NME3 is a gatekeeper for DRP1-dependent mitophagy in hypoxia.

NME3 is a member of the nucleoside diphosphate kinase (NDPK) family localized on the mitochondrial outer membrane (MOM). Here, we report a role of NME3 in hypoxia-induced mitophagy dependent on its active site phosphohistidine but not the NDPK function. Mice carrying a knock-in mutation in the Nme3 gene disrupting NME3 active site histidine phosphorylation are vulnerable to ischemia/reperfusion-induced infarction and develop abnormalities in cerebellar function. Our mechanistic analysis reveals that hypoxia-induced phosphatidic acid (PA) on mitochondria is essential for mitophagy and the interaction of DRP1 with NME3. The PA binding function of MOM-localized NME3 is required for hypoxia-induced mitophagy. Further investigation demonstrates that the interaction with active NME3 prevents DRP1 susceptibility to MUL1-mediated ubiquitination, thereby allowing a sufficient amount of active DRP1 to mediate mitophagy. Furthermore, MUL1 overexpression suppresses hypoxia-induced mitophagy, which is reversed by co-expression of ubiquitin-resistant DRP1 mutant or histidine phosphorylatable NME3. Thus, the site-specific interaction with active NME3 provides DRP1 a microenvironment for stabilization to proceed the segregation process in mitophagy.

Nerve pathology of microangiopathy and thromboinflammation in hereditary transthyretin amyloidosis.

OBJECTIVE: Despite amyloid deposition as a hallmark of hereditary transthyretin amyloidosis (ATTRv) with polyneuropathy, this pathology could not completely account for nerve degeneration. ATTRv patients frequently have vasomotor symptoms, but microangiopathy hypothesis in ATTRv was not systemically clarified. METHODS: This study examined the vascular pathology of sural nerves in ATTRv patients with transthyretin (TTR) mutation of p.Ala117Ser (TTR-A97S), focusing on morphometry and patterns of molecular expression in relation to nerve degeneration. We further applied human microvascular endothelial cell (HMEC-1) culture to examine the direct effect of TTR-A97S protein on endothelial cells. RESULTS: In ATTRv nerves, there was characteristic microangiopathy compared to controls: increased vessel wall thickness and decreased luminal area; both were correlated with the reduction of myelinated fiber density. Among the components of vascular wall, the area of collagen IV in ATTRv nerves was larger than that of controls. This finding was validated in a cell model of HMEC-1 culture in which the expression of collagen IV was upregulated after exposure to TTR-A97S. Apoptosis contributed to the endothelial cell degeneration of microvasculatures in ATTRv endoneurium. ATTRv showed prothrombotic status with intravascular fibrin deposition, which was correlated with (1) increased tissue factor and coagulation factor XIIIA and (2) reduced tissue plasminogen activator. This cascade led to intravascular thrombin deposition, which was colocalized with upregulated p-selectin and thrombomodulin, accompanied by complement deposition and macrophages infiltration, indicating thromboinflammation in ATTRv. INTERPRETATION: Microangiopathy with thromboinflammation is characteristic of advanced-stage ATTRv nerves, which provides an add-on mechanism and therapeutic target for nerve degeneration.

HSP27 Modulates Neuropathic Pain by Inhibiting P2X3 Degradation.

The role of heat shock protein 27 (HSP27), a chaperone, in neuropathic pain after nerve injury has not been systematically surveyed despite its neuroprotective and regeneration-promoting effects. In this study, we found that HSP27 expression in sensory neurons of the dorsal root ganglia (DRG) mediated nerve injury-induced neuropathic pain. Neuropathic pain behaviors were alleviated by silencing HSP27 in the DRG of a rat spinal nerve ligation (SNL) model. Local injection of an HSP27-overexpression construct into the DRG of naïve rats elicited neuropathic pain behaviors. HSP27 interacted with a purinergic receptor, P2X3, and their expression patterns corroborated the induction and reversal of neuropathic pain according to two lines of evidence: colocalization immunohistochemically and immunoprecipitation biochemically. In a cell model cotransfected with HSP27 and P2X3, the degradation rate of P2X3 was reduced in the presence of HSP27. Such an alteration was mediated by reducing P2X3 ubiquitination in SNL rats and was reversed after silencing HSP27 in the DRGs of SNL rats. In summary, the interaction of HSP27 with P2X3 provides a new mechanism of injury-induced neuropathic pain that could serve as an alternative therapeutic target.

Identification of an 85-kb Heterozygous 4p Microdeletion With Full Genome Analysis in Autosomal Dominant Charcot-Marie-Tooth Disease.

Background and Objectives: Charcot-Marie-Tooth disease (CMT) is a syndrome of a hereditary neurodegenerative condition affecting the peripheral nervous system and is a single gene disorder. Deep phenotyping coupled with advanced genetic techniques is critical in discovering new genetic defects of rare genetic disorders such as CMT. Methods: We applied multidisciplinary investigations to examine the neurophysiology and nerve pathology in a family that fulfilled the diagnosis of CMT2. When phenotype-guided first-tier genetic tests and whole-exome sequencing did not yield a molecular diagnosis, we conducted full genome analysis by examining phased whole-genome sequencing and whole-genome optical mapping data to search for the causal variation. We then performed a systematic review to compare the reported patients with interstitial microdeletion in the short arm of chromosome 4. Results: In this family with CMT2, we reported the discovery of a heterozygous 85-kb microdeletion in the short arm of chromosome 4 (4p16.3)[NC_000004.12:g.1733926_1819031del] spanning 3 genes [TACC3 (intron 6-exon 16), FGFR3 (total deletion), and LETM1 (intron 10-exon14)] that cosegregated with disease phenotypes in family members. The clinical features of peripheral nerve degeneration in our family are distinct from the well-known 4p microdeletion syndrome of Wolf-Hirschhorn syndrome, in which brain involvement is the major phenotype. Discussion: In summary, we used the full genome analysis approach to discover a new microdeletion in a family with CMT2. The deleted segment contains 3 genes (TACC3, FGFR3, and LETM1) that likely play a role in the pathogenesis of nerve degeneration.

An induced pluripotent stem cell-based model identifies molecular targets of vincristine neurotoxicity.

To model peripheral nerve degeneration and investigate molecular mechanisms of neurodegeneration, we established a cell system of induced pluripotent stem cell (iPSC)-derived sensory neurons exposed to vincristine, a drug that frequently causes chemotherapy-induced peripheral neuropathy. Sensory neurons differentiated from iPSCs exhibit distinct neurochemical patterns according to the immunocytochemical phenotypes, and gene expression of peripherin (PRPH, hereafter referred to as Peri) and neurofilament heavy chain (NEFH, hereafter referred to as NF). The majority of iPSC-derived sensory neurons were PRPH positive/NEFH negative, i.e. Peri(+)/NF(-) neurons, whose somata were smaller than those of Peri(+)/NF(+) neurons. On exposure to vincristine, projections from the cell body of a neuron, i.e. neurites, were degenerated quicker than somata, the lethal concentration to kill 50% (LC50) of neurites being below the LC50 for somata, consistent with the clinical pattern of length-dependent neuropathy. We then examined the molecular expression in the MAP kinase signaling pathways of, extracellular signal-regulated kinases 1/2 (MAPK1/3, hereafter referred to as ERK), p38 mitogen-activated protein kinases (MAPK11/12/13/14, hereafter referred to as p38) and c-Jun N-terminal kinases (MAPK8/9/10, hereafter referred to as JNK). Regarding these three cascades, only phosphorylation of JNK was upregulated but not that of p38 or ERK1/2. Furthermore, vincristine-treatment resulted in impaired autophagy and reduced autophagic flux. Rapamycin-treatment reversed the effect of impaired autophagy and JNK activation. These results not only established a platform to study peripheral degeneration of human neurons but also provide molecular mechanisms for neurodegeneration with the potential for therapeutic targets.

Genotoxicity and 28-day repeated dose oral toxicity study of garlic essential oil in mice.

Background and aim: Garlic essential oil (GEO) isolated from Garlic (Allium sativum L.) exerts biological activities in disease prevention, particularly in metabolic and liver diseases, and is used for a dietary therapy for centuries. However, due to the side effects associated with the excessive consumption of GEO, there is a need to evaluate the safety of the GEO. Experimental procedure: Ames test using five Salmonella typhimurium strains (TA98, TA100, TA102, TA1535, and TA1537) and Chinese hamster ovary (CHO-K1) cells with or without metabolic activation (S9 system), and mammalian erythrocyte micronucleus test were used to assess the genotoxicity and clastogenic effects of GEO. A repeated dose of GEO (15, 25, and 50 mg/kg body weight, p.o.) were administrated to ICR mice for 28 days to ascertain the subacute toxicity of GEO. Results and conclusions: The results of the Ames test with or without S9 system indicated that GEO did not induce mutagenicity nor have clastogenic effects in CHO-K1 cells with or without S9 activation. Furthermore, GEO did not affect the ratio of immature to total erythrocytes or the number of micronuclei in immature erythrocytes of ICR mice after 24 and 48 h. In a 28-day oral toxicity assessment, GEO (15, 25, and 50 mg/kg body weight, p.o.)-fed ICR mice exhibited normal behaviors, mortality, body weight, daily intake, hematology, clinical biochemistry, and organ weight. GEO shows no genotoxicity, and the no-observed-adverse-effect level (NOAEL) for GEO is considered to be greater than 50 mg/kg bw/day orally for 28 days in mice.

Small Fiber Pathology in CADASIL: Clinical Correlation With Cognitive Impairment.

BACKGROUND AND OBJECTIVES: This study investigated the cutaneous small fiber pathology of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) and its clinical significance, that is, the NOTCH3 deposition in cutaneous vasculatures and CNS neurodegeneration focusing on cognitive impairment. METHODS: Thirty-seven patients with CADASIL and 59 age-matched healthy controls were enrolled to evaluate cutaneous small fiber pathology by quantitative measures of intraepidermal nerve fiber density (IENFD), sweat gland innervation, and vascular innervation. Cognitive performance of patients with CADASIL was evaluated by a comprehensive neuropsychological assessment, and its association with small fiber pathology was tested using multivariable linear regression analysis adjusted for age and diabetes mellitus. We further assessed the relationships of IENFD with cutaneous vascular NOTCH3 ectodomain (NOTCH3ECD) deposition and biomarkers of neurodegeneration including structural brain MRI measures, serum neurofilament light chain (NfL), glial fibrillary acidic protein (GFAP), tau, and ubiquitin carboxy-terminal hydrolase L1. RESULTS: Patients with CADASIL showed reduced IENFD (5.22 ± 2.42 vs 7.88 ± 2.89 fibers/mm, p = 0.0001) and reduced sweat gland (p < 0.0001) and vascular (p < 0.0001) innervations compared with age-matched controls. Reduced IENFD was associated with impaired global cognition measured by Mini-Mental State Examination (B = 1.062, 95% CI = 0.370-1.753, p = 0.004), and this association remained after adjustment for age and diabetes mellitus (p = 0.043). In addition, IENFD in patients with CADASIL was associated with mean cortical thickness (Pearson r = 0.565, p = 0.0023) but not white matter hyperintensity volume, total lacune count, or total microbleed count. Reduced IENFD was associated with cutaneous vascular NOTCH3ECD deposition amount among patients harboring pathogenic variants in exon 11 (mainly p.R544C) (B = -0.092, 95% CI = -0.175 to -0.009, p = 0.031). Compared with those with normal cognition, patients with CADASIL with cognitive impairment had an elevated plasma NfL level regardless of concurrent small fiber denervation, whereas only patients with both cognitive impairment and small fiber denervation showed an elevated plasma GFAP level. DISCUSSION: Cutaneous small fiber pathology correlates with cognitive impairment and CNS neurodegeneration in patients with CADASIL, indicating a peripheral neurodegenerative process related to NOTCH3ECD aggregation.

Distinct Patterns of Amyloid Pathology in Autopsies of Transthyretin S77Y and A97S Amyloidosis: Significance of Symptomatology and Cell Biology.

Hereditary transthyretin (ATTRv) amyloidosis is a systemic disease with amyloid deposition in the peripheral and autonomic nervous systems caused by mutation of transthyretin (TTR) gene. The mutant TTR S77Y is the second prevalent mutation in many countries. In Taiwan, A97S mutant accounts for more than 90% of cases. Although distinct clinical manifestations such as dysphagia, carpal tunnel syndrome, and sudden cardiac death occur, the underlying pathology has not been elucidated. Here, we report the first autopsy cases of ATTRv S77Y and A97S and comprehensively compare the pathology underlying the unique clinical manifestations. This study demonstrated the following: (1) distinct spatial patterns of amyloid deposits in peripheral nerves, with a tendency toward more amyloid deposition in the large peripheral nerves, particularly the median nerves, and scarcely in the sural nerves, and different amyloid distribution in different genotypes; (2) amyloid deposits in the conduction system of the heart in addition to surrounding cardiomyocytes; (3) extensive amyloid deposits in the larynx and gastrointestinal tract, contributing to the unique clinical symptom of dysphagia; and (4) characteristic TTR intracytoplasmic inclusions in the hepatocytes of A97S. The pathology of the first autopsied cases of ATTRv S77Y and A97S provides pathology and mechanisms underlying unique clinical manifestations.

Nonpermissive Skin Environment Impairs Nerve Regeneration in Diabetes via Sec31a.

OBJECTIVE: Although the microenvironment for peripheral nerve regeneration is permissive, such a mechanism is defective in diabetes, and the molecular mediators remain elusive. [Correction added on May 11, 2022, after first online publication: In the preceding sentence, "is ok" was changed to "is defective".] This study aimed to (1) investigate the relationship between skin innervation and collagen pathology in diabetic neuropathy and to (2) clarify the molecular alterations that occur in response to hyperglycemia and their effects on axon regeneration. METHODS: We addressed this issue using two complementary systems: (1) human skin from patients with diabetic neuropathy and to (2) a coculture model of human dermal fibroblasts (HDFs) with rat dorsal root ganglia neurons in the context of intrinsic neuronal factor and extrinsic microenvironmental collagen and its biosynthetic pathways. RESULTS: In diabetic neuropathy, the skin innervation of intraepidermal nerve fiber density (IENFd), a measure of sensory nerve degeneration, was reduced with similar expression of a growth associated protein 43, a marker of nerve regeneration. In contrast, the content and packing of collagen in the diabetic skin became more rigid than the control skin. Sec31a, a protein that regulates the collagen biosynthetic pathway, was upregulated and inversely correlated with IENFd. In the cell model, activated HDFs exposed to high-glucose medium enhanced the expression of Sec31a and collagen I through the activation of transforming growth factor ß, a profibrotic molecule. Sec31a upregulation impaired neurite outgrowth. This effect was reversed by silencing Sec31a expression and neurite outgrowth was resumed. INTERPRETATION: The current study provides evidence that Sec31a plays a key role in inhibiting nerve regeneration in diabetic neuropathy. ANN NEUROL 2022;91:821-833.

Maladaptive motor cortical excitability and connectivity in polyneuropathy with neuropathic pain.

BACKGROUND AND PURPOSE: Sensory symptoms, especially neuropathic pain, are common in polyneuropathy. Conventional diagnostic tools can evaluate structural or functional impairment of nerves but cannot reveal mechanisms of neuropathic pain. Changes in the brain after polyneuropathy may play roles in the genesis of neuropathic pain. METHODS: This cross-sectional study investigated changes of cortical excitability within left primary motor cortex (M1) by measuring resting motor thresholds, short-interval intracortical inhibition (SICI), intracortical facilitation (ICF), and afferent inhibition between polyneuropathy patients and controls, and investigated the correlates of these parameters with neuropathic pain and M1 structural and functional connectivity assessed by diffusion tractography imaging and functional magnetic resonance imaging. RESULTS: Thirty-three painful and 15 nonpainful neuropathic patients and 21 controls were enrolled. There were no differences in intraepidermal nerve fiber density, nerve conduction studies, thermal thresholds, or autonomic functional tests between patients with and without neuropathic pain. Compared to controls, neuropathic patients exhibited similar resting motor thresholds or afferent inhibition, but attenuated SICI and augmented ICF, especially in painful patients. Changes of intracortical excitability in neuropathic patients were correlated with intensities of neuropathic pain, and different presentations of SICI and ICF were noted between patients with and without thermal paresthesia. Additionally, short-latency afferent inhibition at an interstimulus interval of 20 ms was associated with structural connectivity of left M1 with brain areas associated with pain perception. CONCLUSIONS: Maladaptive cortical excitability with altered structural connectivity in left M1 developed after peripheral nerve degeneration and was associated with neuropathic pain and sensory symptoms in polyneuropathy.

Capping Protein Regulator and Myosin 1 Linker 3 (CARMIL3) as a Molecular Signature of Ischemic Neurons in the DWI-T2 Mismatch Areas After Stroke.

Ischemic stroke with a mismatch between diffusion-weighted imaging (DWI) and fluid-attenuated inversion recovery (FLAIR) or T2-weighted images indicates onset within 4.5 h, but the pathological substrates in the DWI-T2 mismatch and T2(+) areas remain elusive. In this study, proteomics was used to explore (1) the protein expression profiles in the T2(+), mismatch, and contralateral areas, and (2) the protein with the highest expression in the T2(+) area in the brains of male Sprague-Dawley rats within 4.5 h after middle cerebral artery occlusion (MCAO). The expression of the candidate protein was further validated in (1) rat brain subjected to MCAO, (2) rat primary cortical neuronal culture with oxygen-glucose deprivation (OGD), and (3) infarcted human brain tissues. This study showed that apoptosis was observed in the T2(+) and mismatch regions and necroptosis in the T2(+) region of rat brains after MCAO. We identified capping protein regulator and myosin 1 linker 3 (CARMIL3) as the candidate molecule in the T2(+) and mismatch areas, exclusively in neurons, predominantly in the cytoplasm, and most abundant in the mismatch area. The CARMIL3(+) neurons and neurites in the mismatch and T2(+) areas were larger than those in the control area, and associated with (1) increased expression of sulfonylurea receptor 1 (SUR1), indicating edema, (2) accumulation of p62, indicating impaired autophagy, and (3) increase in 8-hydroxy-2'-deoxyguanosine (8-OHdG), indicating oxidative stress. The increased expression of CARMIL3 was validated in a cell model of cortical neurons after OGD and in infarcted human brain tissues. In conclusion, this study shows that the mismatch and T2(+) areas within 4.5 h after ischemia are characterized by upregulated expression of CARMIL3 in neurons, particularly the mismatch area, which is associated with neuronal edema, impaired autophagy, and oxidative stress, indicating that CARMIL3 serves as a molecular signature of brain ischemia.

Early changes of nerve integrity in preclinical carriers of hereditary transthyretin Ala117Ser amyloidosis with polyneuropathy.

BACKGROUND AND PURPOSE: Disease-modifying therapies provide new horizons for hereditary transthyretin amyloidosis with polyneuropathy (ATTRv-PN) to slow neuropathic progression. Initiating treatment at the earliest time requires biomarkers reflecting both small- and large-fiber degeneration in carriers. METHODS: This study included examinations of pathology (intraepidermal nerve fiber [IENF] density), physiology (nerve conduction studies, autonomic function test, and nerve excitability), and psychophysics (thermal thresholds) in carriers to compare to healthy controls and asymptomatic diabetic patients. RESULTS: There were 43 carriers (44.2 ± 11.4 years, p.Ala117Ser in 42 carriers), 43 controls (43.4 ± 12.7 years) including 26 noncarrier families, and 50 asymptomatic diabetic patients (58.1 ± 9.5 years). Carriers had lower IENF densities than controls and similar densities as diabetic patients. Median nerve conduction parameters, especially distal motor latency, were the most frequent neurophysiological abnormality in carriers, could differentiate carriers from controls and diabetic patients, were correlated with IENF densities in carriers but not in controls and diabetic patients, and were correlated with nerve excitability parameters in carriers but not in controls. Fifteen carriers (34.9%) with electrophysiological evidence of median nerve entrapment at the wrist had lower IENF densities and more abnormal conduction parameters than carriers without. We defined nerve dysfunction index-the ratio of median distal motor latency to IENF density-which differentiated carriers from controls. CONCLUSIONS: In late-onset ATTRv-PN carriers with predominant p.Ala117Ser, median conduction parameters were the most common neurophysiological abnormalities and served as surrogate signatures of small- and large-fiber impairment. Combination of median distal motor latency and IENF density can reflect early neuropathy in carriers.

Peripheral Neuropathic Pain: From Experimental Models to Potential Therapeutic Targets in Dorsal Root Ganglion Neurons.

Neuropathic pain exerts a global burden caused by the lesions in the somatosensory nerve system, including the central and peripheral nervous systems. The mechanisms of nerve injury-induced neuropathic pain involve multiple mechanisms, various signaling pathways, and molecules. Currently, poor efficacy is the major limitation of medications for treating neuropathic pain. Thus, understanding the detailed molecular mechanisms should shed light on the development of new therapeutic strategies for neuropathic pain. Several well-established in vivo pain models were used to investigate the detail mechanisms of peripheral neuropathic pain. Molecular mediators of pain are regulated differentially in various forms of neuropathic pain models; these regulators include purinergic receptors, transient receptor potential receptor channels, and voltage-gated sodium and calcium channels. Meanwhile, post-translational modification and transcriptional regulation are also altered in these pain models and have been reported to mediate several pain related molecules. In this review, we focus on molecular mechanisms and mediators of neuropathic pain with their corresponding transcriptional regulation and post-translational modification underlying peripheral sensitization in the dorsal root ganglia. Taken together, these molecular mediators and their modification and regulations provide excellent targets for neuropathic pain treatment.

Skin nerve pathology: Biomarkers of premanifest and manifest amyloid neuropathy.

OBJECTIVE: Small-fiber sensory and autonomic symptoms are early presentations of familial amyloid polyneuropathy (FAP) with transthyretin (TTR) mutations. This study aimed to explore the potential of skin nerve pathologies as early and disease-progression biomarkers and their relationship with skin amyloid deposits. METHODS: Skin biopsies were performed in patients and carriers to measure intraepidermal nerve fiber (IENF) density, sweat gland innervation index of structural protein gene product 9.5 (SGII[PGP9.5]) and peptidergic vasoactive intestinal peptide (SGII[VIP]), and cutaneous amyloid index. These skin pathologies were analyzed with clinical disability assessed by FAP stage score (stage 0-4) and compared to neurophysiological and psychophysical tests. RESULTS: There were 70 TTR-mutant subjects (22 carriers and 48 patients), and 66 cases were TTR-A97S. Skin nerve pathologies were distinct according to stage. In carriers, both skin denervation and peptidergic sudomotor denervation were evident: (1) IENF density was gradually reduced from stage 0 through 4, and (2) SGII(VIP) was markedly reduced from stage 1 to 2. In contrast, SGII(PGP9.5) was similar between carriers and controls, but it declined in patients from stage 2. Skin amyloids were absent in carriers and became detectable from stage 1. Cutaneous amyloid index was correlated with SGII(PGP9.5) and stage in a multivariate mixed-effect model. When all tests were compared, only IENF density, SGII(PGP9.5), and cutaneous amyloid index were correlated with stage, and IENF density had the highest abnormal rate in carriers. INTERPRETATION: Biomarkers of sensory and sudomotor innervation exhibited a stage-dependent progression pattern, with sensory nerve degeneration as the early skin nerve pathology. Ann Neurol 2019;85:560-573.

CD40-mediated HIF-1α expression underlying microangiopathy in diabetic nerve pathology.

To understand the pathology and molecular signatures of microangiopathy in diabetic neuropathy, we systemically and quantitatively examined the morphometry of microvascular and nerve pathologies of sural nerves. In the endoneurium of diabetic nerves, prominent microangiopathy was observed, as evidenced by reduced capillary luminal area, increased capillary basement membrane thickness and increased proportion of fibrin(+) blood vessels. Furthermore, capillary basement membrane thickness and the proportion of fibrin(+) blood vessels were correlated with small myelinated fiber density in diabetic nerves. In diabetic nerves, there was also significant macrophage and T cell infiltration, and cluster of differentiation 40 (CD40) expression was increased. The molecular alterations observed were upregulation of hypoxia-inducible factor-1α (HIF-1α), mitogen-activated protein kinase-activated protein kinase 2 (MK2; MAPKAPK2) and phosphatase and tensin homolog (PTEN). In addition, HIF-1α was correlated with small myelinated fiber density and capillary luminal area, while both MK2 and PTEN were correlated with capillary basement membrane thickness. The molecular cascades were further demonstrated and replicated in a cell model of microangiopathy on human umbilical vein endothelial cells (HUVECs) exposed to high-glucose medium by silencing of CD40, PTEN and HIF-1α in HUVECs using shRNA. These data clarified the hierarchy of the molecular cascades, i.e. upregulation of CD40 leading to HIF-1α expression in endothelium and nerve fibers. In conclusion, this study revealed the association of microangiopathy, thrombosis and inflammatory infiltrates with nerve degeneration in diabetic nerves, demonstrating that CD40 is a key molecule for the upregulation of HIF-1α and PTEN underlying the severity of microangiopathy.

Unfractionated bone marrow cells attenuate paraquat-induced glomerular injury and acute renal failure by modulating the inflammatory response.

The aim of this study was to evaluate the efficacy of unfractionated bone marrow cells (BMCs) in attenuating acute kidney injury (AKI) induced by paraquat (PQ) in a mouse model. PQ (55 mg/kg BW) was intraperitoneally injected into C57BL/6 female mice to induce AKI, including renal function failure, glomerular damage and renal tubule injury. Glomerular podocytes were the first target damaged by PQ, which led to glomerular injury. Upon immunofluorescence staining, podocytes depletion was validated and accompanied by increased urinary podocin levels, measured on days 1 and 6. A total of 5.4 × 10(6) BMCs obtained from the same strain of male mice were injected into AKI mice through the tail vein at 3, 24, and 48 hours after PQ administration. As a result, renal function increased, tubular and glomerular injury were ameliorated, podocytes loss improved, and recipient mortality decreased. In addition, BMCs co-treatment decreased the extent of neutrophil infiltration and modulated the inflammatory response by shifting from pro-inflammatory Th1 to an anti-inflammatory Th2 profile, where IL-1ß, TNF-α, IL-6 and IFN-γ levels declined and IL-10 and IL-4 levels increased. The present study provides a platform to investigate PQ-induced AKI and repeated BMCs injection represents an efficient therapeutic strategy.