Comprehensive study of dexamethasone on albumin biogenesis during normal and pathological renal conditions.

CONTEXT: Dexamethasone (DXM) has an anti-immunoinflammatory effect, and is often used in acute kidney injury (AKI). However, the effects of DXM on albumin (ALB) have not been fully studied. OBJECTIVE: To investigate the effects of DXM on ALB production and renal function. MATERIALS AND METHODS: Male Wistar rats were divided into normal and DXM groups (0.25, 0.5, 1 mg/kg for 5 days) (n = 15) for a dose-dependent study. Rats were divided into normal group and DXM groups (0.5 mg/kg for 3, 5, 7 days) (n = 9) for a time-dependent study. In AKI experiment, rats were divided into normal (saline), cisplatin (CP, 5 mg/kg, i.v.), CP + DXM groups (0.25, 0.5 and 1 mg/kg, i.m.) (n = 16). The blood and the organs were isolated for analysis. RESULTS: In normal, serum ALB (sALB) and serum total protein (sTP) increased in DXM group with sALB increased 19.8-32.2% (from small to large dosages); and 30.2-32.5.6% (from 3 to 7 days of DXM); sTP 15.7-22.6% and 14.2-24.3%; urine ALB (uALB) 31.5-392.3%, and 1047.2-1390.8%; urine TP (uTP) 0.68-173.1% and 98.0-504.9%, compared with normal groups. DXM increased the mRNA expression of Cebp and Hnf, suppressing podocin. In AKI, DXM decreased serum BUN (53.7%), serum Cre (73.4%), sALB (30.0%), sTP (18.7%), uALB (74.5%), uTP (449.3%), rescuing the suppressed podocin in kidney. CONCLUSIONS: DXM acts on Cebp and Hnf and promotes ALB production. This finding helps to evaluate the rationale of DXM for kidney injury.

[Study on therapeutic effect and its related mechanism of anemoside B4 on ischemia reperfusion injury induced by renal artery and vein ligation in rats].

The aim of this paper was to investigate the effect and mechanism of anemoside B4 on renal ischemia reperfusion injury in rats. A total of 50 rats were randomly divided into the model group(NS) and anemoside B4 low-dose(1.25 mg·kg~(-1)), medium-dose(2.5 mg·kg~(-1)) and high-dose(5 mg·kg~(-1)) groups after the right kidney was removed and the left kidney was ligated to make the ischemia reperfusion model. Another 10 rats were selected as sham operation group only for normal control group(NS, received normal saline). Automatic biochemical analyzer was used to measure serum blood urea nitrogen(BUN), creatinine(Cre), cerebrospinal fluid(CSF) and urinemicroalbumin(mALB) levels after 5 days of tail vein injection treament. Total urine protein and total urinary albu-min were calculated and kidney samples were collected. Histopathological changes of renal tissues were observed by PAS staining. Western blot analysis was performed to detect the protein expressions of TLR4 and NF-κB in renal inflammatory factors related to NLRP3 pathway and TLR4/NF-κB pathway. The results showed that the levels of BUN, Cre, urinary total protein and urinary total albumin in the model group were significantly increased(P<0.01), with severe renal tubule injury was serious, manifested by obvious expansion of renal tubules, more serious tubular proteins, and some tubular epithelial cells were exfoliated. At the same time, the expression of inflammatory factors related to NLRP3 pathway and TLR4/NF-κB pathway increased significantly(P<0.01 or P<0.05). The levels of BUN, Cre were reduced in different doses of anemoside B4(P<0.05). The levels of total urinary protein and total urinary albumin were decreased in the low and high dose groups of anemoside B4.The level of total urinary albumin in the high-dose group of anemoside B4 was significantly reduced(P<0.05).Renal tubular injury was alleviated, tubular epithelial cell exfoliation was reduced, and the expression of related inflammatory factors was reduced in different degrees(P<0.01 or P<0.05). This study showed that anemoside B4 could alleviate renal ischemia-reperfusion injury in rats. And its mechanism may be related to the inhibition of inflammatory factors related to response mediated by NLRP3 pathway and TLR4/NF-κB pathway by anemoside B4.

Comparison of the TLR4/NFκB and NLRP3 signalling pathways in major organs of the mouse after intravenous injection of lipopolysaccharide.

Context: Lipopolysaccharide (LPS) is often used to induce immunoinflammatory reactions. TLR4/NFκB and NLRP3 signalling are major factors for inflammation. Dexamethasone (DXM) has an anti-immunoinflammatory effect. Objective: To investigate the inflammatory reaction in pathological changes of organs and the expression of inflammatory signalling during LPS infection. Materials and methods: ICR mice were divided into control group (n = 9), LPS group (n = 15) and LPS + DXM group (n = 14). LPS (10 mg/kg) was injected intravenously in LPS group and LPS + DXM group, normal saline was injected to the control group; DXM (0.5 mg/kg) was given by intragastric administration. 12 h after LPS, the blood was collected and the organs were isolated for biochemical analysis, protein expression, and morphological examination. Results: The results showed that BUN, Cre, ALT, AST in the LPS group increased distinctly by 81.42, 67.84, 40.53 and 36.05%, respectively, and CK, ALP, TP and ALB decreased by 71.37, 60.6, 12.57 and 19.73%, respectively, compared with the control group. In the morphologic observation, local necrosis in the liver, arterial vasodilation in the heart and kidney, alveolar secretions and pulmonary interstitial in the lungs, and mucosal shedding in the small and large intestines, the expression of TLR4-NFκB signalling were up-regulated distinctly whereas NLRP3 signalling was less broadly affected. DXM can decrease BUN and Cre, downregulate the expression of TLR4-NFκB signalling, but has no effect on the organ damage based on morphology. Conclusion: Acute injuries induced by LPS are extensive. The inflammatory damage in small and large intestines, liver and kidney was more severe than other organs. TLR4-NFκB signalling was the major response to LPS stress.

[Study on acute kidney injury model induced by renal ischemia-reperfusion in rats].

In this study,in-depth systematic evaluation of rat of acute kidney injury(AKI) caused by renal arteriovenous ligation was conducted to better master and apply this model for drug research. Male SD rats of 2-3 months old were employed in this study.The left kidney was removed,and the right kidney received ligation for 40 min and reperfusion for 24 h. Serum creatinine(Crea),urea nitrogen(BUN) and the renal tissue sections were assayed as the basic indicators to evaluate their renal function. The mRNA expression of inflammatory necrosis factors and apoptotic factors was used to evaluate the mechanism of molecular pathophysiological changes. The results showed that the serum Crea and BUN caused by ligation of both renal arteries and veins were significantly higher than those of rats with renal artery ligation. After renal arteriovenous ligation for 40 min and reperfusion for 24 h in rats,the serum Crea of the rats varied from less than 100 µmol·L-1 to more than 430 µmol·L-1. Among them,5 rats showed less than 100 µmol·L-1 serum Crea,20 rats with 100-200 µmol·L-1 serum Crea and 12 rats with more than 430 µmol·L-1. Rats with serum Crea between 300-430 µmol·L-1 accounted for 66.3%(122/184) of the total number of the experiment rats. After 72 h reperfusion,serum Crea in the group of Crea 370-430 µmol·L-1 continued to increase,while the serum Crea in the group of Crea 200-300 µmol·L-1 and the group of Crea 300-370 µmol·L-1 recovered quickly. No matter serum Crea was elevated or decreased,the renal tubules showed pathological changes such as vacuolar degeneration or even necrosis. The mRNA expression levels of Toll-like receptor(TLR4),tumor necrosis factor(TNF-α) and interleukin(IL-6) in renal tissueswere significantly up-regulated,and the effect was most obvious in the group of serum Crea 370-430 µmol·L-1. The study indicated that the model for AKI caused by renal arteriovenous ligation and reperfusion is easy to operate,and the serum Crea and BUN have the characteristics of continuous increase,beneficial to the observation of drug effects. This acute kidney injury is mainly related to the pathophysiological response of inflammatory necrosis.

Genetic interaction of purinergic P2X7 receptor and ER-α polymorphisms in susceptibility to osteoporosis in Chinese postmenopausal women.

Osteoporosis (OP) is an increasing public health problem worldwide. Genetic factors are considered to be major contributors to the pathogenesis of OP. The aim of this study was to investigate the association of the purinergic P2X7 receptor (P2X7R) and estrogen receptor-α (ER-α) genes with OP risk, and the effect of the possible interaction between the two genes on predisposition to OP in Chinese postmenopausal women. A total of 596 subjects, including 350 OP patients and 246 controls, were recruited in this case-control study. Five functional single-nucleotide polymorphisms (SNPs) in the P2X7R gene (rs2393799, rs7958311, rs1718119, rs2230911, rs3751143) and two ER-α PvuII and XbaI polymorphisms were genotyped and analyzed. Single-gene variant analysis showed that the carriers of the CC genotype of P2X7R rs3751143 revealed an increased OP risk. Haplotype rs1718119G-rs2230911G-rs3751143C also appeared to be a significant 'risk' haplotype with OP. For the ER-α gene, no evidence of significant association of PvuII or XbaI polymorphism with OP risk was found. Moreover, there was a significant gene-gene interaction between P2X7R rs3751143 and ER-α PvuII; the cross-validation consistency was 10/10 and the testing accuracy was 0.5818 (P = 0.0107). A 1.67-fold-increased risk for OP was detected in individuals carrying the genotypes of AC or CC of rs3751143 and Pp or PP of PvuII compared to subjects with AA of rs3751143 and pp of PvuII. Our findings suggest an important association of the P2X7R rs3751143CC genotype and the rs1718119G-rs2230911G-rs3751143C haplotype with an increased OP risk. Also, the P2X7R rs3751143 and ER-α PvuII two-locus interaction confers a significantly high susceptibility to OP in Chinese postmenopausal women.

Purinergic P2X7 receptor functional genetic polymorphisms are associated with the susceptibility to osteoporosis in Chinese postmenopausal women.

Osteoporosis (OP) is a major public health problem worldwide. Genetic factors are considered to be major contributors to the pathogenesis of OP. The purinergic P2X7 receptor (P2X7R) has been shown to play a role in the regulation of osteoblast and osteoclast activity and has been considered as an important candidate gene for OP. A case-control study was performed to investigate the associations of functional single nucleotide polymorphisms (SNPs) in the P2X7R gene (rs2393799, rs7958311, rs1718119, rs2230911, and rs3751143) with susceptibility to OP in 400 Chinese OP patients and 400 controls. Results showed that rs3751143 was associated with OP; in particular, carriers of the C allele and CC/(AC + CC) genotypes were at a higher risk of OP, but no significant association of rs2230911, rs7958311, rs1718119, and rs2393799 with OP risk was observed. Analysis of the haplotypes revealed one haplotype (rs1718119G-rs2230911G-rs3751143C) that appeared to be a significant "risk" haplotype with OP. The rs3751143 polymorphism was associated with osteoclast apoptosis; ATP-induced caspase-1 activity of osteoclasts with AC and CC genotypes is lower than that of osteoclasts with AA genotype in vitro. The findings suggest that the P2X7R rs3751143 functional polymorphism might contribute to OP susceptibility in Chinese postmenopausal women.

LncRNA NONRATT021972 siRNA attenuates P2X7 receptor expression and inflammatory cytokine production induced by combined high glucose and free fatty acids in PC12 cells.

Diabetic neuropathy (DNP) is a frequent chronic complication of diabetes mellitus with potentially life-threatening outcomes. High glucose and elevated free fatty acids (FFAs) have been recently recognized as major causes of nervous system damage in diabetes. Our previous study has indicated extracellular stimuli, such as high glucose and/or FFA stress, may activate the p38 mitogen-activated protein kinase (MAPK) signaling pathway and induce a p38 MAPK-dependent sensitization of the P2X7 receptor and release of inflammatory factors in PC12 cells, while the mechanisms underlying remain to be elucidated. Long noncoding RNAs (lncRNAs) play important roles in diverse biological processes, including activation of a series of pathway signalings. Here, we showed combined high D-glucose and FFAs (HGHF) induced an increment of lncRNA-NONRATT021972 (NONCODE ID, nc021972) in PC12 cells. Nc021972 small interference RNA (siRNA) alleviated HGHF-induced activation of p38 MAPK, expression of the P2X7 receptor, and [Ca(2+)]i increment upon P2X7 receptor activation. Further experiments showed that there existed a crosstalk between nc021972 and the p38 MAPK signaling pathway. Inhibition of p38 MAPK signaling decreased nc021972-induced expression of the P2X7 receptor and [Ca(2+)]i increment upon P2X7 receptor activation. Also, nc021972 siRNA inhibited HGHF-induced PC12 release of TNF-α and IL-6 and rescued decreased cell viability mediated by the P2X7 receptor. Therefore, inhibition of nc021972 may serve as a novel therapeutic strategy for diabetes complicated with nervous inflammatory diseases.

Genetic association analyses of fast plasma glucose level in pre-menopausal Chinese women: potential interaction between osteocalcin and oestrogen receptor α.

BACKGROUND: Fasting plasma glucose (FPG) levels are usually tightly regulated within a narrow physiologic range. Variation of FPG levels is clinically important and is strongly heritable. Several lines of evidence suggest the importance of the oestrogen receptor α (ER-α) and osteocalcin (also known as BGP, for bone Gla protein) in determining FPG; however, whether their polymorphisms are associated with FPG variation is not well understood. AIM: To investigate whether ER-a PvuII and BGP HindIII genetic polymorphisms and their potential interaction are associated with FPG variation. SUBJECTS AND METHODS: The study subjects were 328 unrelated pre-menopausal Chinese women aged 21 years and over (mean age ± SD, 33.2 ± 5.9 years), with an average FPG of 4.92 (SD = 0.81). All subjects were genotyped at the ER-α PvuII and BGP HindIII loci using polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP). RESULTS: The ER-α PvuII genotypes were significantly associated with FPG (p = 0.007). In addition, a significant interaction was observed of the ER-α PvuII polymorphism with BGP HindIII polymorphism on FPG variation (p = 0.013), although the BGP HindIII polymorphism was not shown to be individually associated with FPG. CONCLUSION: The PvuII polymorphism of the ER-α gene and its potential interaction with the HindIII polymorphism of the BGP gene were associated with FPG in pre-menopausal Chinese women.

Anemoside B4 attenuates abdominal aortic aneurysm by limiting smooth muscle cell transdifferentiation and its mediated inflammation.

Abdominal aortic aneurysm (AAA) is a degenerative disease characterized by local abnormal dilation of the aorta accompanied by vascular smooth muscle cell (VSMC) dysfunction and chronic inflammation. VSMC dedifferentiation, transdifferentiation, and increased expression of matrix metalloproteinases (MMPs) are essential causes of AAA formation. Previous studies from us and others have shown that Anemoside B4 (AB4), a saponin from Pulsatilla chinensis, has anti-inflammatory, anti-tumor, and regulatory effects on VSMC dedifferentiation. The current study aimed to investigate whether AB4 inhibits AAA development and its underlying mechanisms. By using an Ang II induced AAA model in vivo and cholesterol loading mediated VSMC to macrophage transdifferentiation model in vitro, our study demonstrated that AB4 could attenuate AAA pathogenesis, prevent VSMC dedifferentiation and transdifferentiation to macrophage-like cells, decrease vascular inflammation, and suppress MMP expression and activity. Furthermore, KLF4 overexpression attenuated the effects of AB4 on VSMC to macrophage-like cell transition and VSMC inflammation in vitro. In conclusion, AB4 protects against AAA formation in mice by inhibiting KLF4 mediated VSMC transdifferentiation and inflammation. Our study provides the first proof of concept of using AB4 for AAA management.

Bioinformatic Analysis Revealed the Essential Regulatory Genes and Pathways of Early and Advanced Atherosclerotic Plaque in Humans.

Atherosclerosis (AS) is a lipid-induced, chronic inflammatory, autoimmune disease affecting multiple arteries. Although much effort has been put into AS research in the past decades, it is still the leading cause of death worldwide. The complex genetic network regulation underlying the pathogenesis of AS still needs further investigation to provide effective targeted therapy for AS. We performed a bioinformatic microarray data analysis at different atherosclerotic plaque stages from the Gene Expression Omnibus database with accession numbers GSE43292 and GSE28829. Using gene set enrichment analysis, we further confirmed the immune-related pathways that play an important role in the development of AS. We are reporting, for the first time, that the metabolism of the three branched-chain amino acids (BCAAs; leucine, isoleucine, and valine) and short-chain fatty acids (SCFA; propanoate, and butanoate) are involved in the progression of AS using microarray data of atherosclerotic plaque tissue. Immune and muscle system-related pathways were further confirmed as highly regulated pathways during the development of AS using gene expression pattern analysis. Furthermore, we also identified four modules mainly involved in histone modification, immune-related processes, macroautophagy, and B cell activation with modular differential connectivity in the dataset of GSE43292, and three modules related to immune-related processes, B cell activation, and nuclear division in the dataset of GSE28829 also display modular differential connectivity based on the weighted gene co-expression network analysis. Finally, we identified eight key genes related to the pathways of immune and muscle system function as potential therapeutic biomarkers to distinguish patients with early or advanced stages in AS, and two of the eight genes were validated using the gene expression dataset from gene-deficient mice. The results of the current study will improve our understanding of the molecular mechanisms in the progression of AS. The key genes and pathways identified could be potential biomarkers or new drug targets for AS management.

Anemoside B4 Inhibits Vascular Smooth Muscle Cell Proliferation, Migration, and Neointimal Hyperplasia.

Vascular smooth muscle cell (VSMC) phenotypic transformation, proliferation, and migration play a pivotal role in developing neointimal hyperplasia after vascular injury, including percutaneous transluminal angioplasty and other cardiovascular interventions. Anemoside B4 (B4) is a unique saponin identified from the Pulsatilla chinensis (Bge.) Regel, which has known anti-inflammatory activities. However, its role in modulating VSMC functions and neointima formation has not been evaluated. Herein, we demonstrate that B4 administration had a potent therapeutic effect in reducing neointima formation in a preclinical mouse femoral artery endothelium denudation model. Bromodeoxyuridine incorporation study showed that B4 attenuated neointimal VSMC proliferation in vivo. Consistent with the in vivo findings, B4 attenuated PDGF-BB-induced mouse VSMC proliferation and migration in vitro. Moreover, quantitative RT-PCR and Western blot analysis demonstrated that B4 suppressed PDGF-BB-induced reduction of SM22α, SMA, and Calponin, suggesting that B4 inhibited the transformation of VSMCs from contractile to the synthetic phenotype. Mechanistically, our data showed B4 dose-dependently inhibited the activation of the phosphatidylinositol 3-kinase (PI3K)/AKT and p38 mitogen-activated protein kinase MAPK signaling pathways. Subsequently, we determined that B4 attenuated VSMC proliferation and migration in a p38 MAPK and AKT dependent manner using pharmacological inhibitors. Taken together, this study identified, for the first time, Anemoside B4 as a potential therapeutic agent in regulating VSMC plasticity and combating restenosis after the vascular intervention.

Tiliroside Ameliorates Ulcerative Colitis by Restoring the M1/M2 Macrophage Balance via the HIF-1α/glycolysis Pathway.

Macrophages polarized to different phenotypes critically contribute to colitis development by coordinating inflammatory and anti-inflammatory processes. Herein, targeting the balance between the pro-inflammatory M1 and the anti-inflammatory M2 macrophage phenotypes can be a novel therapeutic approach for colitis. In the present study, we firstly demonstrated that tiliroside possessed the ability to alleviate the clinical symptoms of colitis as evidenced by decreased disease activity index (DAI) scores, longer colon length, reduced myeloperoxidase (MPO) activity, and improvement of colonic pathological damage in vivo. Furthermore, we showed that tiliroside modulated the balance between M1 and M2 macrophages toward a more anti-inflammatory status in colonic lamina propria but has little effect on the T cell population and epithelial barrier function in colitis mice. The macrophage depletion study further showed the protective effect of tiliroside was macrophage dependent in vivo. Mechanistically, our study demonstrated that tiliroside regulated cellular metabolism by inhibiting aerobic glycolysis in LPS and IFNγ stimulated macrophages. At the molecular level, tiliroside facilitated the proteasomal degradation of HIF-1α and downregulated mRNA expressions of HIF-1α dependent glycolytic enzymes in macrophages. Collectively, our data highlight the aberrant M1/M2 macrophage polarization in the initiation and development of ulcerative colitis and put forth the stage for considering tiliroside as a metabolic regulator in reprogramming macrophage polarization, which may serve as a promising therapeutic approach for treatment of inflammation-associated and metabolic disorders.

Proteomic profiling of the neurons in mice with depressive-like behavior induced by corticosterone and the regulation of berberine: pivotal sites of oxidative phosphorylation.

Chronic corticosterone (CORT) stress is an anxiety and depression inducing factor that involves the dysfunction of glucocorticoid receptor (GR), brain-derived neurotrophic factor (BDNF), and neuronal plasticity. However, the regulation of proteomic profiles in neurons suffering CORT stress is remaining elusive. Thus, the proteomic profiles of mouse neuronal C17.2 stem cells were comprehensively investigated by TMT (tandem mass tag)-labeling quantitative proteomics. The quantitative proteomics conjugated gene ontology analysis revealed the inhibitory effect of CORT on the expression of mitochondrial oxidative phosphorylation-related proteins, which can be antagonized by berberine (BBR) treatment. In addition, animal studies showed that changes in mitochondria by CORT can affect neuropsychiatric activities and disturb the physiological functions of neurons via disordering mitochondrial oxidative phosphorylation. Thus, the mitochondrial energy metabolism can be considered as one of the major mechanism underlying CORT-mediated depression. Since CORT is important for depression after traumatic stress disorder, our study will shed light on the prevention and treatment of depression as well as posttraumatic stress disorder (PTSD).

Anemoside B4 Protects Rat Kidney from Adenine-Induced Injury by Attenuating Inflammation and Fibrosis and Enhancing Podocin and Nephrin Expression.

Anemoside B4 (B4) isolated from Radix Pulsatilla has anti-inflammatory activities in the colon and antitumor effects. However, its role in the prevention and treatment of kidney injury has not been reported. Here, we reported the effects of B4 on chronic kidney injury (CKI) and studied its related mechanism based on an adenine-induced kidney injury model in rats. The results showed that serum BUN (blood urea nitrogen), Crea (creatinine), and urinary proteins increased significantly after oral administration of adenine. Meanwhile, the adenine contents in both renal tissue and urine increased markedly compared with those of normal rats. Moreover, IL-1ß, IL-6, TNFα, and NFκB expression was upregulated in the kidney. Simultaneously, the expression of NLRP3 (the nucleotide-binding and oligomerization domain-like receptor, leucine-rich repeat and pyrin domain-containing 3) in the inflammasome, which consists of Caspase 1, ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain), and IL-18, was significantly upregulated. B4 could significantly decrease BUN and Crea; reduce urinary proteins in rats; suppress the expression of IL-6, IL-1ß, NFκB, NLRP3, Caspase 1, ASC, and IL-18; and increase urinary adenine contents and promote its excretion. In addition, B4 also upregulated the expression of podocin and nephrin, two major podocyte proteins, and reduced the fiber collagen in the renal interstitial, suggesting that B4 could protect the glomerular matrix from adenine injury in addition to its anti-inflammatory effects. The results of this study show new perspective of B4 as a potential drug against adenine-induced renal injury.

LncRNA NONRATT021972 siRNA rescued decreased heart rate variability in diabetic rats in superior cervical ganglia.

Diabetic cardiac autonomic neuropathy (DCAN) is a serious and common complication in diabetes mellitus (DM). Long noncoding RNAs (lncRNAs), an important class of regulatory molecules in diverse biological processes, have attracted considerable interest in DCAN. Our previous study has indicated a lncRNA, NONRATT021972 (NONCODE ID), was enhanced in sympathetic neuronal-like PC12 cells in the setting of high glucose (HG) and high FFAs (HF); its silence was found to significantly alleviate HGHF-induced tumor necrosis factor-α (TNF-α) release in PC12 cells. Here we further explore the effects of NONRATT021972 small interference RNA (siRNA) on heart rate variability (HRV) mediated by superior cervical ganglia (SCG) in diabetic rats and the possible mechanism underlying. We found an increment of NONRATT021972 in SCG of DM rats. Treatment of NONRATT021972 siRNA in DM rats decreased the elevated expression of TNF-α, blocked serine phosphorylation of insulin receptor substrate (IRS) 1 and increased the down-regulated expression of IRS1 in SCG. Meanwhile, NONRATT021972 siRNA rescued decreased HRV in DM rats. Therefore, inhibition of NONRATT021972 may serve as a novel therapeutic strategy for preventing the development of DCAN.

Trans-resveratrol attenuates high fatty acid-induced P2X7 receptor expression and IL-6 release in PC12 cells: possible role of P38 MAPK pathway.

Diabetic neuropathy (DNP) is the most common chronic complication of diabetes. Elevated free fatty acids (FFAs) have been recently recognized as major causes of inflammation and are relevant to the functional changes of nerve system in diabetes. Trans-resveratrol (RESV), a polyphenolic natural compound, has long been acknowledged to have anti-inflammation properties and may exert a neuroprotective effect on neuronal damage in diabetes, while the mechanisms underlying are largely unknown. Our previous study on differential PC12 cells cultured with high FFAs has shown chronic FFAs overload increased PC12 interleukin (IL)-6 release mediated by P2X7 receptor, a ligand-gated cation channel activated by extracellular adenosine triphosphate (ATP); a high FFA-induced activation of P38 mitogen-activated protein kinase (MAPK) pathway was pointed to be a potential underlying mechanism. Data from this study indicated that RESV, in a dose-dependent manner, reduced high FFA-induced IL-6 release by impeding the activation of P2X7 receptor, as shown by the results that both high FFA-elevated P2X7 receptor messenger RNA (mRNA) and protein expression as well as high FFA-evoked [Ca(2+)]i in response to 3'-O-(4-benzoyl) benzoyl-ATP (a selective P2X7 receptor agonist) were significantly attenuated. Meanwhile, high FFA-induced activation of P38 MAPK, an essential prerequisite for high FFA-activated P2X7 receptor and subsequent IL-6 release, was also dose-dependently abrogated by RESV. Furthermore, RESV may hamper the activation of P38a MAPK (one paramount P38 isoform) via forming hydrogen bonding with Thr175 residue, surrounding the two residues (Thy180 and Tyr182) essential for canonical activation of P38a MAPK. Taken together, RESV could inhibit high FFA-induced inflammatory IL-6 release mediated by P2X7 receptor through deactivation of P38 MAPK signaling pathway. All these results outline the potential mechanisms involved in the neuroprotective roles of RESV and highlight the clinical application of RESV in treatment of inflammation in relation to DNP.