morphogenesis and remodeling, not merely setting individual cell identity. Moreover, a number of the key results in this field have shown that bioelectric circuits are non-local – regions of the body make decisions based on bioelectric events at a considerable distance. [100] [103] [104] Such non-cell-autonomous events suggest distributed network models of bioelectric control [216] [217] [218] new computational and conceptual paradigms may need to be developed to understand spatial information processing in bioelectrically-active tissues. It has been suggested that results from the fields of primitive cognition and unconventional computation are relevant [217] [219] [68] to the program of cracking the bioelectric code. Finally, efforts in biomedicine and bioengineering are developing applications such as wearable bioreactors for delivering voltage-modifying reagents to wound sites, [165] [164] and ion channel-modifying drugs (a kind of electroceutical) for repair of birth defects [89] and regenerative repair. [170] Synthetic biologists are likewise starting to incorporate bioelectric circuits into hybrid constructs. [220]

Virus-Cell Interactions

Viruses adapt to their hosts in large part by evolving to interact efficiently with host cells in initiating infection and producing large amounts of virus. The vires spreads to different organs of the host and in this process causes tissue damage. Strains of a virus (e.g., variola major and variola minor) differ in their vimlence or ability to cause fatal disease. The differences in virulence may be due to changes in the rapidity of virus replication and spread, the amounts of vires produced, the ability to damage the cells in which the vires replicates, or the ability to evade the immune response of the host. In addition, orthopoxvirus tissue tropism genes have been identified in vaccinia vires and cowpox (C7L, K1L, and CHOhr), and the morphogenesis of the multiple forms of orthopoxvirus particles is coming better understood [43, 44]. The genetic basis of orthopoxvirus infections may thereby be revealed. Infection of human cells grown in tissue culture could begin to provide answers to some of the following questions:

Finally, judging from what is known about other poxviruses, modulation of host immune responses is highly likely to contribute to the virulence of the virus. Infection of immune system cells could make it possible to assess direct effects on such cells, and incubation of human immune system cells with proteins secreted by infected cells could allow identification of potentially unique interactions between viral proteins and mediators of the antiviral immune response. These interactions could be used to identify important and potentially unique aspects of the human response to virus infections.

Lysosomal storage disorders: The cellular impact of lysosomal dysfunction

Lysosomal storage diseases (LSDs) are a family of disorders that result from inherited gene mutations that perturb lysosomal homeostasis. LSDs mainly stem from deficiencies in lysosomal enzymes, but also in some non-enzymatic lysosomal proteins, which lead to abnormal storage of macromolecular substrates. Valuable insights into lysosome functions have emerged from research into these diseases. In addition to primary lysosomal dysfunction, cellular pathways associated with other membrane-bound organelles are perturbed in these disorders. Through selective examples, we illustrate why the term �llular storage disorders” may be a more appropriate description of these diseases and discuss therapies that can alleviate storage and restore normal cellular function.

Lysosomal storage disorders: A brief overview

Inborn errors of metabolism are a common cause of inherited disease (Burton, 1998), of which lysosomal storage diseases (LSDs) are a significant subgroup (Platt and Walkley, 2004 Fuller et al., 2006 Ballabio and Gieselmann, 2009). The combined incidence of LSDs is estimated to be approximately 1:5,000 live births (Fuller et al., 2006), but the true figure is likely greater when undiagnosed or misdiagnosed cases are accounted for. Common to all LSDs is the initial accumulation of specific macromolecules or monomeric compounds inside organelles of the endosomal𠄺utophagic–lysosomal system. Initial biochemical characterization of stored macromolecules in these disorders led to the implication of defective lysosomal enzymes as a common cause of pathogenesis (Hers, 1963 Winchester, 2004). Although most LSDs result from acidic hydrolase deficiencies (Winchester, 2004), a considerable number of these conditions result from defects in lysosomal membrane proteins or non-enzymatic soluble lysosomal proteins (Saftig and Klumperman, 2009). Therefore, LSDs offer a window into the normal functions of both enzymatic and non-enzymatic lysosomal proteins.

Clinical phenotypes of LSDs

The age of clinical onset and spectrum of symptoms exhibited amongst different LSDs vary, depending on the degree of protein function affected by specific mutations, the biochemistry of the stored material, and the cell types where storage occurs. Apart from lysosomal diseases involving substrate storage in bone and cartilage (e.g., the mucopolysaccharidoses Table 1 ) most babies born with these conditions appear normal at birth. The classical clinical presentation of an LSD is a neurodegenerative disease of infancy/childhood (Wraith, 2002), but adult-onset variants also occur (Spada et al., 2006 Nixon et al., 2008 Shapiro et al., 2008). A health surveillance program tasked with diagnosing all neurodegenerative disease cases in UK children has so far revealed that lysosomal disorders are amongst the most commonly confirmed diagnoses of neurodegeneration (45% of cases) and will provide a robust frequency of infantile/juvenile onset cases as the study gathers more data over the coming years (Verity et al., 2010). Key molecular and clinical features of the storage diseases mentioned in this review are summarized in Table 1 . In addition, detailed medical descriptions on the various disorders are available on the Online Metabolic and Molecular Bases of Inherited Disease (OMMBID) website (Valle et al., 2012).

Table 1.

The causes of lysosomal storage diseases, the organelles affected, and major sites of pathology

Mechanism of lysosomal storageDisease examplesLysosomal protein defect (gene symbol)Substrate(s) storedMajor peripheral organ systems affectedCNS pathology
Lysosomal enzyme deficienciesAspartylglucosaminuriaAspartylglucosaminidase (glycosylasparaginase, AGA)aspartylglucosamine (N-acetylglucosaminyl-asparagine)Skeleton, connective tissue+
Fabryα-Galactosidase (GLA)(Lyso-)GlobotriaosylceramideKidney, heart
Gaucher types 1, 2, and 3β-Glucocerebrosidase (GBA)Glucosylceramide, glucosylsphingosineSpleen/liver, bone marrow+ a
GM1-gangliosidosisβ-Galactosidase (GLB1)GM1-ganglioside, oligosaccharidesSkeleton, heart+
Krabbe (globoid cell leukodystrophy)Galactocerebrosidase (GALC)GalactosylceramideHeart+
Metachromatic leukodystrophyArylsulfatase A (ARSA)Sulfogalactosylceramide +
MucopolysaccharidosesEnzymes involve in mucopolysaccharide catabolismMucopolysaccharidesCartilage, bone, heart, lungs+ b
Multiple sulfatase deficiencySUMF1 (Formylglycine-generating enzyme needed to activate sulfatases)Multiple, including sulfated glycosaminoglycansSpleen/liver, bone, skin+
Pompeα-Glucosidase (GAA)GlycogenSkeletal muscle
Sandhoffβ-hexosaminidase A and B (HEXB)GM2-ganglioside +
Trafficking defect of lysososomal enzymesMucolipidosis type II (I-cell disease)N-acetyl glucosamine phosphoryl transferase α/β (GNPTAB)Carbohydrates, lipids, proteinsSkeleton, heart+
Mucolipidosis type IIIA (pseudo-Hurler polydystrophy)N-acetyl glucosamine phosphoryl transferase α/β (GNPTAB)Carbohydrates, lipids, proteinsSkeleton, heart+/−
Defects in soluble non-enzymatic lysosomal proteinsNiemann-Pick disease type C2NPC2 (soluble cholesterol binding protein)Cholesterol and sphingolipidsLiver+
Defects in lysosomal membrane proteinsCystinosisCystinosin (cysteine transporter, CTNS)CystineKidney, eye
Danon diseaseLysosomal-associated membrane protein 2, splicing variant A (LAMP2)Glycogen and other autophagic componentsCardiac and skeletal muscle+
Free sialic acid storage disorderSialin (sialic acid transporter, SLC17A5)Free sialic acidLiver/spleen, skeleton+
Mucolipidosis IVMucolipin-I (MCOLN1)Mucopolysaccharides and lipidsEye+
Niemann-Pick disease type C1NPC1 (membrane protein involved in lipid transport)Cholesterol and sphingolipidsLiver+
Enigmatic lysosomal disordersNeuronal ceroid lipofuscinoses (NCLs, including Batten disease)Disparate group of diseases with genetic defects in apparently unrelated genes, not all of which are associated with the lysosomal system. Not known if these genes cooperate in common cellular pathways.Autofluorescent lipofuscin is a common feature, with convergent clinical signs, e.g., visual system defects/blindness +

Listed are the diseases discussed in the main text. Mucopolysaccharidoses and neuronal ceroid lipofuscinoses refer to collections of related disorders.

Relatively few lysosomal diseases lack pathology in the central nervous system (CNS Wraith, 2004). In the majority of LSDs, CNS involvement is common and neurodegeneration can occur in multiple brain regions (e.g., thalamus, cortex, hippocampus, and cerebellum). Neuropathology in LSDs involves unique temporal and spatial changes, which often entails early region-specific neurodegeneration and inflammation, before global brain regions are affected. The main reasons for this are threefold: (1) specific storage metabolites exert differential effects on neuronal subtypes, (2) varying proportions of macromolecules are synthesized in different neuronal populations, and (3) there is differential neuronal vulnerability to storage (e.g., Purkinje neurons degenerate in many of these diseases leading to cerebellar ataxia). Activation of the innate immune system is also prevalent in the brain of LSDs, which directly contributes to CNS pathology (Vitner et al., 2010). Astrogliosis (activation of astrocytes) is another common feature of LSDs, which damages neurons through an inflammatory process known as glial scarring (Jesionek-Kupnicka et al., 1997 Vitner et al., 2010). The additive detrimental effects that astrogliosis has on neuron function is recapitulated in animal models of lysosomal diseases (Farfel-Becker et al., 2011 Pressey et al., 2012).

A notable non-neuronopathic LSD is Type 1 Gaucher disease (β-glucocerebrosidase deficiency), which is a relatively common LSD, particularly within the Ashkenazi Jewish community. The major cell type affected by glucosylceramide storage in this disease is the macrophage (“Gaucher cells”), whose dysfunction affects the production and turnover of cells belonging to the hematopoietic system. Gaucher cells infiltrate into various organs and affect the immune system, bone strength, spleen, and liver function.

A key question currently challenging this field is how endosomal–lysosomal storage leads to pathogenesis and how expanding this knowledge will improve treatment for patients (Bellettato and Scarpa, 2010 Cox and Cachón-González, 2012). This review aims to delineate regulatory systems and organelles that become disrupted in these disorders, highlighting the complexity of cellular storage, its consequences on pathogenesis, and implications for therapy.

Endosomal𠄺utophagic–lysosomal function and dysfunction in storage diseases

Lysosomes play a central role in processing the clearance of cellular substrates from multiple routes within the endosomal𠄺utophagic–lysosomal system ( Fig. 1 ). Lysosomes are acidic organelles that contain enzymes required for the degradation of macromolecules, and efflux permeases that facilitate the inside-out translocation of small molecules generated through macromolecule catabolism. In comparison to endosomes and autophagosomes, lysosomes are smaller in size, are highly enriched in particular transmembrane proteins and hydrolytic enzymes (including proteases, glycosidases, nucleases, phosphatases, and lipases), have a higher buoyant density, an electron-dense appearance by transmission electron microscopy, and a high proton and Ca 2+ content (Luzio et al., 2007 Saftig and Klumperman, 2009 Morgan et al., 2011). Lysosomes differ from endosomes in their degree of acidification and more abundant levels of lysosomal membrane proteins (LMPs) such as LAMP1 and LAMP2. Most nascent lysosomal enzymes bind to mannose-6-phosphate receptors (M6PRs) in the trans-Golgi network (TGN), which traffic the enzymes to early and late endosomes (Ghosh et al., 2003). Lysosomes in turn receive these enzymes when endosomal–lysosomal fusion occurs. Notably, dense lysosomes do not contain M6PRs. Acidotropic reagents such as Lysotracker are useful for labeling lysosomes however, the mildly acidic interiors of late endosomes and autophagosomes also allows Lysotracker to label these organelles to varying degrees (Bampton et al., 2005).

Lysosomes as catabolic centers of the cell. Lysosomes utilize four distinct pathways for the degradation of cellular material. (A) Macroautophagy begins with the formation of isolation membranes that sequester regions of the cytosol that include denatured proteins, lipids, carbohydrates, and old/damaged organelles into encapsulated vesicles known as autophagosomes. The dynamic kinetics of autophagosome production and clearance by lysosomes is known as autophagic flux. (B) Endosomal degradation by lysosomes predominantly targets late endosomes/multivesicular bodies. Fusion between late endosomes and lysosomes can occur by (i) full fusion/degradation or (ii) kiss-and-run content mixing, where transient endosomal docking occurs. (C) Microautophagy involves the pinocytosis of cytosolic regions surrounding lysosomes. (D) Chaperone-mediated autophagy (CMA) selectively targets proteins with a KFERQ motif for delivery to lysosomes using Hsc-70 as its chaperone and LAMP-2A as its receptor.

The biogenesis and functioning of endosomal and autophagosomal pathways is controlled by transcription factor EB (TFEB), which regulates the expression of 471 genes that constitute the CLEAR (coordinated lysosomal expression and regulation) gene network (Sardiello et al., 2009 Palmieri et al., 2011). Recent work indicates that non-active TFEB is highly phosphorylated and associates with late endosomes/lysosomes (Roczniak-Ferguson et al., 2011). Autophagy-inducing conditions (e.g., deprivation of glucose or amino acids) result in reduced and altered TFEB phosphorylation, leading to its translocation into the nucleus (Pe༚-Llopis et al., 2011) and transcriptional expression of CLEAR genes (Palmieri et al., 2011).

Degradation of endosomal and autophagosomal material takes place upon exchange of content (via transient “kiss-and-run” contacts) or fusion with lysosomes, forming endolysosomes (Tjelle et al., 1996 Bright et al., 1997, 2005 Mullock et al., 1998) and autolysosomes (Jahreiss et al., 2008 Fader and Colombo, 2009 Orsi et al., 2010), respectively ( Fig. 1, A and B ). Lysosomes can be regarded as storage compartments for acidic hydrolases that enter cycles of fusion and fission with late endosomes and autophagosomes, while the digestion of endocytosed and autophagic substrates takes place primarily in endolysosomes and autolysosomes (Tjelle et al., 1996 Luzio et al., 2007). Under physiological conditions, endolysosomes and autolysosomes are transient organelles.

Cells deficient in lysosomal hydrolytic enzymes, lysosomal membrane proteins, or non-enzymatic soluble lysosomal proteins accumulate excessive levels of undegraded macromolecules (enzyme deficiency) or monomeric catabolic products (efflux permease deficiency) and contain numerous endo/autolysosomes ( Fig. 2 ). When very high levels of macromolecules/monomers accumulate in endo/autolysosomes, they inhibit catabolic enzymes and permeases that are not genetically deficient, which results in secondary substrate accumulation (Walkley and Vanier, 2009 Lamanna et al., 2011 Prinetti et al., 2011). For example, lysosomal proteolytic capacity is reduced in fibroblasts from various LSDs, such as mucopolysaccharidoses I and VI, and GM1-gangliosidosis, which are themselves not caused by protease deficiency (Kopitz et al., 1993). The accumulation of primary and secondary substrates sets off a cascade of events that impacts not only the endosomal𠄺utophagic–lysosomal system, but also other organelles, including mitochondria, the ER, Golgi, peroxisomes ( Fig. 3 ), and overall cell function ( Fig. 4 ).

Subtypes of storage organelles accumulate in LSDs. In different LSDs, cells display a unique spectrum of dysfunctional organelles depending on the specific lysosomal enzyme or non-enzymatic protein affected. (A) In primary LSDs, deficiencies in degradative enzymes prevent the clearance of autophagic and endocytic substrates, resulting in the accumulation of (i) autolysosomes (LC3-II (+), LAMP-1 (+)), (ii) endolysosomes (CI-MPR (+), LAMP-1 (+)), and (iii), in the case of certain lipase deficiencies, lipid-rich multilamellar bodies (CI-MPR (+), LAMP-1 (+)). (B) In a secondary storage disease such as Niemann-Pick type C1, lysosomal enzyme function remains intact, but impaired heterotypic fusion of autophagic and endocytic organelles with lysosomes results in the accumulation of (iv) autophagosomes (LC3-II (+), LAMP-1 (−)), (v) late endosomes (CI-MPR (+), active cathepsin D (−)), and (vi) endosome-derived multilamellar bodies (lipid-rich, CI-MPR (+), active cathepsin D (−)). Note: many primary storage diseases also accumulate organelles seen in secondary storage diseases (see text).

Summary of organelles affected in LSDs. Also shown are selective examples of LSDs. See Table 1 and main text for details.

Hypothetical cascade of events in LSD pathology. How gene mutations in lysosomal enzymes and non-enzymatic lysosomal proteins could lead to LSDs. Endo/autolysosomal events are confined to the darker shaded background, whereas processes taking place in the cytoplasm that affect autophagosomes, the ER, Golgi, peroxisomes, and mitochondria are on the lighter background. Processes depicted have been observed in a number of LSDs but do not necessarily apply to all LSDs.

Autophagic pathways.

The autophagic (“self-eating”) pathway constitutively targets intracellular cytosolic components for lysosomal degradation, and is essential for maintaining cellular energy and metabolic homeostasis (Kuma and Mizushima, 2010 Singh and Cuervo, 2011). To date, three distinct forms of autophagy have been characterized: macroautophagy, microautophagy, and chaperone-mediated autophagy ( Fig. 1, A, C, and D ). All three autophagic processes culminate in lysosomal degradation however, routes taken by substrates to the lysosome differ between each form. Macroautophagy involves the bulk sequestration of cytosolic regions into double- or multi-membrane bound autophagosomes, which are trafficked to lysosomes for content digestion ( Fig. 1 A ). A diverse range of cellular material is degraded via macroautophagy, including lipids, carbohydrates and polyubiquitinated proteins, RNA, mitochondria, and fragments of the ER (Eskelinen and Saftig, 2009). The most characterized protein associated with autophagosomes is the lipidated (phosphatidylethanolamine) form of microtubule-associated protein light chain 3 (MAP-LC3), known as LC3-II, which is generated early in the autophagic process but degraded in the final phase of autophagic digestion.

Autophagic flux (the rate at which autophagic vacuoles are processed by lysosomes) is reduced in most LSDs (Ballabio, 2009 Ballabio and Gieselmann, 2009 Raben et al., 2009). This is evident from the combined elevation of autophagic substrates and autophagosome-associated LC3-II. LSD cells often display increased numbers of LC3(+) organelles, of which only a subgroup carry lysosomal markers, suggesting that both autophagosomes and autolysosomes persist in these conditions. For example, in mouse models of Batten disease (a neuronal ceroid lipofuscinosis [NCL] disorder Table 1 ), most LC3-positive compartments are not positive for LAMP1 (Koike et al., 2005), and in multiple sulfatase deficiency and juvenile neuronal ceroid lipofuscinosis, LC3 and LAMP1 are predominantly localized in separate organelles, which is even more pronounced after starvation (Cao et al., 2006 Settembre et al., 2008). Endosome–lysosome and autophagosome–lysosome fusion is also impaired in mucolipidosis type IIIA and multiple sulfatase-deficient mouse embryonic fibroblasts (Fraldi et al., 2010).

Microautophagy does not involve de novo synthesis of nascent vacuoles, but rather occurs via the direct pinocytosis of cytosolic material by lysosomes ( Fig. 1 C ). The membrane dynamics regulating microautophagy are similar to those involved in the formation of intra-luminal vesicles (ILVs) found in multivesicular bodies/late endosomes (Sahu et al., 2011). Currently, little is known about the repercussions of lysosomal storage on microautophagy, but this process appears to be impaired in primary myoblasts from patients with the muscle-wasting condition Pompe disease (Takikita et al., 2009).

Chaperone-mediated autophagy (CMA) is a selective form of autophagic proteolysis that targets proteins containing a KFERQ motif for degradation (Dice et al., 1990 Cuervo and Dice, 2000). The eponymous chaperone that recognizes and binds to proteins destined for CMA is the heat shock cognate protein of 70 kD (Hsc70). Substrate-bound Hsc70 docks on lysosomes via contact with lysosomal-associated membrane protein 2A (LAMP-2A), allowing entry of proteins into lysosomes ( Fig. 1 D ). Mutations in LAMP-2A cause Danon disease, and specifically affect CMA (Eskelinen et al., 2003 Fidziańska et al., 2007). CMA is also known to be impaired in mucolipidosis IV, where mutations in transient receptor potential mucolipin-1 (MCOLN1) lead to reduced amounts of LAMP-2A and substrate uptake into lysosomes (Venugopal et al., 2009).

Lysosome reformation.

Both endolysosomes and autolysosomes extend tubular structures where lysosomal hydrolases and LMPs concentrate (Tjelle et al., 1996 Bright et al., 1997, 2005 Pryor et al., 2000 Yu et al., 2010). At the ends of these tubules, [LC3(−), LAMP1(+)] vesicles bud off and acidify, maturing into dense lysosomes, a fission process referred to as lysosome reformation. This event completes each cycle of endocytic and autophagic degradation, yielding dense lysosomes that are available to fuse with newly generated endosomes and autophagosomes.

Efficient processing of endo/autolysosomal substrates is essential for lysosome reformation. This is well illustrated in a study that monitored exogenous sucrose metabolism in rat kidney fibroblasts (Bright et al., 1997). Sucrose is a disaccharide composed of the monosaccharides glucose and fructose, and is itself indigestible by cells. In this study, sucrose-filled endosomes fused with lysosomes and formed large endolysosomes, which accumulated in the cytosol. A depletion of dense-core lysosomes was seen under these conditions however, dissolution of the accumulated sucrose by uptake of exogenous invertase resulted in the reappearance of dense-core lysosomes. This study and another more recent one from Yu et al. (2010) indicate that lysosome biogenesis does not occur de novo, but is rather born out of a reformation/budding from endolysosomes. Lysosome reformation appears to be defective in sialic acid storage disease as skin fibroblasts from diseased individuals lack dense lysosomes, while lysosomal enzymes persist in intermediate or light organelles (Schmid et al., 1999).

Interestingly, impairment of lysosome reformation appears to be the primary cellular defect in Niemann-Pick type C2 (NPC2)-deficient cells, indicating that the NPC2 protein has a crucial role in this process (Goldman and Krise, 2010). Considering that NPC1 and NPC2 deficiencies have the same pathological consequences (Niemann-Pick type C disease Table 1 ), this suggests that lysosome reformation is as essential as endosome/autophagosome–lysosome fusion, which is impaired in NPC1-deficient cells.

Recent reports have provided a mechanistic link between the failure of endo/autolysosomal clearance and the deficit of lysosome reformation. Central to this pathway is mTOR, a serine/threonine kinase that has an overarching role in coordinating cellular metabolism with nutritional status (Laplante and Sabatini, 2012). During the course of the autophagic process, mTOR goes through a cycle of phosphorylation-dependent inactivation and reactivation, with the latter being required for autophagic lysosome reformation (Yu et al., 2010). In turn, mTOR reactivation depends on the completion of autolysosomal substrate digestion, and sufficient levels of luminal amino acids (Zoncu et al., 2011). Limited information is currently available on the extent of lysosome reformation and mTOR reactivation in LSDs. However, inadequate autolysosomal degradation may preclude mTOR reactivation and, hence, also impede lysosome reformation, leaving affected cells deprived of dense lysosomes. Consequently, in addition to stalled autolysosomes, autophagosomes may persist due to a deficiency of dense lysosomes, explaining the low level of colocalization of autophagosomal and lysosomal markers. mTOR activity is reduced in the brain of a mouse model of juvenile neuronal ceroid lipofuscinosis (Cao et al., 2006), in fibroblasts from mucopolysaccharidosis type I S, Fabry disease and aspartylglucosaminuria subjected to starvation-induced autophagy (Yu et al., 2010), in NPC1- and NPC2-knockdown human umbilical vein endothelial cells (Xu et al., 2010), and in MCOLN1-deficient Drosophila pupae (Wong et al., 2012), but not in brain samples from Sandhoff, GM1-gangliosidosis, and NPC1 mice (Boland et al., 2010). Considering the myriad of cellular signaling pathways that mTOR is involved in (Laplante and Sabatini, 2012), it may be necessary to differentiate mTOR activity in affected cell populations of different brain regions. In addition, electron microscopy remains a powerful tool for the ultrastructural classification of autophagosomes and autolysosomes in LSD cells, and could also be used to monitor the extent of lysosome reformation.

Mitochondrial dysfunction and cytoplasmic protein aggregation.

In LSDs, a reduction of autophagic flux has a major impact on mitochondrial function and on cytoplasmic proteostasis. Constitutive macroautophagy maintains mitochondrial quality by selectively degrading dysfunctional mitochondria via a process known as mitophagy (Kim et al., 2007). Mitochondrial proteins are consistently found in the proteomes of highly purified autolysosomes, especially subunits of the mitochondrial ATPase (Schrr et al., 2010). Reduced autophagic flux in LSDs leads to the persistence of dysfunctional mitochondria, which is highly pronounced in Batten’s disease neurons (Ezaki et al., 1996). Several LSDs (mucolipidosis types IV, IIIA [pseudo-Hurler polydystrophy], and II [I-cell disease], late infantile neuronal ceroid lipifuscinosis [CLN2], mucopolysaccharidosis VI, and GM1 gangliosidosis) display mitochondrial abnormalities, including replacement of the extended filamentous mitochondrial network with high numbers of relatively short mitochondria, and loss of mitochondrial calcium-buffering capacity and membrane potential (Jennings et al., 2006 Settembre et al., 2008 Takamura et al., 2008 Tessitore et al., 2009). Studies into aging and autophagosome formation have shown that mitochondria are involved in signaling pathways regulating apoptosis and innate immunity, and that reduced autophagic flux and subsequent accumulation of dysfunctional, reactive oxygen species–generating mitochondria renders cells more sensitive to apoptotic and inflammatory stimuli (Terman et al., 2010 Green et al., 2011 Nakahira et al., 2011 Zhou et al., 2011). Therefore, the aberrant functioning of mitochondria may be responsible for apoptosis and inflammation in the CNS of multiple LSDs.

In addition, a lack of autophagy completion in LSDs leads to the persistence of ubiquitinated and aggregate-prone polypeptides in the cytoplasm, including p62/SQSTM1, α-synuclein, and Huntingtin protein (Ravikumar et al., 2002 Suzuki et al., 2007 Settembre et al., 2008 Tessitore et al., 2009). Alpha-synuclein itself contributes to neurodegeneration by reducing the efficiency of autophagosome formation (Winslow et al., 2010), and is also a main component of Lewy bodies that are notably elevated in Parkinson’s disease and other forms of dementia. Diminished quality control of cytosolic proteins may thus also contribute to LSD pathology.

Impairment of autophagy and escalation of cytoplasmic protein aggregation are shared between neurodegenerative LSDs and more common neurodegenerative disorders, such as Alzheimer’s, Parkinson’s, Huntington’s disease, and amyotrophic lateral sclerosis (ALS Garc໚-Arencibia et al., 2010 Wong and Cuervo, 2010). Mutations in presenilin-1, which cause a familial form of Alzheimer’s disease, is known to impair lysosomal clearance of autophagosomes (Esselens et al., 2004 Wilson et al., 2004 J.H. Lee et al., 2010). Different mechanisms have been proposed to explain how the partial loss of presenilin function impairs autophagic flux. Reports from J.H. Lee et al. (2010) indicate that presenilin 1 is need for the glycosylation and subsequent delivery of V0a1 protein to lysosomes, where it forms a subunit of lysosomal v-ATPase. This in turn is thought to impair lysosomal proteolysis by raising their pH above an optimal acidity of pH4𠄵. Alternatively, another recent report has indicated that mutations in presenilin 1 lead to a loss of lysosomal calcium regulation, which in turn affects fusion and clearance of autophagosomes (Coen et al., 2012). However, considering both groups confirmed that presenilin 1 mutations affect autophagic flux, Alzheimer’s disease is beginning to emerge as a neurodegenerative disorder that may share similarities in terms of underlying pathogenic mechanisms with lysosomal storage disorders.

Efflux of molecules from endo/autolysosomes.

Some storage molecules in LSDs (glycoconjugates, amino acids, or insoluble lipids) escape from cells and can be detected in blood and/or urine, which can be utilized for diagnostic purposes (Meikle et al., 2004). While glycoconjugates derived from storage cells in multiple tissues could escape as solutes in blood and urine, lipids extracted from urine are believed to be membrane associated and predominantly exosomal (Pisitkun et al., 2004).

At the cellular level, a big question that remains to be resolved concerns the way in which storage molecules escape the lysosomal system and affect the function of other organelles and cellular systems (Elleder, 2006). Theoretically, lipids can undergo redistribution within cells via membrane trafficking, fusion, or via altered trafficking pathways characteristic of these diseases (Chen et al., 1999). Endolysosomal macromolecules may also be disseminated via membrane contact sites between endolysosomes and the ER (Eden et al., 2010 Toulmay and Prinz, 2011), and by extracellular secretion of endolysosomal content, including exosome release. For example, primary kidney cells from arylsulfatase A�icient mice secrete the accumulating lipid (sulfogalactosylceramide) into the culture medium (Klein et al., 2005), and NPC1-deficient cells release higher amounts of cholesterol-rich exosomes (Chen et al., 2010 Strauss et al., 2010). Accordingly, the possibility needs to be considered that exosomes containing storage molecules are taken up by recipient cells, and that these macromolecules and lipids affect recipient cell function by distributing to the plasma membrane and other organelles outside the endolysosomal system (Simons and Raposo, 2009).

Due to the extraordinarily high levels of lipids in the endo/autolysosomal system, even a minor redistribution to other cellular membranes could have functional implications. Over the past few years, multiple examples have emerged suggesting that this not only occurs but can actively contribute to the pathogenic cascade (Vitner et al., 2010). A key challenge is to demonstrate experimentally that particular storage macromolecules are indeed ectopically present in the membrane of other organelles. This is technically challenging due to the limitations of conventional cell fractionation techniques. Currently, the presence of storage components in non-lysosomal sites is either inferred indirectly or evidence has been provided by immunostaining methods. To date, the best examples come from studying the effects of lipid storage in the ER (Sano et al., 2009 Futerman, 2010).

Lysosomal calcium homeostasis.

Endosomes and lysosomes are regulated calcium stores (Morgan et al., 2011) that release calcium in response to the second messenger nicotinic acid adenine dinucleotide phosphate (NAADP Churchill et al., 2002). NPC1 disease is unusual in having a profound block in late endosome–lysosome fusion (Kaufmann et al., 2009 Goldman and Krise, 2010), a process known to be calcium dependent (Lloyd-Evans et al., 2008). In NPC1 patient cells and cultured cells deficient in NPC1 protein, calcium levels within acidic organelles are approximately 30% of wild-type cells (Lloyd-Evans et al., 2008 H. Lee et al., 2010). NPC1 cells do respond to NAADP, but, due to the reduced luminal calcium levels, release less calcium, thus leading to the fusion deficiency associated with this disorder (Lloyd-Evans et al., 2008). Therefore, NPC1 disease demonstrates that acidic calcium stores play a central role in the regulation of fusion and trafficking within the endocytic system itself (Morgan et al., 2011).

Endoplasmic reticulum defects.

In addition to the endoplasmic reticulum (ER) being the major site of the secretory pathway responsible for protein folding/quality control and N-glycosylation, it is also a regulated calcium store. The lipid and protein content of the ER is tightly regulated to maintain its essential quality-control functions. Surprisingly, very few examples of ER stress (e.g., unfolded protein response) have been reported among LSDs, with GM1 gangliosidosis being the only sphingolipid storage disorder in which this has been demonstrated to date (Tessitore et al., 2004 Sano et al., 2009 Vitner et al., 2010). Instead, the major impact in lipid storage disorders is on ER calcium regulation (Futerman and van Meer, 2004 Futerman, 2010). ER calcium homeostasis is perturbed in the sphingolipid storage disorders, Gaucher disease, GM1 and GM2 gangliosidoses, and Niemann-Pick type A (Ginzburg and Futerman, 2005), leading to elevated cytosolic calcium. In these diseases, the characteristic lipids being stored, glucosylceramide, GM1 and GM2 ganglioside, and sphingomyelin, respectively, may hypothetically escape from endolysosomes and affect ER calcium channel function. Interestingly, the mechanisms leading to defective ER calcium homeostasis are specific to each disorder and have recently been reviewed (Vitner et al., 2010). In turn, aberrant ER calcium regulation may impact mitochondria through ER–mitochondria contact sites, resulting in mitochondrial calcium excess and an induction of mitochondria-mediated apoptosis, as seen in GM1 gangliosidosis (Sano et al., 2009).

The Golgi.

Dysfunction of the Golgi is a common feature of many lipid storage disorders, and has traditionally been thought to arise from alterations in sphingolipid trafficking from the Golgi to the lysosome (Pagano et al., 2000). However, recently Golgi involvement has been demonstrated in mucopolysaccharidosis IIIB (Sanfillipo B syndrome Vitry et al., 2010). Surprisingly, this study did not find any evidence that the endocytic and autophagic pathways were affected in Sanfillipo B syndrome instead, they noticed that large storage bodies were enriched in the Golgi matrix protein, GM130, which is required for vesicle tethering in pre- and cis-Golgi compartments. Furthermore, the morphology of the Golgi apparatus was altered in cells with distended cisternae connected to LAMP1-postive storage bodies. This study therefore suggests that Golgi biogenesis may be affected in this disease and further studies will shed light on the molecular mechanisms that underpin Golgi involvement in this neurodegenerative disorder.


There are reports of peroxisomal dysfunction occurring in some lipid lysosomal storage diseases, including Krabbe (globoid cell leukodystrophy Haq et al., 2006) and NPC1 disease (Schedin et al., 1997). In Krabbe disease, the major storage lipid galactosylceramide is converted into its lysosomal metabolite, galactosylsphingosine, which down-regulates the peroxisome proliferator�tivated receptor-α (PPAR-α). Loss of PPAR-α and subsequent cell death can be prevented using an inhibitor of secretory phospholipase A2, suggesting a novel therapeutic approach for Krabbe disease (Haq et al., 2006). In the NPC1 disease mouse model, peroxisomes appear normal at the ultrastructural level but have decreased peroxisomal β oxidation of fatty acids and catalase activity, which is an early event in disease pathogenesis (Schedin et al., 1997). In peroxisomal biogenesis disorders such as Zellweger syndrome and infantile Refsum disease, a-series gangliosides (e.g., GM1, GM2) and their precursor GM3 ganglioside are stored. As these gangliosides are common secondary storage metabolites in many LSDs, this raises the possibility that peroxisomal dysfunction underpins secondary ganglioside storage in LSDs and merits systematic study to test this hypothesis. How peroxisomal function affects ganglioside metabolism remains unknown but may be part of a broader lipid regulatory network in mammalian cells.

Cellular metabolic stress.

Considering that both endocytic and autophagic pathways are essential for maintaining cellular metabolic homeostasis, the diminished efflux of monomeric products from endo/autolysosomes is likely to induce a state of metabolic insufficiency, where key catabolic intermediates are unavailable to enter a variety of metabolic recycling pathways (Schwarzmann and Sandhoff, 1990 Walkley, 2007). For example, in some cell types, the majority of nascent glycosphingolipids are synthesized from endolysosome-derived sphingoid bases derived from ceramide catabolism (Tettamanti, 2004 Kitatani et al., 2008). Multiple endolysosomal exoglycosidases, including glucocerebrosidase, which is deficient in Gaucher disease, are involved in this process (Kitatani et al., 2009). The lack of reutilized sphingolipids/fatty acids that normally result from endolysosomal degradation would place such cells under significant metabolic stress. This may also apply to NPC disease, which is a particularly complex and enigmatic storage disease caused by mutations in either the NPC1 or NPC2 genes, with resulting storage of several lipids species including cholesterol and various sphingolipids (Lloyd-Evans and Platt, 2010). The NPC1 protein is an integral membrane protein of late endosomes that may function to efflux sphingosine (protonated at acidic pH) out of endolysosomes and into the sphingolipid salvage pathway or undergo phosphorylation to sphingosine-1-phosphate (S1P), raising the possibility that S1P deficiency contributes to NPC1 disease pathogenesis (Lloyd-Evans et al., 2008 Lloyd-Evans and Platt, 2010).

Therapeutic implications

Over the past two decades there has been a remarkable expansion in the number of therapeutic strategies for LSDs that target different cellular organelles ( Table 2 ). The first treatment that led to a licensed commercial product was enzyme replacement therapy (ERT) for type 1 Gaucher disease. The discoveries leading to that seminal therapeutic advance were recently reviewed by Roscoe Brady, who pioneered this approach (Brady, 2010). This therapy “replaces” the defective enzyme in the lysosome by delivering a fully functional wild-type enzyme that is endocytosed into macrophages via the macrophage mannose receptor. Wild-type glucocerebrosidase was initially purified from human placenta (now recombinant products are used) and typically given to patients every two weeks by intravenous infusion (Charrow, 2009). This strategy leads to a remarkable degree of therapeutic benefit and has transformed the lives of patients with this debilitating peripheral storage disease (Charrow, 2009). This success catalyzed the development of ERT for Fabry disease (Schiffmann and Brady, 2006 Angelini and Semplicini, 2012), Pompe disease (Angelini and Semplicini, 2012), and several of the mucopolysaccharide storage disorders (Kakkis, 2002). However, the clinical limitations of ERT are two-fold. First, product delivery is invasive and time-consuming to deliver, and second, lysosomal enzymes do not cross the blood𠄻rain barrier to any significant extent, so cannot effectively treat CNS disease, which is characteristic of most LSDs. To circumvent this problem, bone marrow (BM) transplantation from healthy donors has been evaluated in some of these diseases. Microglia are of BM origin and over time a few donor-derived monocytes enter the CNS and serve as local sites of wild-type enzyme production, which can be taken up via secretion-recapture by neighboring host cells. On the whole, BM transplantation is only effective if it is performed in early infancy, does not show efficacy in all LSDs, and is not curative (Wraith, 2001). Further complications include the need for human leukocyte antigen (HLA) matched donors, the high rate of mortality associated with recipients, and the lack of standardization amongst different BMT regimens in different clinical centers.

Table 2.

Status of approved treatments and experimental therapies for LSDs with selected bibliography

TherapyTarget organelleIn vitro POCIn vivo POCClinical trialsRegulatory approvalReferences
Enzyme replacement (ERT)Lysosome++++Brady, 2006b Neufeld, 2011
Bone marrow transplantation (BMT)Lysosome+++N/AKrivit, 2002 Brady, 2006a
Substrate reduction therapy (SRT)Golgi++++Platt and Butters, 2004 Platt and Jeyakumar, 2008 Cox, 2010
Enzyme enhancement therapy (EET)ER/lysosome+In progressOkumiya et al., 2007 Fan, 2008
Gene therapy (GT)Nucleus++In progressGritti, 2011 Tomanin et al., 2012
Stop codon read-throughNucleus+Brooks et al., 2006
Calcium modulation therapy (CMT)ER++Lloyd-Evans et al., 2008
Enhanced exocytosis therapy (ExT)Exosome+Strauss et al., 2010 Medina et al., 2011
Chaperone therapy by HSp70 (CT)Lysosome+Kirkegaard et al., 2010
Proteostasis regulation therapy (PRT)ER+Balch et al., 2008 Mu et al., 2008
Cholesterol removal using cyclodextrin in NPC1 diseaseLysosome++Davidson et al., 2009 Ward et al., 2010 Aqul et al., 2011

Another therapy to be developed and subsequently approved for LSDs was substrate reduction therapy using the oral small molecule imino sugar drug, miglustat (Lachmann, 2006). This has been approved for type 1 Gaucher disease (worldwide) for over a decade, and in 2009 for treating neurological manifestations in Niemann-Pick type C disease (now approved in most countries/regions, except the USA Patterson et al., 2007). Miglustat targets the Golgi enzyme, glucosylceramide synthase (Platt et al., 1994), and by partially inhibiting glycosphingolipid biosynthesis it reduces the catabolic burden of these molecules on lysosomes that cannot digest them. It has the potential to be used in diseases with glycosphingolipid storage, as miglustat inhibits the first committed step in the biosynthesis of this family of lipids. Also, miglustat crosses the blood𠄻rain barrier, hence its disease-modifying benefit in Niemann-Pick type C disease (Patterson et al., 2007). Like all drugs, this compound has side effects, the primary one being inhibition of disaccharidases, which can lead to gastrointestinal symptoms, particularly in the first 1𠄲 months of therapy. More recently, eliglustat tartrate (Genz-112638) has entered clinical trials in type 1 Gaucher disease as an oral substrate reduction therapy. As this drug has a different chemistry to miglustat, it also has a different side-effect profile (Cox, 2010).

There are currently several alternative therapeutic strategies that have shown utility in tissue culture models and/or in animal models of these diseases and are summarized in Table 2 . Many of these approaches target non-lysosomal organelles. No doubt as more is known about pathogenic cascades and their impact on cellular organelles, additional creative approaches to treatment will emerge and undergo pre-clinical testing. Due to the severity and complexity of these disorders it is likely that ultimately a combination therapy will be needed to target multiple steps/organelles in the pathogenic cascade.


In conclusion, we have provided some selective examples illustrating the complexity of how lysosomal dysfunction impinges upon multiple aspects of cell biology, often in unanticipated ways (summarized in Fig. 3 ). Many questions remain unanswered at the present time, and some of these are highlighted in Box 1. However, the study of these rare diseases ( Table 1 ) fills two voids in our knowledge, namely providing fundamental insights into lysosomal biology and in leading to novel approaches to generate next-generation therapeutic interventions for treating these truly fascinating yet devastating disorders ( Table 2 ). It is clear that although storage is primarily initiated in the late endosomal𠄺utophagic–lysosomal system, it induces a pathogenic cascade that impacts on multiple cellular systems and organelles, suggesting that conceptually we should view these diseases as cellular storage disorders and use this broader knowledge for the design of therapeutic interventions.

Box 1. Open Questions

• How does storage affect other aspects of lysosomal function, independent of the primary storage metabolite?

• How does storage trigger innate immune activation?

• How does lysosomal storage affect cell signaling?

• How do storage lipids escape the lysosome and affect the function of other organelles?

• What is the hierarchy of the pathogenic cascade in these diseases, which steps should be targeted for optimal therapy?

• Do the genetic defects in the neuronal ceroid lipofuscinoses (NCL disorders) cause convergent symptoms by chance, or are the disparate genes functioning in common cell biological pathways?

Formation of Reactive Oxygen Species and Cellular Damage

Reactive oxygen species (ROS) are molecules containing an oxygen atom with an unpaired electron in its outer shell. As ROS are formed, they become very unstable due to the unpaired electron now residing in the outermost shell. The unstable forms of oxygen are sometimes called free radicals.

How do ROS actually get generated in cells? One way is via cellular respiration driven by the electron transport chain in the mitochondria. The electron transport chain is responsible for generating ATP, the main source of energy for a cell to function. A key molecule that helps “jump start” the electron transport chain, is NADH (or nicotinamide adenine dinucleotide), which serves as the electron donor (i.e., the H in the NADH). NADH is often referred to as a “coenzyme”, even though it is not an enzyme (a protein).

NADH is present in all cells–it is generated by many biochemical reactions. One way that NADH gets generated in large quantities is when alcohol is metabolized (or oxidized) to form acetaldehyde and then to acetic acid. During the metabolism of alcohol, the enzyme alcohol dehydrogenase (ADH) and NAD + convert alcohol to acetylaldehyde, generating NADH. A second enzyme, aldehyde dehydrogenase (ALDH) and NAD + convert acetaldehyde to acetic acid, generating even more NADH. In these reactions, the coenzyme NAD + is reduced to NADH (and alcohol and acetaldehyde are oxidized).

Review the oxidation of alcohol by alcohol dehydrogenase (ADH)

To learn more about the oxidation of alcohol by ADH, you can participate in a virtual reality game called “DiVE into Alcohol” at

Now there is plenty of NADH available to “jump start” mitochondrial respiration. NADH moves from the cytosol into the mitochondria where it donates an electron to the electron transport chain. The electron transport chain consists of a group of proteins (and some lipids) that work together to pass electrons “down the line”. Finally in the presence of oxygen, ATP is formed, providing energy for many cellular functions.

However, some electrons can “escape” the electron transport chain and combine with oxygen to form a very unstable form of oxygen called a superoxide radical (O2•-). The superoxide radical is one of the reactive oxygen species (ROS).

The superoxide radical is a type of free radical. Free radicals have a lone electron in their outer electron orbital and they are very reactive molecules because they tend to donate single electrons (e-) or steal e- from other molecules. Free radicals can be destructive to cellular components. Free radicals often have a • shown to indicate the lone e-.

Our cells have ways to protect themselves from the damaging effects of these reactive molecules. For example, our cells are able to maintain low levels of the superoxide radicals with the help of the enzyme superoxide dismutase (SOD). SOD helps reduce superoxide to form hydrogen peroxide (H2O2), which is then converted (detoxified) by the enzyme catalase to water and O2.

However, sometimes the levels of superoxide rise, for example after alcohol exposure (which generates a lot of NADH). Thus, more hydrogen peroxide is formed and can’t be detoxified by the limited amount of catalase. Instead hydrogen peroxide becomes reduced by iron (Fe 2+ ) (normally present in cells), which donates an electron to produce the hydroxyl radical (•OH), a very nasty molecule. It is extremely reactive, and it’s a great oxidizing agent. The hydroxyl radical oxidizes cellular components such as lipids, proteins, and DNA by literally stealing an e- (associated with an H atom) from them, damaging cells.

Figure: The metabolism (i.e., oxidation) of alcohol produces NADH, which acts as an electron donor for the electron transport chain (molecules designated with roman numerals). Electrons (e-) that “leak out” of the electron transport chain (stars at I and III) combine with oxygen to produce superoxide radicals (O2•-). Through a series of reactions the superoxide radicals generate hydroxyl (OH•) radicals. Oxygen radicals are circled in red.

Metabolism can become imbalanced.

Cells have to adapt to the amount of nutrients that are available. So if there is an imbalance with the cell's ability to sense or process nutrients, that causes problems.

With age, cells become less accurate at detecting the amount of glucose or fat that's in the body, so some fats and sugars don't get properly processed. Aging cells accumulate an excessive amount of fats not because older people ingest a lot of fat, but because cells don't digest it properly. This can affect the insulin and IGF-1 pathway, which play a role in diabetes.

This is why age-related diabetes is fairly common — older adults' bodies can no longer properly metabolize all the things they eat.


Oedema forms when fluid is allowed to move from one body fluid compartment to another. This is generally the result of an underlying condition.

There are two main forms of oedema - interstitial and intracellular:

- In interstitial oedema, a number of common pathways can be identified, caused by changes in capillary dynamics (hypertension, heart failure, malnutrition, renal disorders), blocked lymphatic system (cancer), or by stimulation of the inflammatory response (trauma, injury, infection)

- Intracellular oedema results in a lack of oxygen supply to cells (such as with pressure ulcers or myocardial infarction), leading to cellular hypoxia. Alterations occur to the plasma membrane due to the reduction in adenosine triphosphate.

The two types of oedema formation are not mutually exclusive, as the development of one can lead to formation of the other. An accumulation of fluid in the interstitial space or in the cells can destroy adjacent healthy cells and prevent regeneration of damaged tissues. The whole process can be exaggerated by the stimulation of the stress response, further stimulating inflammation, resulting in increased damage and a worsened condition.

Buckman, R.F., Badellino, M.M., Goldberg, A. (1992)Pathophysiology of hemorrhagic hypovolemia and shock. Trauma Quarterly 8: 4, 12-27.

Carelock, J., Clark, A.P. (2001)Heart failure: pathophysiologic mechanisms. American Journal of Nursing 101: 12, 26-33.

Edelstein, C.L., Ling, H., Schrier, R.W. (1997)The nature of renal cell injury Kidney International 51: 5, 1341-1351.

Edwards, S. (2000)Maintaining optimum nutrition. In: Manley, K., Bellman, L. Surgical Nursing: Advancing practice. Edinburgh: Churchill Livingstone.

Edwards, S. (2001)Regulation of water, sodium and potassium: implications for practice. Nursing Standard 15: 22, 36-42.

Edwards, S. (2002)Physiological insult/injury: pathophysiology and consequences. British Journal of Nursing 11: 4, 263-274.

Edwards, S. (2003)The formation of oedema. Part 1: pathophysiology, causes and types. Professional Nurse 19: 1, 29-31.

Germann, W.J., Stanfield, C.L. (2002)Principles of Human Physiology. San Francisco, Ca: Benjamin Cummings.

Huddleston, V. (1992)The inflammatory/immune response: implications for the critically ill. In: Huddleston, V. (ed.). Multisystem Organ Failure: Pathophysiology and clinical implications. St. Louis, Mo: Mosby Year Books.


    NDSS. (n.d.). What causes Down syndrome? Retrieved June 11, 2012, from Parker, S. E., Mai, C. T., Canfield, M. A., Rickard, R., Wang, Y., Meyer, R. E., et al. (2010). Updated national birth prevalence estimates for selected birth defects in the United States, 2004–2006. Birth Defects Research. Part A, Clinical and Molecular Teratology, 88, 1008–1016. Genetic and Rare Diseases Information Center (GARD). (2012). Down syndrome. Retrieved June 11, 2012, from

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