Diabetic peripheral neuropathy (DPN) is a common complication of diabetes characterized by oxidative stress and neuroinflammation. Quercetin, a natural flavonoid with antioxidant properties, has shown potential in treating various nerve injuries, but its mechanisms in DPN remain unclear. This research aims to explore the potential therapeutic mechanisms of quercetin in DPN through network pharmacology and experimental validation. In this study, streptozotocin (STZ)-induced diabetic rats were treated with quercetin for 12 weeks. Behavioral tests and histopathological examination were performed to evaluate nociceptive behaviors and sciatic nerve pathology, and biochemical and molecular assays were used to assess oxidative stress, inflammation, and myelin-related markers. Bioinformatics and network pharmacology were used to predict relevant targets and pathways, followed by in vivo validation in sciatic nerve tissues and in vitro verification using high glucose-exposed RSC96 Schwann cells. Quercetin improved pain thresholds and sciatic nerve morphology without significantly altering blood glucose or body weight. Moreover, quercetin restored the remyelination-related gene expression and attenuated oxidative stress and inflammatory responses in DPN rats. Bioinformatics and network pharmacology analyses indicated that the TXNIP/NLRP3 signaling played a key role. Subsequent in vivo and in vitro experiments further supported that quercetin reduced TXNIP expression and downregulated NLRP3 inflammasome–related gene expression. Our results indicate that quercetin attenuates DPN-like manifestations and mitigates oxidative stress and neuroinflammation. Its potential mechanism may be related to the regulation of TXNIP/NLRP3 inflammasome–related signaling, which lays a foundation for further research on DPN treatment.

We aimed to explore the associations between serum retinol and all-cause mortality among people with prediabetes and diabetes. The study included 2582 participants with prediabetes and 1654 with diabetes aged ≥40 years from the National Health and Nutrition Examination Survey 2001-2006. Serum retinol was collected from laboratory tests and categorized into five groups, including <50, 50-60, 60-70, 70-80, and ≥80 μg/dL. Deaths were obtained by linkage to National Death Index up to December 31, 2019. Cox proportional hazards models were used to estimate the associations between serum retinol and all-cause mortality. During the follow-up, 993 participants with prediabetes died and 874 participants with diabetes died. There were U-shaped associations between serum retinol and mortality among participants with prediabetes and diabetes, separately. Among participants with prediabetes, compared to serum retinol levels of 50-60 μg/dL, the hazard ratio (HR) (95% confidence interval [CI]) of mortality was 1.40 (95% CI 1.11 to 1.76) and 1.26 (95% CI 1.00 to 1.57) for serum retinol <50 or ≥80 μg/dL, respectively. Among participants with diabetes, compared to serum retinol levels of 50-60 μg/dL, the hazard ratio (HR) (95% confidence interval [CI]) of mortality was 1.25 (95% CI 0.96 to 1.62) and 1.21 (95% CI 0.91 to 1.62) for serum retinol <50 or ≥80 μg/dL, respectively. The U-shaped associations between serum retinol and mortality still existed among participants aged ≥60 years with prediabetes or diabetes but were not statistically significant among those aged 40-59 years with prediabetes or diabetes. In conclusion, both low and excessive serum retinol tended to be with higher mortality risk among people with abnormal blood glucose.

Tu-Xian decoction (TXD), a traditional Chinese medicine (TCM) formula, has been frequently administered to manage diabetic cognitive impairment (DCI). Despite its widespread use, the mechanisms underlying TXD’s protective effects on DCI have yet to be fully elucidated. As a significant regulator in neurodegenerative conditions, death-associated protein kinase-1 (DAPK-1) serves as a focus for understanding the action of TXD. This study was designed to whether TXD mediates its beneficial outcomes by inhibiting DAPK-1. To this end, a diabetic model was established using Sprague-Dawley (SD) rats through a high-fat, high-sugar (HFHS) diet regimen, followed by streptozotocin (STZ) injection. The experimental cohort was stratified into six groups: Control, Diabetic, TC-DAPK6, high-dose TXD, medium-dose TXD, and low-dose TXD groups. Following a 12-week treatment period, various assessments—including blood glucose levels, body weight measurements, Morris water maze (MWM) testing for cognitive function, brain magnetic resonance imaging (MRI), and histological analyses using hematoxylin-eosin (H&E), and Nissl staining—were conducted. Protein expression in the hippocampus was quantified through Western blotting analysis. The results revealed that TXD significantly improved spatial learning and memory abilities, and preserved hippocampal structure in diabetic rats. Importantly, TXD administration led to a down-regulation of proteins indicative of neurological damage and suppressed DAPK-1 activity within the hippocampal region. These results underscore TXD's potential in mitigating DCIvia DAPK-1 inhibition, positioning it as a viable therapeutic candidate for addressing this condition. Further investigation into TXD's molecular mechanisms may elucidate new pathways for the treatment of DCI.

Background & aims: Little is known about adherence to lifestyle advice from health professionals among people with high blood cholesterol, and its relationship with cardiovascular disease (CVD). We aimed to examine the proportion of adherence to lifestyle advice and its effect on lipid profile and CVD among people with high cholesterol.Methods: Within the National Health and Nutrition Examination Survey 1999-2010, the study included adults aged >= 20 years who were recommended to improve lifestyles in diet (N = 6645), bodyweight (N = 4797), or exercise (N = 5594) due to their high cholesterol. Adherent status was self-reported through questionnaires. Lipid measurements were collected from laboratory tests. 10-year Atheroscle-rotic CVD (ASCVD) risk was estimated by using pooled cohort risk equations for participants aged 40-79 years. CVD mortality up to December 31, 2019 was obtained from the National Death Index.Results: The percentages of adherents were 80.9%, 80.7%, and 72.7% for eating less fat, controlling weight, and increasing exercise, respectively. The percentages of adherents for controlling weight and increasing exercise significantly increased from 1999 to 2000 to 2009-2010 (both P trend <0.05). Adherents had a higher high-density lipoprotein cholesterol (HDL-C) and lower total cholesterol/HDL-C ratio than non -adherents (P for difference <0.05 for all). Participants adhering to increasing exercise had a lower odds ra-tio of 10-year intermediate-to-high ASCVD risk (Odd ratio 0.73, 95% confidence interval [CI] 0.56-0.95) and a lower CVD mortality (Hazard ratio 0.70, 95% CI 0.51-0.97) than non-adherents.Conclusions: Lifestyle advice from health staff obtained relatively high and increasing acceptability. Given the better lipid profile and lower CVD risk of adherents, health professionals should be encouraged to recommend lifestyle modifications for adults with high cholesterol in clinical practice.(c) 2022 The Author(s). Published by Elsevier Ltd on behalf of European Society for Clinical Nutrition and Metabolism. This is an open access article under the CC BY license (http://creativecommons.org/licenses/ by/4.0/).

The AMPK/PGC-1α pathway-mediated mitochondrial dysfunction has been supposed to play a crucial role in pathogenesis of diabetic peripheral neuropathy (DPN). The present study investigated the neuroprotective potential of quercetin, a natural AMPK activator. Streptozotocin (STZ)-induced diabetic rats that developed DPN phenotype were orally administrated with quercetin (30 and 60 mg/kg per day) for 6 weeks. The morphologic changes in the sciatic nerves (SN), the pathological structure of neurons in dorsal root ganglion (DRG), and the expressions of myelin proteins were assessed. The ATP content and the mitochondrial ultrastructure were measured. Furthermore, key proteins in the AMPK/PGC-1α pathway were determined. As a result, quercetin administration at both doses improved the paw withdrawal threshold, nerve conduction velocity, and the pathologic changes in SN and DRG of DPN rats. The expressions of myelin basic protein and myelin protein zero were also increased by quercetin. The oxidative stress, decreased ATP generation, and morphological changes of mitochondria were corrected by quercetin. In vitro study found that quercetin treatment significantly decreased the high-glucose-induced generation of reactive oxygen species, as well as attenuated the mitochondrial morphologic injuries and oxidative DNA damages of RSC96 cells. Quercetin treatment promoted the expressions of phosphorylated AMPK, PGC-1α, SIRT1, NRF1, and TFAM under hyperglycemic state in vivo and in vitro. This study revealed that the neuroprotective effect of quercetin was mainly related to mitochondrial protection by activation of the AMPK/PGC-1α pathway for the first time and proved quercetin as a potential therapeutic agent in the management of diabetic neuropathy.

Abstract Ribes himalense Royle ex Decne. (family Saxifraaceae, subfamily Grossulariaceae, genus Ribes) is a wild berry fruit with illustrated health‐promoting features, which widely distributed in Northwest China are deficiently exploited. This study aimed to assess the potential of a Ribes himalense as a source of natural bioactive compounds through characterizing its nutraceutical characteristics, phytochemicals properties, and antioxidant ability. Fresh berries were quantitatively analyzed for proximate composition, minerals, vitamins, amino acids, total polyphenols, total flavonoids, anthocyanins, procyanidin, and polysaccharides contents through China National Food Safety Standard; the characterization and identification of extracts of wild berries obtained with ethanol 30%, ethanol 50%, and ethanol 95% were firstly performed by UPLC‐Triple‐TOF‐MS2. Furthermore, antioxidant activity of the ethanol extract was evaluated via different assay methods such as DPPH, ABTS, and FRAP. The results indicated that the most important bioactive composition was procyanidin (0.72%), polyphenols (0.49%), total flavonoids (0.38%), vitamin C (64.6 mg/100g FW), and K (218.44 mg/100 g FW), and a total of 95 compounds were detected with polyphenols, flavonoids, and proanthocyanidins as the dominant, and also ethanol extract possessed stronger antioxidant activity. These results suggested that Ribes himalense fruit has great potential in protecting human health, with the focus on the development of functional products.

Jinmaitong (JMT) is a compound prescription of traditional Chinese medicine that has been used to treat diabetic neuropathic pain (DNP) for many years. Here, we investigated the effects of JMT on the activation of NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome and pyroptosis in Dorsal root ganglia (DRG) of diabetic rats. Streptozotocin (STZ)-induced diabetic rats were gavaged with JMT (0.88 g/kg/d) or alpha-lipoic acid (ALA, positive control, 0.48 mmol/kg/d) for 12 weeks. Distilled water was administered as a vehicle control to both diabetic and non-affected control rats. Blood glucose levels and body weights were measured. Behavioral changes were tested with mechanical withdrawal threshold (MWT) and tail-flick latency (TFL) tests. Morphological injury associated with DRG was observed with hematoxylin and eosin (H&E) and Nissl's staining. mRNA and protein levels of NLRP3 inflammasome components (NLRP3, ASC, caspase-1), downstream IL-1β and gasdermin D (GSDMD) were evaluated by immunohistochemistry, quantitative real time-PCR and western blot. The results showed that JMT had no effect on blood glucose levels and body weights, but significantly improved MWT and TFL behavior in diabetic rats, and attenuated morphological damage in the DRG tissues. Importantly, JMT decreased the mRNA and protein levels of components of NLRP3 inflammasome, including NLRP3, ASC and caspase-1. JMT also down-regulated the expression of IL-1β and GSDMD in the DRG of DNP rats. In addition, ALA treatment did not perform better than JMT. In conclusion, JMT effectively relieved DNP by decreasing NLRP3 inflammasome activation and pyroptosis, providing new evidence supporting JMT as an alternative treatment for DNP.

Neuroinflammation contributes significantly to the pathogenesis of diabetic peripheral neuropathy (DPN). Quercetin reportedly exerts neuroprotective effects in DPN. Here, we aimed to evaluate the potential anti-inflammatory effects of quercetin in a DPN rat model. Eight weeks after streptozotocin administration, diabetic rats were treated with quercetin (30 and 60 mg/kg/day orally) for 6 weeks. We assessed the mechanical withdrawal threshold (MWT), nerve conduction velocity (NCV) and morphological changes in sciatic nerves. Additionally, we measured the levels of tumour necrosis factor-alpha (TNF-α), interleukin (IL)-1β, and IL-6 by ELISA and the expression of TLR4, MyD88, and NF-κB in sciatic nerves by western blotting and immunohistochemical assays. Our results revealed that blood glucose levels and body weight were unaltered following quercetin treatment. However, quercetin improved MWT (p < 0.05), NCV (p < 0.05), and pathological changes in the sciatic nerves of DPN rats. Quercetin significantly alleviated the increased expression of TNF-α (p < 0.05) and IL-1β (p < 0.001). Furthermore, high-dose quercetin administration significantly downregulated the expression of TLR4 (p < 0.001), MyD88 (p < 0.001), and NF-κB (p < 0.001) in sciatic nerves of DPN rats. Our findings revealed that quercetin could reduce the levels of inflammatory factors in DPN rats, possibly mediated via the downregulation of the TLR4/MyD88/NF-κB signalling pathway. Collectively, these results suggest that although quercetin did not decreased blood glucose levels or reversed the reduced body weight, it showed anti-inflammatory and neuroprotective effects, which was beneficial for the treatment of DPN.

Ethnopharmacological relevance: The genus Ribes Linn., which belongs to the Grossulariaceae family, contains 160 species distributed mainly in temperate and cold regions of the Northern Hemisphere. There are 59 species in southwest, northwest and northeast China. Some species of Ribes have been used as traditional and local medicines for the treatment of glaucoma, cardiovascular disease, stomachache, hepatitis, hyperlipidemia, hypertension and other ailments. However, the data provided in recent years have not been collated and compared. Aim of the study: This review aims to summarize the current status of ethnopharmacological uses, phytochemistry, pharmacology, clinical applications, and pharmacokinetics of the genus Ribes to better understand the therapeutic potential of the genus Ribes in the future and hope to provide a relatively novel perspective for further clinical application on the genus. Materials and methods: The literature on Ribes was collected through a series of scientific search engines including Elsevier, ACS, Springer, Web of Science, PubMed, Google Scholar, Baidu Scholar, Wiley, China National Knowledge Infrastructure (CNKI) and books. Results: Ribes species have been used for detoxification, glaucoma, cardiovascular disease, stomachache, hepatitis, hyperlipidemia, hypertension and other ailments. These plants mainly contain phenolic glycosides, flavonoids, proanthocyanidins, polysaccharides, etc. Most traditional uses are related to biological activity and have been confirmed by modern research. Pharmacological studies in vitro and in vivo revealed that the extracts and pure compounds possessed significant hypolipidemic, antioxidant, anti-inflammatory, antitumor, antibacterial, and antiviral activity, eyesight protection and other effects. Conclusions: The traditional uses, phytochemistry, pharmacology, pharmacokinetics, and clinical applications described in this article explained that the Ribes species has numerous activities, and these findings will promote further action in the area of mechanism research. However, very few preclinical and clinical studies have focused on the toxicology and pharmacokinetics of crude extracts and pure compounds from the genus Ribes. Moreover, several clinical evidence to support the health benefits of Ribes plants. The development of new medicines based on Ribes species as ingredients may be restricted. The pharmacological activity, clinical efficacy and safety of Ribes species need to be verified by systematic and comprehensive preclinical studies and clinical trials.

Background Traditional Chinese medicine (TCM) has long been used in the treatment of diabetic cognitive impairment (DCI), and Tu-Xian mixture is one of the prescriptions. This study used a network pharmacology method to predict the active ingredients, potential targets and pathways of Tu-Xian mixture in the treatment of DCI.Methods The potential active ingredients of Tu-xian mixture were obtained by querying databases such as TCMSP, drug targets were screened through Pharmmapper, ETCM, and UniProt databases, and disease target protein groups were collected using TTD, OMIM, and CTD databases. Therapeutic targets were obtained by comparative analysis. Then use WebGestalt to complete GO enrichment analysis and KEGG pathway analysis.Results A total of 51 active ingredients and 619 potential targets were obtained, among which there were 33 key targets for diabetes and 18 key targets for cognitive impairment. After GO analysis and KEGG pathway analysis, steroid hormone biosynthesis and γ-aminobutyric acid signaling pathway may be the important metabolic pathways for Tu-Xian mixture to perform therapeutic effects.Conclusions This work promoted the comprehension of Tu-Xian mixture in the treatment of DCI, and further experimental verification of the prediction results was still needed to support its clinical application.

Objective To investigate the association between serum adiponectin levels and diabetic peripheral neuropathy (DPN) in Chinese type 2 diabetes (T2D) patients. Design and Methods Two hundred nineteen T2D patients aged 40–79 years were divided into two groups according to whether they had DPN. The systemic levels of five biomarkers were measured using a human adipokine multiplexed bead-based immunoassay. Diabetic peripheral neuropathy diagnostic criteria included both common DPN symptoms and neurological screening tests. Results Most features of DPN (n=98) and non-DPN patients (n=121) are similar, but the DPN patients were slightly older, had longer diabetes duration, higher hemoglobin (Hb) A1c, lower estimated glomerular filtration rates (eGFR), less exercise, and used lipid-lowering drugs more often. Serum adiponectin levels of DPN patients were higher than that of non-DPN patients (8.13 vs. 9.63 mg/ml, P = 0.004). Serum adiponectin levels were positively associated with DPN after adjusting for age, gender, body mass index, hypertension, HbA1c, alcohol intake, smoking status, physical activity, log-transformed low density lipoprotein cholesterol, lipid-lowering drug usage, eGFR, and diabetes duration {odds ratio (OR) 1.72 [95% confidence interval (CI) 1.02-2.89], P = 0.041}. The OR refers to a doubling in biomarkers. Conclusions Serum adiponectin levels were higher in DPN patients compared to non‑DPN patients in this Chinese T2D population. Serum adiponectin levels were positively associated with DPN presence, independent of multiple confounders.

Objective To investigate the role of thioredoxin interacting protein(TXNIP)/ nucleotides-binding oligomerization domain-like receptor protein(NLRP)3 inflammasome in the sciatic nerve of streptozotocin(STZ)-induced diabetic rats. Methods The diabetic rat model was established by single intraperitoneal injection of STZ.The rats with matched sex and age were taken as normal control group.The blood glucose and body weight were monitored.The mechanical withdrawal threshold was measured by von Frey filaments at 12 weeks after the model was established.At 12 weeks,the rats were sacrificed and the sciatic nerves were separated for Luxol fast blue staining,the expressions of TXNIP,NLRP3,caspase-1,and interleukin(IL)-1β were detected by immunohistochemistry and Western blot method,and the levels of IL-1β and IL-18 in serum were measured by enzyme-linked immunosorbent assay(ELISA). Results The expression of TXNIP protein in the sciatic nerve of diabetic rats was 3.78±0.08,which significantly increased than that in the normal control group(0.99±0.06)(t=26.980,P<0.0001).Compared with the normal control group(0.97±0.05),the expression of NLRP3 protein in the diabetic group(2.44±0.16)was significantly higher(t=8.885,P<0.0001).The expression of cleaved caspase-1 was 4.45±0.19 in the diabetic group and 1.08±0.06 in the normal control group,and the difference was significant(t=16.900,P<0.0001).The expression of IL-1β protein in the diabetic group(4.50±0.16)was significantly higher than that(1.19±0.08)in the normal control group(t=18.630,P<0.0001).Compared with the normal control group,the levels of IL-1β [(110.50±8.80)pg/ml vs.(17.97±3.18)pg/ml,t=9.892,P<0.0001] and IL-18 [(591.70±8.78)pg/ml vs.(160.70±8.33)pg/ml,t=35.620,P<0.0001] in the serum of diabetic rats significantly increased. Conclusion The pathogenesis of diabetic peripheral neuropathy may be related to increased expression of TXNIP,activation of NLRP3 inflammasome,and downstream inflammation,which may provide a new target for diabetic peripheral neuropathy therapy.

Background Jinmaitong (JMT) has been used to prevent and treat diabetic peripheral neuropathy (DPN) for decades. The present study aimed to elucidate the effects of JMT on thioredoxin-interacting protein (TXNIP) and Nod-like receptor protein 3 (NLRP3) inflammasome activation in the streptozotocin (STZ)-induced rat model. Methods The diabetic rat model was induced by a single intraperitoneal injection of 55 mg/kg STZ. The rats were divided into 3 groups (n = 8–10 per group): diabetic control, JMT (0.876 g/kg/d), and alpha-lipoic acid (ALA; 100 mg/kg/d). Body weight and blood glucose levels were monitored every 4 weeks for 12 weeks. Mechanical allodynia and myelin sheath injury of sciatic nerves (SNs) were assessed using the mechanical withdrawal threshold (MWT) test and Luxol fast blue staining. Serum T-superoxide dismutase (T-SOD), malondialdehyde (MDA), and catalase (CAT) levels were measured using commercially available kits. TXNIP/NLRP3 inflammasome proteins, including TXNIP, NLRP3, pro-caspase-1, and cleaved -caspase-1, and the downstream protein interleukin (IL)-1β, were measured using immunohistochemistry and Western blot. Gasdermin D (GSDMDC1) protein expression was analyzed using Western blot, and serum IL-1β and IL-18 levels were detected using ELISA. Results JMT did not significantly affect body weight or level of fasting blood glucose but improved mechanical allodynia and myelin sheath injury of SNs at 12 weeks following treatment. Moreover, JMT increased serum levels of the anti-oxidative enzymes CAT and T-SOD, and decreased MDA levels. Both JMT and ALA decreased expression of TXNIP, NLRP3, and cleaved-caspase-1 protein. JMT and ALA also decreased IL-1β, IL-18, and GSDMDC1 protein expression. Conclusion The current study demonstrated that TXNIP/NLRP3 inflammasome activation is involved in the molecular mechanisms underlying JMT’s protective effects in the STZ-induced diabetic rat model, which provides novel evidence to support the future clinical use of JMT.

Objective To summarize the distribution and clinical characteristics of Chinese med- icine (CM) syndrome types in 660 patients with diabetic peripheral neuropathy (DPN). Methods Totally 660 inpatients at Department of Traditional Chinese Medicine, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences were recruited from Jan 2000 to Dec 2014. Their first diagnoses were DNP. The distributions of their syndrome types were observed. Clinical characteristics in patients with different syndrome types were compared. Meanwhile, Logistic regression analysis was performed in independent variable by taking syndrome types of CM as quartering regression variables. Results The ratio of syndrome types was sequenced from high to low as yin deficiency blood stasis syndrome [39.24% (259/660)], yang deficiency blood stasis syndrome [29.39% (194/60)], phlegm stasis in collaterals syndrome [19. 24% (127/660) ] , yin deficiency induced wind stirring syndrome [12. 12% (80/ 660) ]. There was no significant difference in the constituent ratio of CM syndrome patterns among groups with different courses of diabetes (P >0. 05). The ratio of yang deficiency blood stasis syndrome had an increasing trend as the course increased. There was significant difference in HbAlc, fasting C pep- tide (FCP) , systolic blood pressure (SBP) , total cholesterol (TC) , 24 h total urinary protein (24 h UCP) , serum creatinine (SCr), blood urea nitrogen (BUN) among patient groups with different CM syndrome types (P <0. 05). Compared with yang deficiency blood stasis syndrome, HbAlc increased, SBP,SCr,BUN and 24 hUCP decreased in yin deficiency blood stasis syndrome with statistical difference (P = 0. 006, 0. 002,0. 001 ,0. 001, and 0. 007; P <0. 05) ; 24 h UCP also decreased in yin deficiency induced wind stirring syndrome (P =0. 34, P <0. 05). Multiclassified Logistic regression showed that when taking yin deficiency blood stasis syndrome as reference, HbAlc was a protective factor of yang deficiency blood stasis syn- drome, 8 h urinary albumin excretion (UAE) was a risk factor. Both TC and SCr were risk factors for yin deficiency induced wind stirring syndrome. SCr was a risk factor for phlegm stasis in collaterals syndrome. Conclusions Poor control of blood glucose in DPN patients might be related with yin deficiency blood sta- sis syndrome. Patients with yang deficiency blood stasis syndrome might have longer course of disease, and were correlated with poorer control of SBP and renal function. DPN patients complicated diabetic ne- phropathy were more liable to have yang deficiency blood stasis syndrome.

目的 原代培养胚胎大鼠背根神经节神经元(DRGn),并观察其生长特性.方法 取E15 SD胎鼠的背根神经节,用胰蛋白酶消化法分离成单细胞,通过密度梯度离心法和差速贴壁法进行分离纯化,在无血清培养基中培养,用抗神经元特异性烯醇化酶(NSE)抗体鉴定神经元.结果 纯化培养神经元生长状态良好,纯度可达93％左右.结论 该方法具有较好的可操作性和可重复性,用于DRGn的相关实验研究.

β-cell failure coupled with insulin resistance plays a key role in the development of type 2 diabetes mellitus (T2DM). Changed adipokines in circulating level form a remarkable link between obesity and both β-cell failure and insulin resistance. Some adipokines have beneficial effects,whereas others have detrimental properties. The overall contribution of adipokines to β-cell failure mainly depends on the interactions among adipokines. This article reviews the role of individual adipokines such as leptin,adiponectin,and resistin in the function,proliferation,death,and failure of β-cells. Future studies focusing on the combined effects of adipokines on β-cells failure may provide new insights in the treatment of T2DM.

目的 研究糖尿病周围神经病变(DPN)的中医证候特点及其影响因素.方法 收集695例DPN患者的病历资料,根据统一的辨证标准辨为气虚证、阴虚证、阳虚证、血瘀证、痰湿证、气滞证6个单证,将单一证候组合形成复合证型,利用统计学方法对数据进行分析.结果 血瘀证(78.71％)、阴虚证(63.31％)比例较高,阳虚证比例随病程延长逐渐增加;血瘀证比例随年龄的增长和病程延长而增加,阳虚证、血瘀证均与糖尿病病程具有显著相关性(r =0.122和0.115,P＜0.01);不同病程组证型构成比差异有统计学意义(P＜0.01),随着病程的延长,气虚、阴虚、气阴两虚证比例有降低趋势,阳虚、阴阳两虚证所占比例增加.阳虚证较非阳虚证患者神经传导功能异常率明显增高(P＜0.05);体重指数与痰湿证、糖化血红蛋白与阴虚证、空腹血糖与气虚证均有显著相关性(r=0.181和0.105和0.117,P＜0.05或0.01).结论 探讨糖尿病周围神经病变中医证候特点,可以为DPN的中医辨证提供临床依据.

OBJECTIVE:To examine the mechanism underlying the beneficial role of cinnamaldehyde on oxidative damage and apoptosis in high glucose (HG)-induced dorsal root ganglion (DRG) neurons in vitro.METHODS:HG-treated DRG neurons were developed as an in vitro model of diabetic neuropathy. The neurons were randomly divided into five groups: the control group, the HG group and the HG groups treated with 25, 50 and 100 nmol/L cinnamaldehyde, respectively. Cell viability was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and apoptosis rate was evaluated by the in situ TdT-mediated dUTP nick end labeling (TUNEL) assay. The intracellular level of reactive oxygen species (ROS) was measured with flow cytometry. Expression of nuclear factor-kappa B (NF-κB), inhibitor of κB (IκB), phosphorylated IκB (p-IκB), tumor necrosis factor (TNF)-α, interleukin-6 (IL-6) and caspase-3 were determined by western blotting and real-time quantitative reverse transcription polymerase chain reaction (RT-PCR). Expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and hemeoxygenase-1 (HO-1) were also measured by western blotting.RESULTS:Cinnamaldehyde reduced HG-induced loss of viability, apoptosis and intracellular generation of ROS in the DRG neurons via inhibiting NF-κB activity. The western blot assay results showed that the HG-induced elevated expressions of NF-κB, IκB and p-IκB were remarkably reduced by cinnamaldehyde treatment in a dose-dependent manner (P <0.01). The HG-induced over-expression of NF-κB p65 mRNA was remarkably attenuated after cinnamaldehyde treatment in a dose-dependent manner (P <0.01). However, the expressions of Nrf2 and HO-1 were not upregulated. Treatment with cinnamaldehyde not only attenuated caspase-3 activation and the caspase cleavage cascade in DRG neurons, but also lowered the elevated IL-6, TNF-α, cyclo-oxygenase and inducible nitric oxide synthase levels, indicating a reduction in inflammatory damage.CONCLUSIONS:Cinnamaldehyde protected DRG neurons from the deleterious effects of HG through inactivation of NF-κB pathway but not through activation of Nrf2/HO-1. And thus cinnamaldehyde may have potential application as a treatment for DPN.

Diabetic peripheral neuropathy (DPN) is a common chronic complication of diabetes. Jinmaitong (JMT), a Traditional Chinese Medicine, improves certain symptoms of DPN, such as limb pain and numbness. The aim of the present study was to investigate the effects of JMT on DNA oxidative damage and apoptosis in the sciatic nerve of diabetic rats. The rats were divided into a normal and a diabetic group. Diabetes was induced using streptozotocin (60 mg/kg). The diabetic model (DM) rats received vitamin C (0.05 g/kg/day) or JMT [low-dosage (L), 0.44 g/kg/day; medium-dosage (M), 0.88 g/kg/day or high-dosage (H), 1.75 g/kg/day]. After 16 weeks, the mechanical pain threshold of the rats was evaluated. The expression of 8-hydroxy-deoxyguanosine (8-OHdG), nicotinamide adenine dinucleotide phosphate (NADPH) oxidase p22phox, B-cell lymphoma 2 (Bcl-2), caspase 3 and cleaved-poly(ADP-ribose) polymerase 1 (PARP-1) in the sciatic nerve tissues was measured using the reverse transcription-quantitative polymerase chain reaction, immunohistochemistry and western blotting. JMT had no effect on body weight and fasting blood glucose levels. Following treatment, the rats in the JMT groups had an improved pain threshold compared with the DM controls (JMT-L, 52.9±6.5 g; JMT-M, 74.7±9.3 g; and JMT-H, 61.7±2.0 g vs. DM control, 35.32±12.06 g; all P<0.01), while the threshold in the JMT-M rats was similar to that of normal controls (P>0.05). 8-OHdG and NADPH oxidase p22phox expression was significantly decreased in the three JMT groups compared with that in the DM controls (all P<0.05). Following JMT treatment, Bcl-2 levels were increased, while caspase 3 and cleaved-PARP-1 levels were decreased compared with those in the DM controls (all P<0.01). In conclusion, JMT may reduce DNA oxidative damage to the sciatic nerve in diabetic rats, as well as regulate genes involved in peripheral neuronal cell apoptosis, suggesting that JMT could be used to prevent or treat DPN in diabetic patients.

Diabetes is the most common cause of peripheral neuropathy in the world. Most patients with diabetes have neuropathy1, and the prevalence of neuropathy in patients with prediabetes is rising. Diabetic neuropathy is a major cause of disability and health care costs2,3. Patients with diabetes are prone to peripheral nerve disorders, which may present as pain, numbness, tingling, weakness, and difficulties with balance. Thus far, there is still no approved therapy for treating diabetic neuropathy. Peripheral neuropathy can be attributed to numerous mechanisms, including sorbitol accumulation, oxidative-nitrosative stress, 12/15-lipoxygenase activation, inflammation, the polyol pathway, and poly (adenosine diphosphate [ADP]-ribose) polymerase (PARP) overactivation4,5. Among these mechanisms, oxidative-nitrosative stress, which interacts with many other pathways implicated in diabetic neuropathy, is often considered to be the most critical6. Hyperglycemia can increase levels of reactive oxidative species (ROS) through both enzymatic and nonenzymatic processes. Once absorbed by cells, sugar molecules serve as substrates for cellular respiration pathways (ie, glycolysis and the tricarboxylic acid cycle) that produce reducing equivalents that then drive oxidative phosphorylation in mitochondria, a process that produces ROS. Glucose autoxidation creates free radicals that can damage cellular proteins and mitochondrial DNA7. Cells possess an array of antioxidant defense machinery to prevent or counterbalance damage caused by reactive radicals. A family of enzymes exists that detoxify highly reactive radicals to protect cells against excessive oxidative stress. Highly toxic metabolites are generated from phase I detoxification reactions and must be detoxified quickly by phase II detoxifying enzymes, such as glutathione S-transferase, UDP glucuronyltransferase, heme oxygenase-1 (HO-1), NADPH-quinone oxidoreductase, and microsomal epoxide hydrolase. A build-up of phase I metabolites can wreak havoc on cells. The transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nrf-2) is recognized as a critical factor in the induction of phase II enzymes8. Oxidative stress results in the activation of NF-κB, which triggers the activation of inflammatory signaling pathways7. NF-κB is a rapid-acting pleiotropic transcription factor that regulates a diverse group of genes, including genes that encode inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and proinflammatory cytokines. In experimental diabetic neuropathy models, ROS-activated NF-κB and its oxidant products have been shown to enhance the expressions of iNOS, COX-2, tumor necrosis factor-alpha (TNF-α), and interleukin (IL)-69,10,11. Oxidative stress and inflammation have been shown to interact with each other and to be inseparably linked to diabetic neuropathy. Treatments that are currently prescribed to lessen neuropathic pain have undesirable secondary effects and situational variable efficacy12. Therefore, researchers are searching for better diabetic neuropathy drug options. Flavonoids are an attractive treatment option, given their efficacy in clearing ROS and their ability to protect against neuronal damage13. The flavonoid quercetin is the most potent scavenger of ROS, including superoxide, and reactive nitrogen species14. Furthermore, quercetin reduces low-density lipoprotein oxidation and suppresses iNOS activity, consequently reducing damage from nitric oxide superoxide anions15. The axons of dorsal root ganglion neurons (DRG neurons) are afferents that relay sensory information to the central nervous system. With their exuberant metabolism and sensitivity to high glycemia, DRG neurons are an experimental target of diabetic neuropathy16,17. Even short-term hyperglycemia can produce oxidative damage and apoptosis in DRG neurons17. Inflammatory factors play an important role in DRG neuronal damage, especially in painful neuropathy9,18,19. In previous studies, lower glucose concentrations induced DRG neuron death in culture, whereas total glucose levels above 35 mmol/L produced hyperglycemic insult leading to DRG neuron death20. To produce a hyperglycemic insult, 20 mmol/L additional glucose (yielding a total 45 mmol/L glucose) were added to the media for the period specified in individual experiments. In this study, we evaluated the effects of quercetin (2.5, 5, and 10 mmol/L) on high glucose (HG)-treated DRG neurons, in terms of cell viability, ROS levels, and apoptosis. The expression levels of NF-кB, Nrf2/HO-1, proinflammatory mediators (IL-6 and TNF-α), iNOS, COX-2, and caspase-3 were also evaluated. Dissociated DRG cells were cultured in poly-D-lysine-precoated 96-well or 6-well clusters (Costar, Corning, NY) at 5×104 cells/well in 0.1 mL for 96-well clusters or 2×106 cells/well in 2 mL for 6-well clusters. DRG cells were cultured in media at 37 °C with 5% CO2 and maintained in media containing 20 μmol/L FUDR for another 24 h to inhibit the growth of non-neuronal cells. The purity of neuronal cells was confirmed by fluorescent labeling of microtubule-associated protein 2 (MAP-2, Merck-Millipore) and a 4',6-diamidino-2-phenylindole (DAPI, Zhongshan Golden Bridge Biotechnology Co, Ltd, Beijing, China) fluorescent counterstain (Figure 1). Neurons were cultured for an additional 24 h under different experimental conditions before examination. Neurobasal medium was used as the control group medium because its glucose concentration is 25 mmol/L, which is optimal for the survival and growth of DRG neurons. An additional 20 mmol/L glucose was added to the medium for the HG groups17. The cells were randomly divided into five groups: control (CON, 25 mmol/L glucose), HG (45 mmol/L glucose), and HG plus 2.5 mmol/L quercetin (Q2.5), 5 mmol/L quercetin (Q5), or 10 mmol/L quercetin (Q10). The choice of quercetin concentrations was based on our unpublished results. Based on our preliminary results, ROS in the HG group were elevated from 1 h, peaked at 4 h, and thereafter declined; therefore, 4 h was chosen as the time point for analysis. After 4 h of HG treatment, cells were harvested and treated with 5 μmol/L DCFH-DA at 37 °C for 30 min in the dark. After DCFH-DA treatment, the culture medium was removed, and the cells were washed with 0.1 mmol/L phosphate-buffered saline (PBS, pH 7.4, Invitrogen) three times to remove excess probe. Fluorescence intensity was analyzed by flow cytometry with histogram plots. ROS values were calculated relative to unlabeled control cells. For immunodetection, blots were incubated with primary antibodies overnight at 4 °C and then washed and incubated with horseradish peroxidase (HRP)-conjugated anti-mouse IgG (H+L) (1:5000, Zhongshan Golden Bridge Biotechnology Co, Ltd, Beijing, China) or HRP-conjugated anti-rabbit IgG (H+L) (1:3000, Zhongshan Golden Bridge Biotechnology Co, Ltd, Beijing, China) for 1 h. ECL Super Signal® West Pico Chemiluminescent Substrate (Pierce) was applied to enable visualization of the bands. The relative levels of each protein to β-actin were determined by densitometry analysis with ImageJ software. In this study, we obtained evidence showing that when DRG neurons are exposed to HG, they exhibit decreased cell viability and increased levels of ROS and apoptosis. They also show increased activation of NF-кB and elevated levels of proinflammatory cytokines (IL-6 and TNF-α) and other inflammatory-related enzymes (iNOS and COX-2). Treatment with quercetin attenuated HG-induced apoptosis, and this effect was concomitant with an inhibition of NF-κB, Nrf-2/HO-1, and caspase-3 activity. These effects were associated with the downregulated expressions of IL-6, TNF-α, iNOS, and COX-2. The effects of quercetin on DRG neurons were dose-dependent and detectable at both the protein and mRNA levels. The present study complements prior research showing decreases in the nuclear localization of NF-кB subunits in HG-exposed DRGs being employed as an in vitro analog of peripheral neuropathy. The model is clinically relevant, in that DRGs are nodes of sensory afferent somata, and diabetes-associated numbness and pain in patients can be attributed to dysfunction and apoptosis of DRG neurons16. Under these conditions, the changes in NF-кB may be attributable, at least in part, to impaired mitochondrial ROS scavenging due to a deficiency of manganese superoxide dismutase. Importantly, enhancement of NF-кB activity in cultured HG-exposed DRG neurons, similar to that shown here, has been shown by others to ameliorate suboptimal neurite outgrowth29. Additionally, Chiarugi30 demonstrated that drug-dependent inhibition of neuronal NF-кB activity induced selective activation of caspase-9 and -3 and mitochondrial release of cytochrome c. Given their high susceptibility to hyperglycemia, DRG neurons are an attractive target for diabetic neuropathy treatment. In the last decade, researchers have found that DRG neurons in culture died through programmed cell death when exposed to elevated levels of glucose, and confirmed that ROS are an important component of this induction of cell death17. Subsequent research revealed additional mechanisms that were contributing to this cell death, including respiratory chain dysfunction, mitochondrial DNA damage, and inflammation9,31,32. Advanced glycation end products (AGEs) and oxidized lipoproteins activate inflammatory signaling, which leads to oxidative and nitrosative stress, which in turn aggravates the inflammation4,33. Therefore, clearance of oxidative and nitrosative stress and inhibition of inflammation have been considered good strategies for the treatment of diabetic neuropathy34. Prior studies have shown that quercetin has a good capacity for scavenging ROS and yielding neuroprotection. Quercetin has been shown to inhibit LPS-induced upregulation of both iNOS and COX-2, as well as expression of p38 mitogen-activated protein kinase (p38-MAPK), TNF-α, and NF-кB15,35,36. Additionally, quercetin has been reported to reduce DNA fragmentation, the Bax/Bcl-2 ratio, nuclear translocation of apoptosis-inducing factor, and PARP cleavage37. Moreover, Arredondo et al38 recently found that quercetin could induce Nrf-2 nuclear translocation and increase the γ-glutamylcysteine ligase catalytic subunit gene expression, and these effects were associated with prevention of neuronal death from oxidant exposure. Thus, the present findings combined with prior findings suggest that quercetin works through multiple pathways to protect neurons from oxidative damage. Researchers interested in controlling the production of free radicals are particularly interested in antioxidant enzymes, especially superoxide dismutases, catalases, and glutathione peroxidases. Nrf-2 is referred to as the “master regulator” of the antioxidant response because it modulates the expression of hundreds of genes, including not only antioxidant enzymes, but also numerous genes involved in diverse processes such as immune and inflammatory responses39. Under quiescent conditions, the repressor of Nrf-2, Kelch-like ECH-associated protein 1 (Keap1), binds Nrf-2 and thereby sequesters it in the cytoplasm. When Keap1 encounters ROS, it dissociates from Nrf-2, leaving Nrf-2 less prone to proteasomal degradation and free to translocate to the nucleus, where it interacts with the antioxidant response element to increase the expression of many antioxidant and detoxifying enzymes and proteins (eg, HO-1, NAD(P)H dehydrogenase quinone 1, glutathione S-transferase, and superoxide dismutase). Although brief (<3 h), mild oxidative stress stimulates nuclear transfer of Nrf-2 to produce a protective effect on DRG neurons. Long-term hyperglycemia inhibits the expression of Nrf-2 and depresses the expression of HO-19,40. Our findings suggest that quercetin alleviates the negative long-term effects of HG by upregulating the expressions of Nrf-2 and HO-1. NF-κB can be activated by many factors, including, in addition to hyperglycemia, hypoxia, ischemia, and infection. Exposure to HG can affect NF-κB expression through various pathways, with hyperglycemia-related ROS being a major factor41,42. Hyperglycemia can trigger phosphatidyl inositide-3-kinase/Akt signaling and induce NF-кB-related upregulation of COX-2, which in turn triggers proapoptotic caspase-3 activity43. Other researchers have found that NADPH oxidase-related ROS-induced apoptosis is mediated via JNK-dependent activation of NF-кB in cardiomyocytes exposed to HG44. Additionally, AGE-induced ROS generation also triggers NF-кB activation and subsequent TNF-α production to induce apoptosis45. Treatment with an NF-кB inhibitor has been reported to prevent AGE-induced iNOS expression by interfering with p38 MAPK activity46. In conclusion, the present experiments demonstrated that apoptosis of DRG neurons was increased under HG (diabetes model) conditions, and this effect was attenuated in the presence of quercetin. Our data further suggest that these protective effects of quercetin may be attributable to influences on the Nrf-2/HO-1 pathway, scavenging ROS, and inhibition of NF-кB activation. The possibility that NF-кB may play a critical role in the pathogenesis of neuronal dysfunction due to hyperglycemia suggests that the inhibition of local NF-кB activity may be a beneficial strategy for preventing or alleviating diabetic neuropathy complications. Furthermore, our findings of the protective effects of quercetin on DRG neurons in a hyperglycemic environment point to quercetin as a potential medicine for diabetic neuropathy. Yue SHI, Xiao-chun LIANG, and Hong ZHANG designed the research; Yue SHI and Hong ZHANG performed the research; Qun-li WU, Ling QU, and Qing SUN contributed new reagents and analytic tools; Yue SHI analyzed the data; and Yue SHI wrote the paper. The authors would like to thank the Department of TCM at PUMCH and the Institute of Basic Medical Sciences of Chinese Academy of Medical Sciences, Beijing, China for supporting this research. We thank Medjaden Bioscience Limited for assisting in the preparation of this manuscript.