Saori Ogasawara

Correction of protein kinase C activity and macrophage migration in peripheral nerve by pioglitazone, peroxisome proliferator activated-γ-ligand, in insulin-deficient diabetic rats

Pioglitazone, one of thiazolidinediones, a peroxisome proliferator‐activated receptor (PPAR)‐γ ligand, is known to have beneficial effects on macrovascular complications in diabetes, but the effect on diabetic neuropathy is not well addressed. We demonstrated the expression of PPAR‐γ in Schwann cells and vascular walls in peripheral nerve and then evaluated the effect of pioglitazone treatment for 12 weeks (10 mg/kg/day, orally) on neuropathy in streptozotocin‐diabetic rats. At end, pioglitazone treatment improved nerve conduction delay in diabetic rats without affecting the expression of PPAR‐γ. Diabetic rats showed suppressed protein kinase C (PKC) activity of endoneurial membrane fraction with decreased expression of PKC‐α. These alterations were normalized in the treated group. Enhanced expression of phosphorylated extracellular signal‐regulated kinase detected in diabetic rats was inhibited by the treatment. Increased numbers of macrophages positive for ED‐1 and 8‐hydroxydeoxyguanosine‐positive Schwann cells in diabetic rats were also corrected by the treatment. Pioglitazone lowered blood lipid levels of diabetic rats, but blood glucose and nerve sorbitol levels were not affected by the treatment. In conclusion, our study showed that pioglitazone was beneficial for experimental diabetic neuropathy via correction of impaired PKC pathway and proinflammatory process, independent of polyol pathway.

Methylcobalamin effects on diabetic neuropathy and nerve protein kinase C in rats.

Eur J Clin Invest 2011; 41 (4): 442–450

Increased susceptibility to ischemia and macrophage activation in STZ-diabetic rat nerve

Ischemic vulnerability in diabetic nerve plays a paramount role in the development of diabetic neuropathy, yet little is known of the underlying mechanism. Diabetes enhances the inflammatory response to ischemia and reperfusion. We investigated pathological characteristics of nerve fibers and endoneurial macrophages along the length of sciatic-tibial nerves before and after ischemia (60 to 90 min) and reperfusion (6h to 7 days) in 8 weeks of STZ-induced diabetic rats. Without ischemia, diabetic nerves revealed significantly increased the density of Iba-1-positive endoneurial macrophages when compared with controls. Most of macrophages appeared slim and triangular in shape, but in diabetic nerves, some were rounded with bromodeoxyuridine (BrdU) incorporation, suggesting proliferating macrophages. Seventy-five minutes of ischemia is the minimal ischemic time to cause pathological changes in diabetic nerves. Following 90 min of ischemia and 6h of reperfusion in diabetic rats, the number of Iba-1-positive endoneurial macrophages was increased significantly at the thigh level of sciatic nerve when compared with those before ischemia. Endoneurial macrophages in diabetic nerves increased in number further significantly after 24 and 48 h of reperfusion and underwent morphological alterations; swollen and rounded including phagocytosis. After 90 min of ischemia and 7 days of reperfusion, severe pathological alterations, e.g., demyelination and endoneurial edema at proximal nerves and axonal degeneration distally, were observed in diabetic nerves, while control nerves showed normal morphology. We conclude that macrophage proliferation occurs in STZ-diabetic nerves. The acute inflammatory response after ischemia and reperfusion was intensified in diabetic nerves. Activation of resident macrophages and infiltration by recruited macrophages could be casually linked to ischemic susceptibility in diabetic nerve.

The role of the polyol pathway in acute kidney injury caused by hindlimb ischaemia in mice

The polyol pathway, a collateral glycolytic process, previously considered to be active in high glucose milieu, has recently been proposed to play a crucial role in ischaemia/reperfusion tissue injury. In this study, we explored the role of the polyol pathway in acute kidney injury (AKI), a life‐threatening condition, caused by hindlimb ischaemia, and determined if inhibition of the polyol pathway by aldose reductase (AR) inhibitor is beneficial for this serious disorder. Mice 8 weeks of age rendered hindlimb ischaemic for 3 h by the clipping of major supporting arteries revealed marked muscle necrosis with accumulation of sorbitol and fructose in ischaemic muscles. Serum concentrations of blood urea nitrogen (BUN), creatinine phosphokinase (CPK), creatinine, tumour necrosis factor (TNF)‐α as well as interleukin (IL)‐6 were all elevated in these mice. Treatment with AR inhibitor (ARI) effectively suppressed muscle necrosis and accompanying inflammatory reactions and prevented renal failure. Similar to ARI‐treated mice, AR‐deficient mice were protected from severe ischaemic limb injury and renal failure, showing only modest muscle necrosis and significant suppression of serum markers of renal failure and inflammation. Thus, these findings suggest that the polyol pathway is implicated in AKI caused by ischaemic limb injury and that AR may be a potential therapeutic target for this condition. Copyright

A spontaneously immortalized Schwann cell line from aldose reductase-deficient mice as a useful tool for studying polyol pathway and aldehyde metabolism

The increased glucose flux into the polyol pathway via aldose reductase (AR) is recognized as a major contributing factor for the pathogenesis of diabetic neuropathy, whereas little is known about the functional significance of AR in the peripheral nervous system. Spontaneously immortalized Schwann cell lines established from long‐term cultures of AR‐deficient and normal C57BL/6 mouse dorsal root ganglia and peripheral nerves can be useful tools for studying the physiological and pathological roles of AR. These cell lines, designated as immortalized knockout AR Schwann cells 1 (IKARS1) and 1970C3, respectively, demonstrated distinctive Schwann cell phenotypes, such as spindle‐shaped morphology and immunoreactivity to S100, p75 neurotrophin receptor, and vimentin, and extracellular release of neurotrophic factors. Conditioned media obtained from these cells promoted neuronal survival and neurite outgrowth of cultured adult mouse dorsal root ganglia neurons. Microarray and real‐time RT‐PCR analyses revealed significantly down‐regulated mRNA expression of polyol pathway‐related enzymes, sorbitol dehydrogenase and ketohexokinase, in IKARS1 cells compared with those in 1970C3 cells. In contrast, significantly up‐regulated mRNA expression of aldo‐keto reductases (AKR1B7 and AKR1B8) and aldehyde dehydrogenases (ALDH1L2, ALDH5A1, and ALDH7A1) was detected in IKARS1 cells compared with 1970C3 cells. Exposure to reactive aldehydes (3‐deoxyglucosone, methylglyoxal, and 4‐hydroxynonenal) significantly up‐regulated the mRNA expression of AKR1B7 and AKR1B8 in IKARS1 cells, but not in 1970C3 cells. Because no significant differences in viability between these two cell lines after exposure to these aldehydes were observed, it can be assumed that the aldehyde detoxification is taken over by AKR1B7 and AKR1B8 in the absence of AR.

Suppression of Neuropathy Development in Diabetic Rage-Deficient Mice Is Associated with Absence of M1/M2 Macrophage Skewing in the Sciatic Nerve

Infiltrating macrophage (Mφ) into sciatic nerve (SN) is assumed one of the pathogenesis of diabetic polyneuropathy (DPN). In particular, skewing infiltrating Mφ toward pro-inflammatory (M1) Mφ, but not anti-inflammatory (M2) Mφ, possibly contributes to dysfunction of SN in diabetic state. However, the impacts of Mφ into dorsal root ganglia (DRG) in DPN are not yet well explored. For this purpose, male C57BL/6 mice (W) and receptor for AGEs (RAGE) null mice (R) were recruited and rendered diabetic (D) with STZ injection. After 8 weeks, nerve conduction velocity (NCV) and tail flick test were performed. Subsequently, mice were sacrificed to dissect SN and DRG, which were evaluated in the pathology and mRNA expression by qPCR. WD showed significant delay of NCV and increase in threshold for thermal sensation (p Disclosure S. Osonoi: None. H. Mizukami: Research Support; Self; Sanwa Kagaku Kenkyusho Co., Ltd., Mitsubishi Tanabe Pharma Corporation. S. Ogasawara: None. K. Takahashi: None. K. Sango: None. S. Yagihashi: None.

Neuropathy in the spontaneously hypertensive rat: An electrophysiological and histological study

Introduction: Hypertension is identified as a risk factor for development of polyneuropathy. In this study we examined nerve conduction and morphological alteration of peripheral nerves in spontaneously hypertensive rats (SHR). Methods: Motor nerve conduction velocity (MNCV) in the sciatic–tibial nerve and sensory nerve conduction velocity (SNCV) in the sural nerve were measured. Pathological investigations included spinal cord, dorsal root ganglion, and hindlimb nerves in SHR and Wistar–Kyoto rats (WKY) aged 4–64 weeks. Results: Blood pressure was significantly higher in SHR than WKY animals at 4 weeks and elevated further with aging. MNCV and SNCV were significantly slower in SHR compared with WKY after age 24 weeks. Prominent morphological changes in SHR nerves included axonal atrophy and myelin splitting. SHR also had endoneurial microangiopathy with reduplication of basement membrane. Conclusions: SHR showed slowed nerve conduction velocity and pathological abnormalities of hindlimb nerves. Sustained severe hypertension may cause axonal atrophy and endoneurial microangiopathy. Muscle Nerve 54: 756–762, 2016