This article is available online at http://www.jlr.org experience a loss of sensation that occurs in a stockingand-glove distribution and has a severe impact on the individual's quality of life and societal productivity (3-7). The progressive loss of sensory function results from distal-toproximal peripheral nerve damage and dysfunction of sensory neurons. Although the pathogenesis of DPN is not fully understood, recent studies indicate that DPN pathogenesis is directly linked to a continuum of metabolic factors associated with dyslipidemia (8-10).Plasma concentrations of free saturated FAs (SFAs) are commonly elevated in T2D (11). In general, SFAs are classified as inducers of lipotoxicity (12, 13), mitochondrial dysfunction (14-17), and apoptosis (17); however, recent evidence indicates that hydrocarbon chain length of SFAs defines the level of intracellular lipotoxicity (18,19). Moreover, the Western diet, characterized by increased intake of foods with high levels of long-chain SFAs (LCSFAs), including myristate (C14:0), palmitate (C16:0), and stearate (C18:0), and low levels of beneficial medium-chain SFAs (MCSFAs), such as laurate (C12:0) (20-22), is a driving factor in the onset of dyslipidemia and T2D (23). MCSFAs are reported to prevent lipotoxicity and increase mitochondrial energy production (18,24,25). Likewise, equimolar caloric intake of MCSFAs and LCSFAs increases mitochondrial energy expenditure and leads to oxidative metabolic pathways as opposed to the metabolic dysfunction triggered by LCSFAs (24,26). SFAs are also directed to mitochondrial oxidative pathways through hydrocarbon chain length-dependent mechanisms; LCSFAs are targeted into the mitochondrial matrix for -oxidation through a transport system consisting of palmitoyltransferases (18,24,27), whereas MCSFAs are transported independently into the mitochondria, allowing for more efficient modulation of mitochondrial energy production (18,24). Hence, chain length of SFAs plays a critical role in regulating mitochondrial Abstract Dyslipidemia associated with T2D leads to diabetic neuropathy, a complication characterized by sensory neuronal dysfunction and peripheral nerve damage. Sensory dorsal root ganglion (DRG) neurons are dependent on axonal mitochondrial energy production facilitated by mitochondrial transport mechanisms that distribute mitochondria throughout the axon. Because long-chain saturated FAs (SFAs) damage DRG neurons and medium-chain SFAs are reported to improve neuronal function, we evaluated . Chain length of saturated fatty acids regulates mitochondrial trafficking and function in sensory neurons. J. Lipid Res. 2019. 60: 58-70. Supplementary key words diabetes • dyslipidemias • apoptosis • palmitate • stearate • laurate • myristate • mitochondrial depolarizationDiabetic peripheral neuropathy (DPN) is a common complication that affects up to 30% of patients with prediabetes and 50% of T2D patients (1, 2). Patients with DPN

Neuropathy is the most common complication of prediabetes and diabetes and presents as distal-to-proximal loss of peripheral nerve function in the lower extremities. Neuropathy progression and disease severity in prediabetes and diabetes correlates with dyslipidemia in man and murine models of disease. Dyslipidemia is characterized by elevated levels of circulating saturated fatty acids (SFAs) that associate with the progression of neuropathy. Increased intake of monounsaturated fatty acid (MUFA)-rich diets confers metabolic health benefits; however, the impact of fatty acid saturation in neuropathy is unknown. This study examines the differential effect of SFAs and MUFAs on the development of neuropathy and the molecular mechanisms underlying the progression of the complication. Male mice Mus musculus fed a high-fat diet rich in SFAs developed robust peripheral neuropathy. This neuropathy was completely reversed by switching the mice from the SFA-rich high-fat diet to a MUFA-rich high-fat diet; nerve conduction velocities and intraepidermal nerve fiber density were restored. A MUFA oleate also prevented the impairment of mitochondrial transport and protected mitochondrial membrane potential in cultured sensory neurons treated with mixtures of oleate and the SFA palmitate. Moreover, oleate also preserved intracellular ATP levels, prevented apoptosis induced by palmitate treatment, and promoted lipid droplet formation in sensory neurons, suggesting that lipid droplets protect sensory neurons from lipotoxicity. Together, these results suggest that MUFAs reverse the progression of neuropathy by protecting mitochondrial function and transport through the formation of intracellular lipid droplets in sensory neurons.

Diabetic neuropathy (DN) is a highly prevalent complication of type 2 diabetes that is associated with dyslipidemia and causes length-dependent sensory nerve damage in the lower extremities. The primary sensory neurons, dorsal root ganglion (DRG) neurons, extend bundles of axons into the peripheral nerves and transmit sensory information from the limbs. These neurons require a constant supply of mitochondrial derived energy maintained by trafficking of the mitochondria throughout the length of the DRG axon. We have shown that dyslipidemia-associated saturated fatty acid (FA) palmitate impairs mitochondrial trafficking whereas unsaturated FA oleate prevents this impairment. In the current study, we assessed whether FAs alter trafficking of synaptic vesicles in DRG axons to determine whether palmitate impairs mitochondrial trafficking specifically or global organellar transport. Primary DRG neurons were treated with physiological concentrations of FAs ranging from 31.25-250 µM saturated FA palmitate, unsaturated FA oleate, and oleate/palmitate mixtures. Mitochondria and synaptic vesicles showed a significant dose-dependent reduction in motility with increasing concentrations of saturated FA palmitate. However, mitochondria and synaptic vesicles retained motility with treatments of unsaturated FA oleate compared to the controls. Similarly, palmitate induced a trending decrease in velocity of bi-directional motile mitochondria and synaptic vesicles, whereas oleate treatments did not alter mitochondrial and synaptic vesicle velocity. Finally, co-incubation of oleate prevents palmitate-induced inhibition of mitochondrial and synaptic vesicle trafficking. These results suggest that high concentrations of saturated FAs lead to the dysfunction of global organellar transport throughout the DRG axon associated with the development and progression of DN. Disclosure G. LoGrasso: None. A. Rumora: None. J.A. Haidar: None. S.I. Lentz: None. E.L. Feldman: None.

Peripheral neuropathy is a prevalent complication of diabetes and prediabetes characterized by distal-to-proximal peripheral nerve damage. Dyslipidemia is a contributory factor that underlies neuropathy development in type 2 diabetes and prediabetes. However, the molecular pathways that underlie sensory neuron dysfunction in peripheral neuropathy associated with diabetes and prediabetes are unknown. Our recent lipidomic and transcriptomic analyses identified altered metabolism of polyunsaturated omega-6 fatty acid linoleic acid in the peripheral nerves of prediabetic mice with neuropathy. However, the molecular effect of linoleic acid on sensory neurons remains unknown. Since sensory dorsal root ganglion (DRG) neurons are dependent on trafficking mechanisms to transport healthy mitochondria throughout the length of the axon, we evaluated the effect of linoleic acid on axonal mitochondrial trafficking and function in DRG neurons. Primary DRG neurons from adult C57BL/6J mice were treated with physiological concentrations of linoleic acid ranging from 31.25-250 µM for 24 hours. Following treatment, we evaluated the effect of linoleic acid treatments on mitochondrial trafficking, mitochondrial membrane potential, ATP level, and apoptosis in DRG neurons. Interestingly, all concentrations of linoleic acid ranging from 31.25-250 µM preserved mitochondrial membrane potential and ATP level in DRG neurons. In line with these results, linoleic acid treatments did not stimulate neuronal apoptosis. However, physiological diabetic concentrations of 250 µM linoleic acid significantly impaired mitochondrial trafficking in the axon of DRG neurons. Overall, these results suggest that elevated levels of linoleic acid impair axonal mitochondrial trafficking without effecting mitochondrial function, and that defects in mitochondrial trafficking of sensory neurons may contribute to neuropathy pathogenesis in diabetes and prediabetes. Disclosure A. Rumora: None. G. LoGrasso: None. J.D. McGrath: None. S.I. Lentz: None. E.L. Feldman: Consultant; Self; Novartis Pharmaceuticals Corporation. Funding American Diabetes Association (7-12-BS-045 to E.L.F.); National Institutes of Health (1R24082841, 1DP3DK094292); National Institute of Diabetes and Digestive and Kidney Diseases (T32DK101357, F32DK112642, K99DK119366, P30DK020572); NeuroNetwork for Emerging Therapies; A. Alfred Taubman Medical Research Institute

Diabetic neuropathy (DN) is a common complication of type 2 diabetes characterized by peripheral nerve damage and sensory loss. Dorsal root ganglion (DRG) sensory neurons require axonal mitochondrial transport to produce axonal ATP for neuronal function. In DN, DRG neurons exhibit axonal mitochondrial dysfunction and bioenergetic failure. A correlation between DN progression and dyslipidemia suggests that increased plasma saturated fatty acids (FAs) and decreased unsaturated FAs may play a role in the progression of DN. In this study, we evaluated the effect of saturated FA palmitate and monounsaturated FA oleate on mitochondrial trafficking and mitochondrial function in DRG neurons. Primary DRG neurons were treated with physiological concentrations of palmitate, oleate, and oleate/palmitate mixtures. The impact of these treatments was assessed by measuring mitochondrial motility and mitochondrial membrane depolarization using TMRM staining. Treatment with 62.5 μM to 250 μM palmitate induced a significant and dose-dependent decrease in axonal mitochondrial transport whereas oleate treatments from 31.25 to 250 μM did not alter the percentage of motile mitochondria. Palmitate treatments also resulted in a reduction of mitochondrial membrane potential in DRG neurons treated with 125 to 250 μM palmitate. Conversely, DRG neurons treated with 125 to 250 μM oleate maintained their mitochondrial membrane potential. Based on the differential effects of oleate and palmitate, we next evaluated whether oleate supplementation could prevent palmitate-induced inhibition of mitochondrial trafficking. Mixtures of oleate/palmitate at 1:1 or 2:1 molar ratios prevented palmitate-induced decreases in mitochondrial trafficking and mitochondrial membrane depolarization. These results indicate that monounsaturated FAs prevent saturated FA induced mitochondrial dysfunction in DRG neurons associated with dyslipidemia and DN. Disclosure A. Rumora: None. G. LoGrasso: None. J.A. Haidar: None. J. Dolkowski: None. S.I. Lentz: None. E.L. Feldman: None.