Abstract
Diabetes mellitus is a chronic disease and it has been associated with several complications affecting peripheral and central nervous system. Proper axonal transport is crucial for neuronal maintenance and function. Alterations in axonal transport have been correlated with the progression of several neurodegenerative diseases. Axonal transport impairment induces functional changes in the distal axon leading to its degeneration. This might occur due to alterations in neuronal cytoskeleton and motor proteins, impaired ATP supply or neuroinflammatory processes which disable molecular motors to undertake transport along the axon.
Studies in experimental models of diabetes have demonstrated that axonal transport may be impaired. The majority of research on this topic has addressed nerve dysfunction under diabetes focusing in changes occuring at peripheral nervous system. Nevertheless, studies at central nervous system, namely in the retina and the hippocampus, suggest that axonal transport is impaired in diabetes and may play a role in diabetic retinopathy and diabetic encephalopathy. Alterations in several components of the transport machinery, changes in neuronal cytoskeleton and changes in axonal transport of distal trophic signals to thecell body of neuronshave been described under diabetes. This article reviews currently state of knowledge about axonal transport defects in diabetes which might contribute to the progression of several diabetic complications.
Keywords: Diabetes; axonal transport; cytoskeleton; kinesin; dynein; neurodegeneration
Abbreviations
ATP: Adenosine triphosphate
CNS: Central nervous system
KIF: Kinesin superfamily protein
NF: Neurofilament
NGF: Nerve growth factor
NT-3: Neurotrophin-3
PNS: Peripheral nervous system
TNF: Tumor necrosis factor
RGC: Retinal ganglion cell
STZ: Streptozotocin
Introduction
Diabetes mellitus is a group of metabolic diseases characterized by chronic hyperglycemia that result from defects in the body’s ability to produce and/or use insulin. Insulin is produced by the pancreatic β-cells and helps glucose from the bloodstream entering into the cells in order to be converted into energy. In diabetes, there is an inadequate supply of glucose to the cells and glucose remains circulating in the bloodstream. Global diabetes incidence is increasing drastically and it is estimated that 642 million people will have diabetes in 2040 (1).
Axonal transport is the fundamental route in neurons by which both soma and synapse can communicate. Throughaxonal transport the synapse is supplied with proteins and lipids synthesized at the cell body allowing proper intracellular neural transmission, mitochondria is transported along axons for local energy supply, and aggregatedand misfolded proteins are cleared from the distal synapse by transporting them to the soma where they get degraded(2). Moreover, axonal transport allows neurons responding effectively to distal trophic signals or stress insults (2).
Impairment of axonal transport has been emerging as a common factor in several neurodegenerative disorders (3) and is also known to be affected in experimental models of diabetes (4). Here, we review the current state of knowledge about axonal transport impairment in diabetes, focusing on the various components and mechanisms that control such transport both at peripheral (PNS) and central nervous systems (CNS).
Diabetes mellitus and axonal transport
Role of axonal cytoskeleton
Axonal transport impairments and alterations of the cytoskeleton have been associated with numerous types of peripheral neuropathy and also central neurodegenerative diseases (5). Axonal transport takes place along the cellular cytoskeleton which provides structural support to the neuron. The neuronal cytoskeleton is composed by three major components , namely microtubules, intermediate filaments and microfilaments, which can be affected by diabetes.
Microtubules
Microtubules are the main cytoskeleton component responsible for the polarity of the axon. Microtubule minus end defined by the α-tubulin sideis located proximally, nearer to the soma, whereas the the plus end is defined by β-tubulin side, which is located distally, closer to the nerve terminal(6). The polarity of microtubules and consequently of the axon is given by this orientation and therefore directs motors protein to undergo anterograde (toward the plus end) or retrograde (toward the minus end) transport (Figure 1). Conversely, in dendrites, microtubules are found in mixed polarity. Microtubules are essential for axonal transport and any changes in their components may lead to impaired axonal transport under diabetes.
Diabetic neuropathy involves a decrease in axon caliber, axonal transport impairment, and a reduced capacity of nerve regeneration, which are dependent on axonal cytoskeleton integrity for proper nerve function (4). Reduced synthesis of tubulin mRNA and an elevated non-enzymatic glycation of peripheral nerve tubulin was described. Particularly, it was demonstrated that after eight weeks of diabetes T alpha 1 alpha-tubulin mRNA is reduced in streptozotocin (STZ)-induced diabetic rats (7), and an increase in tubulin glycation was detected in the sciatic nerve of STZ-induced diabetic rats after two weeks of diabetes duration, which may contribute to axonal transport abnormalities by impairment of microtubule function (8, 9). Brain tubulin is also glycated in early experimental diabetes, consequently affecting its ability to form microtubules (10). Nevertheless, this finding was not replicated in subsequent studies, where it was demonstrated that glycation was not associated with inhibition of microtubule assembly (8, 11). In the sural nerves of diabetic patients it was detected an increase in advanced glycation end products accumulation in cytoskeletal proteins (12), suggesting that axonal cytoskeletal proteins glycation may play a role in axonal degeneration polyneuropathy in diabetes.
Tau is a microtubule associated protein important for the stability of axonal microtubules. Tau hyperphosphorylation impairs its binding to microtubules, changing the trafficking route for molecules which may ultimately lead to synaptic degeneration (13, 14). Diabetes induces tau hyperphosphorylation in the brain, as for example in the hippocampus (15), and proteolytic tau cleavage (16), being both processes occuring in Alzheimer’s disease (17). Hyperglycemia and insulin dysfunction may induce tau modifications, and therefore may play a role for the increased incidence of Alzheimer’s disease in diabetic patients (16). Tau modification impairs axonal transport through microtubule arrangement disruption and by blocking axonal trafficking route, which can culminate in synaptic function changes and consequent neurodegeneration (18, 19). In Alzheimer’s disease, glycation of tau may stabilize paired helical filaments aggregation leading to tangle formation (20). It is likely that similar processes may be occuring under diabetes.
Neurofilaments
Neurofilaments (NF) are the intermediate filaments (10 nm) found specifically in neurons that assemble from three subunits based on molecular weight: NF-L (70 kDa), NF-M (150 kDa), and NF-H (200 kDa) (21). Neurofilaments lack overall polarity upon assembly and mainly provide neuronal structural stabilization and regulate axonal growth (22). Aggregation of neurofilaments is a common marker of neurodegenerative diseases (23). Abnormal NF expression, processing, and structure may contribute to diabetic neuropathy, since reduced synthesis of NF proteins or formation of incorrectly associated NFs could severely disrupt the axonal cytoskeleton (24).
Neurofilament mRNAs are selectively reduced in diabetic rats and alterations on post-translational modification of NF proteins have been detected. A reduction of myelinated fiber size is correlated with axonal NFs loss in peripheral nerves of STZ-induced diabetic rats (25, 26), and mRNAs levels encoding for NF-L and NF-H are reduced in the same animal model of diabetes (7). Moreover, changes on the expression of several NF-associated protein kinases isoforms may also contribute to diabetes-induced changes (4). Several protein kinases regulate NF phosphorylation status, being NFs hyperphosphorylation a hallmark of several neurodegenerative diseases. Abnormal NF phosphorylation has been described in sensory neurons of animal models of type 1 diabetes (27). Moreover, in the spinal cord of diabetic rats there is increased phosphorylation of NF-H, (28). Additionally, changes on the activity of Cdk5 and GSK-3β kinases have been described to alter the phosphorylation status of NFs in an animal model of type 1 diabetes. Specifically, in dorsal root ganglion neuronsincreased phosphorylation of GSK-3β correlated linearly with increased phosphorylation of NF-H, while decreasing activity of Cdk5 is associated with reduced phosphorylation of NF-M, which may result in progressive deficits of axonal function (29).
Microfilaments
Microfilaments (or actin filaments) are the thinnest filaments of the cytoskeleton, having 6 nm in diameter, providing both stability and dynamics to neurons. In neurons, actin filaments are packed into networks and stabilized by interacting proteins (22). Microfilaments play a role in spine formation and spine volume stabilization (30), with the dynamics of actin leading to the formation of new synapses as well as increased cell communication. The actin cytoskeleton controls several cellular processes. In animal models of diabetes there is an impairment of slow axonal transport of cytoskeletal elements like tubulin and NF proteins (slow component a), and polypeptides such as actin (slow component b) (31-33). Actin undergoes glycation in the brain of STZ-induced diabetic rats and the appearance of glycated actin is prevented by administration of insulin (9, 34).
More recently, it was investigated if the receptor for advanced glycation end-product (RAGE) is involved in axonal transport impairment via interaction with its cytoplasmic domain binding partner mDia1, which is involved in actin structure modifications. Slow axonal transport in the peripheral nerves is indeed affected by diabetes, but in a RAGE-independent manner (35). Moreover, mDia1 axonal transport is impaired, suggesting that diabetes-induced changes affecting actin binding proteins are early events in the course of the pathology (35), and forward the hypothesis that mDia1 axonal transport impairment might be correlated with the extent of actin glycation (34).
Taken together, these studies in experimental diabetes indicate that post-translational modifications, as well as altered expression of cytoskeletal proteins (tubulin, neurofilament and actin), may interfere with cytoskeletal assembly, contributing to altered axonal transport and subsequent nerve dysfunction.
Motor proteins and diabetes
Intracellular transport is carried out by three sets of molecular motors: myosins, kinesins and dyneins. Myosins move on microfilaments and are thought to be responsible for short range transport, whereas kinesin and dynein proteins use microtubules as tracks for long distance transport and are capable of recognizing the microtubule polarity (36). A number of studies have shown that most kinesin-family motors move towards the plus-end of the microtubules that are usually used to deliver cargos towards the synapse. On the contrary, dynein moves in the opposite direction for transport toward the cell body (37). Kinesins and dyneins have “motor domains” that travel along the microtubules in a distinct direction by using the energy resultant from ATP hydrolysis.
Kinesins are microtubule based anterograde intracellular transport motors (Figure 1). Newly synthesized components at cell body indispensable for neuronal function and maintenance are transported anterogradely from the soma to nerve terminals. Ultrastructural studies have demonstrated many small vesicles, tubulo-vesicular structures, mitochondria and dense-core vesicles move along the axon by fast anterograde transport (38). Components transported in fast anterograde transport are required not only for the supply and turnover of intracellular membrane compartments such as mitochondria, but also for the transport of proteins required for the maintenance of axonal metabolism (3). Conventional kinesins usually form a protein dimer of two identical heavy chains and each of them binds to a kinesin light chain. The heavy chain contains a highly conserved globular head called motor domain, which includes a microtubule-binding site and an ATP-binding/hydrolysis site, a short, flexible neck linker, a stalk domain, which has a long, central coiled-coil region for dimerization and a tail domain for light chain and cargo binding (39). Kinesin move in a hand-over-hand mechanism over long distances along the microtubuleprior to detaching, allowing a very efficient cargo transport (40).
In opposition to anterograde axonal transport, which can be carried out by several different kinesin motor proteins, cytoplasmic dynein is the major motor protein responsible retrograde transport (Figure 1). Cytoplasmic dynein is a multi-subunit complex containing light chains, light intermediate chains and two heavy chains that bind microtubules. The heavy chains contain ATPase activity and are therefore responsible for generating movement along the axon microtubules, while the other chains are involved in cargo binding and binding to dynactin (41). The overall motility tendency is pointed toward microtubule minus end, but the ability of dynein to change directions may allow it to contour obstacles encountered along the axon during transport. Studies on dynein demonstrate that this motor protein wanders along the microtubule, and frequently takes backward steps (42). Also, fluorescently labeled dynein-dynactin complexes present bidirectional motility towards both microtubule ends, showing the flexibility of dynein to avoid obstacles in the cell. (43).
Changes in motor protein content and distribution have been reported under experimental diabetes. In STZ-induced diabetic rats, KIF5B content is increased in the sciatic nerve, as well as KIF5B and SYD (scaffold protein which interacts with kinesin) mRNA levels in spinal cord sensory and motor neurons (44). At CNS level, diabetes decreases the content of the kinesin KIF1A in the retina. Analyzing its immunoreactivity in all retinal layers it was demonstrated that with the exception of the photoreceptor layer, its immunoreactivity is decreased in STZ-induced diabetic rats after 8 weeks of diabetes. KIF5B kinesin immunoreactivity was also changedobserved in the retina of those animals, being increased at the photoreceptor and outer nuclear layers, and decreased in the inner plexiform and ganglion cell layers, whereas no significant changes were detected regarding dynein immunoreactivity in the retina (45).
Cones photoreceptors lacking KIF3A progressively degenerate concomitantly with changes in photopic electroretinogram (46). Additionally, rod photoreceptors lacking KIF3A degenerate rapidly between 2 and 4 weeks (46). In a model retinal neurotoxicity induced by N-methyl-D-aspartate occurs an early upregulation of KIF5B levels in the retina, but a significant downregulation occurs in the optic nerve, indicating that a depletion of KIF5B may precedes axonal degeneration of the optic nerve (47). These studies highlight the importance of proper kinesin function in the visual pathway. Therefore, it is expected that any imbalance in their content due to diabetes might have an impact in axonal transport in the retina.
In the hippocampus of STZ-induced diabetic rats, increased expression and immunoreactivity of KIF1A and KIF5B was detected, but no changes were found concerning dynein (48). In hippocampal cultures incubated with high glucose, mimicking hyperglycemic conditions, there is an increased number of fluorescent KIF1A and synaptotagmin-1 accumulations, and decreased KIF5B, synaptophysin and SNAP-25 immunoreactivity in the axons of hippocampal neurons (48).
Together, the observations of alterations in kinesin motor proteins in the retina and hippocampus triggered by diabetes, indicate that anterograde axonal transport mediated by KIF1A and KIF5B may be impaired in retinal and hippocampal neurons, and may therefore underlie changes already detected in synaptic protein levels in nerve terminals induced by diabetes (48-50).
Axonal transport impairments under diabetes in the PNS
Neurons are highly polarized cells, making them dependent on active intracellular transport. Inhibition of axonal transport rapidly leads to funtional impairments in the distal axon and to “dying back” axonal degeneration. Any impairment of axonal transport is considered an early and possibly causative event in many neurodegenerative diseases.
In experimental diabetes, several studies have demonstrated that axonal transport in PNS is affected by this disease. One of the earliest structural changes detected was a decrease in the calibre of myelinated axons in the rat, suggesting that inhibition of axonal transport of proteins is an early event, possibly the first change to occur, leading to the development of peripheral nerve abnormalities (51). As previously mentioned, several components of the cytoskeleton are affected in diabetes, and the transport of slow components a and components b is impaired (32, 52). Reduced rate of slow axonal transport of proteins occurs in motor fibres of the sciatic nerve of STZ-induced diabetic rats (31, 33, 53, 54). These alterations were correlated with reductions in axonal transport of NF proteins, tubulins and actin. NF protein transport has been shown to be affected at an early stage (53), which could lead to changes in the speed of nerve conduction, since NFs preserve axonal calibre (55). Also, a reduction in the slow axonal transport of cytoskeletal proteins may therefore lead to proximal swelling and distal deterioration of axons in diabetic nerve (26, 56, 57) (Figure 2).
Abnormal synthesis or post-translational modifications of the axonal cytoskeleton can not only influence axonal growth and caliber, but also impair nerve regeneration (58-60). Changes on retrograde signaling from the injury site to the soma were described in experimental diabetes (61, 62), and the delivery of growth factors are also known to be affected (63, 64).
It has been shown that experimental diabetes leads to a decrease in the axonal transport of isotopically labelled glycoproteins and proteins in several types of peripheral nerves (65-69). Studies regarding transport rates along sciatic nerves of STZ-induced diabetic rats suggest that impairment of the retrograde axonal transport is the first event leading to neuronal changes in diabetes (65, 70), followed by impairment of the anterograde axonal transport of structural proteins (66). Fast axonal transport is impaired at sciatic nerve and alterations found in the transport and synthesis of protein and glycoconjugates may be correlated to the early decrease in axon calibre and conduction velocity in peripheral nerves of STZ-induced diabetic rat (68). Regarding retrograde axonal transport, diabetes also induces a decrease in peripheral axon organelle speed in sciatic nerve and dorsal and ventral nerve roots at 1 month of diabetes (71). Moreover, axonal transport of receptors in the sciatic nerves of STZ-induced diabetic rats is also affected, which may contribute to neurological abnormalities detected in diabetic patients (72). Decreased fast axonal transport rates in the sciatic nerve of diabetic animals can be reversed by insulin administration (67). Insulin treatment is able to prevent transport impairments occurring in diabetic rats and reverse an already slowed transport velocity (73). In short-term experimental diabetes, impairments in both anterograde and retrograde axonal transport of 6-phosphofructokinase activity were also detected, which were prevented by insulin treatment (74). Moreover, decreased tubulin and actin transport rates are counteracted by ganglioside treatment suggesting that cytoskeleton proteins may interact with gangliosides and conferring a pharmacological effect (75).
More recently, it was demonstrated that hyperglycemia impairs axonal transport in mice olfactory receptor neurons. Increased oxidative stress in STZ-induced wild type diabetic mice activates the p38 MAPK-tau signaling cascades attenuating axonal transport rates in the olfactory system, whereas in STZ-induced superoxide dismutase-overexpressing mice, in which superoxide levels are decreased, axonal transport deficits are reversed (76).
Diabetes also induces changes in the axonal transport of distal trophic factors to the soma. Increasing evidence shows that retrograde transport impairments of nerve growth factor (NGF) accounts for some functional deficits occurring in experimental diabetic neuropathy. Expression deficits in both NGF and its high-affinity receptor, trkA, were described in animal models of diabetes, resulting in NGF retrograde axonal transport deficits . As a consequence, NGF-dependent sensory neurons support decrease, concomitant with reduced neuropeptide expression by these neurons, such as substance P and calcitonin gene-related peptide (77). NGF and neurotrophin-3 (NT-3) retrograde transport was also found to be decreased in the cervical vagus nerve in diabetic animals (78). In the sciatic nerve of STZ-induced diabetic rats, a clear reduction in the retrograde transport of NGF is detected (79). In STZ-induced diabetic rats the relative levels of phosphorylated (activated) c-Jun N-terminal kinase (JNK) and p38 retrogradly transported in sciatic nerve are increased compared with age-matched controls. Treatment of diabetic animals with NT-3 prevented the activation of JNK and p38, in sciatic nerve suggesting the involvement of JNK and p38 as intermediaries in the transport of diabetes-related stress signals along the axons caused by a deficiency in neurotrophic support, since the periphery to the cell body (80). Moreover, it was also proposed that changes in PI3 kinase/Akt signaling cascade may in part contribute for the retrograde axonal transport impaiments of neurotrophins in the vagus nerve of STZ-induced diabetic rats (81).
Axonal transport impairments under diabetes in the CNS
Although most studies regarding changes in axonal transport triggered by diabetes have been focused in changes occurring at peripheral nervous system, evidences from studies in the optic nerve, spinal cord and brain also indicate that axonal transport is impaired at the central nervous system.
Analysis of slow axonal transport in the optic nerve of STZ-induced diabetic rats (4-6 weeks diabetes duration) and BioBreeding rats with spontaneous diabetes (2.5-3.5 months duration), showed impaired transport of NF subunits, tubulin and actin (26, 31), further evidencing that cytoskeleton proteins transport is altered under diabetes. Moreover, a delay in the transport of 60, 52 and 30 kDa polypeptides, likely to be the glycolytic enzymes aldolase, neuron-specific enolase and pyruvate kinase, respectively, was also detected in both animal models of diabetes (31).
In the brain of STZ-induced diabetic rats, altered electrophoretic mobility of actin from a cytoskeletal protein preparation was detected, suggesting the presence of a product of nonenzymatic glycation (9). Furthermore, diabetes also leads to an hyperphosphorylation of spinal cord neurofilament proteins. These post-translational modifications of neuronal cytoskeletal proteins in the spinal cords and brains of diabetic rats may contribute, as already referred, to the impairment of axonal transport and consequent neuronal dysfunction in experimental diabetes (9).
In the retina of diabetic rabbits, a reduction in slow axonal transport was observed, with a concomitant reduction in fast axonal transport and protein synthesis, whereas protein degradation remains unchanged (82, 83). These evidences suggest that changes in axonal transport in the optic nerve may be indicative of decreased protein synthesis in retinal ganglion cells (RGCs) (84). Studies using fluoro-gold, a neuronal retrograde tracer, demonstrated a decreased accumulation of this tracer in RGCs of STZ-induced diabetic rats, thus suggesting that retrograde axonal transport from the dorsal lateral geniculate nucleus in the brain to the RGCs in the retina is impaired (85, 86). Nevertheless, the accumulation of fluoro-gold in RGCs of OLETF rats, an animal model of type II diabetes with obesity, is not significantly decreased compared with control rats, suggesting that, the impairment of retrograde axonal transport in RGCs is higher in type 1 than in type 2 diabetic rats (85). Changes in polyol pathway may play a role in the gradual impairment of retrograde axonal transport in the optic nerve of STZ-induced diabetic rats, suggesting that it may be a consequence of metabolic changes associated with diabetes (87). Besides changes in retrograde transport, more recently, it was found that anterograde transport in RGCs (from the retina to the superior colliculus) is impaired in STZ-diabetic animals (88, 89).
In summary, changes already described regarding axonal transport in the retina and visual pathway under diabetes (86, 88) may be correlated with alterations in motor protein content and distribution previously detected in the retina (45). .
Mitochondria axonal transport deficits under diabetes
Since neurons are highly polarized cells with long axons, axonal transport implies the localization of functional mitochondria along the axons to supply motor proteins with the required ATP. Proper ATP levels supply is mainly produced by mitochondria via oxidative phosphorylation. The majority of mitochondria are produced in the neuronal soma and then transported along microtubules by motor proteinsuntil they reach areas with high ATP demands (90). Since synapses locate distally, they depend on effective mitochondria axonal transport to fulfill local ATP requirements. Generally, kinesin motor proteins carry out anterograde mitochondrial transport, while dyneins retrogradely transport them. However, mitochondria have saltatory and bidirectional movement,and almost never move in a single direction (90). Their transport involves pauses for short and long periods , having a net mitochondrial velocity of 0.3-2.0 μm/s (91), and may be transported both anterogradely and retrogradely, suggesting that mitochondria are simultaneously coupled to oppositely directed molecular motors (91) (Figure 3). Mitochondria can abruptly change their direction of movement towards regions with high metabolic demand and pause locally to provide ATP.
Mitochondrial axonal transport deficits are described to be involved in the pathogenesis of several neurological disorders (92).
Kinesin KIF5 motor proteins are described to drive axonal transport of mitochondria. In neuronal cultures of KIF5A-/- knockout mice, the velocity of mitochondrial transport is decreased both in anterograde and retrograde direction (93). A decrease in the number of mitochondria axonally located will likely reduce ATP supply to motors proteins leading to impairments both in anterograde and retrograde movement of mitochondria and vesicles (41). Mitochondrial dysfunction may therefore play a significant role in the progression of neurodegenerative diseases like Alzheimer’s disease, Huntington’s disease and amyotrophic lateral sclerosis (94-97).
Regarding mitochondria alterations in diabetes, it was already described that nerve degeneration is mediated by changes in mitochondrial ultrastructure and physiology (98). Furthermore, alterations in mitochondrial trafficking has also been suggested as a mediator of neurodegeneration in diabetic nerves (98), yet, as far as we know, the impact of diabetes on mitochondria axonal transport in the CNS remains to be elucidated. In a previous work, we addressed the question if mitochondria could be affected by exposure to elevated glucose (mimicking hyperglycemia) in the axons of hippocampal neurons. NWe did not found any significant change in the number of accumulations related with mitochondria, intensity of fluorescence or their distribution , when compared to control condition (48). Nevertheless, since diabetes is a multifactorial pathology, mitochondria may be affected in diabetes by other factors, such as the lack of insulin, which may also have an impact in mitochondria transport. Neuroinflammation also plays a role in diabetes complications and may lead to changes in axonal transport of mitochondria. It was reported that tumor necrosis factor (TNF) induces mitochondria clustering around the nucleus of L929 cells by impairing kinesin-mediated transport (99).
Neuroinflammation and axonal transport impairments
Neuroinflammation may contribute to changes in axonal transport (Figure 4). Pro-inflammatory cytokines, such as interleukin-1β and TNF, are upregulated in the hippocampus of diabetic BB/Wor rats (100) and STZ-induced diabetic animals (101). TNF impairs kinesin activity by hyperphosphorylation of kinesin light chain, suggesting that activation TNF receptor-1 signaling pathways may regulate – axonal transport mediated by kinesin motor proteins(102). Moreover, exposure of hippocampal neurons to TNF enhances the phosphorylation of JNK in neuritis, reducing the mobile fraction and inhibiting axonal transport of mitochondria and synaptophysin through dissociation of KIF5B from axon microtubules(103). Evidence also suggests that axonal transport may be impaired by microglia-derived nitric oxide, inhibiting axonal transport of synaptic vesicle precursors containing synaptotagmin and synaptophysin in hippocampal neurons (104).. Also, hydrogen peroxide, an inflammatory mediator, is also able to alter axonal transport of mitochondria in hippocampal cultures (105) and cerebellar slice cultures (106). Additional studies are required to clarify if similar pathways are also beingactivated in diabetes, therefore contributing for previously detected changes in axonal transport (Figure 4).
Conclusions
Defects in axonal transport are early pathogenic events that may contribute for the development and progression of diabetic complications such as diabetic neuropathy, retinopathy and encephalopathy. Disruption of axonal transport can occur through several ways, including alterations in the cytoskeleton, molecular motors, cargoes (via inhibition of their attachment to motors), mitochondrial damage, and through microglia-mediated release of inflammatory mediators (via inhibition of motor attachment to microtubules). Nevertheless, little is known regarding the cellular and molecular mechanisms associated with axonal transport impairments under diabetes. Advances in the understanding of such mechanisms, may be crucial to develop novel neuroprotective strategies for diabetic complications.
Figure Legends
Figure 1. Fast axonal transport. In axons, microtubules are oriented with the plus ends facing toward the synapse and the minus ends toward the cell body. The majority of motors of the kinesin family unidirectionally move toward the microtubule plus end, mediating transport in the direction of the synapse (anterograde). In the opposite direction, the molecular motor cytoplasmic dynein moves to the microtubule minus end and mediates transport of most cargoes toward the cell body (retrograde) Many components essential for neuronal function and maintenance, such as small vesicles, mitochondria and dense-core vesicles are transported anterogradely, whileretrograde transport carries mainly mitochondria and vesicular cargoes such as signaling endosomes.
Figure 2. Morphologic changes occurring in diabetic axons. Diabetic axons present an increased number of microtubules and neurofilaments proximally, as well as increased cross-sectional area near the cell body. On the other hand, axonal cross-sectional area near the synapse is decreased, and the total number of microtubules and neurofilaments also decrease distally. As a consequence of cytoskeleton and nerve terminals derangements, motor proteins fail to bind microtubule end, contributing for the impairment of axonal transport.
Figure 3. Axonal mitochondrial transport. In neurons, mitochondria can be observed to undergo dynamic, bidirectional transport along neuronal axons, changing direction frequently, pausing or being persistently stationary. This dynamic transportpatterns is due to the fact that mitochondria arecoupled to both kinesin and dynein motor proteins, which are opposite directed motors..
Figure 4. Microglia-driven inflammation and axonal transport impairment. In hippocampal cultures, nitric oxide (NO) released from activated microglia impairs axonal transport of synaptic vesicles. Tumor necrosis factor (TNF) produced by activated glial cells in inflammatory or degenerative conditions affects neurites by acting on the kinesin-microtubule complex and impairing axonal transport of mitochondria and synaptophysin in hippocampal cultures. Moreover, hydrogen peroxide (H2O2), a common mediator elevated during inflammation, also inhibits axonal transport.
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