α-硫辛酸对2型糖尿病患者尿podocalyxin及MCP-1排泄的影响

α-硫辛酸（alpha—lipoic acid，ALA）是一种存在于线粒体的酵素，通过清除氧自由基、还原体内抗氧化系统、螯合金属离子等发挥强抗氧化作用。临床上已被广泛用于治疗糖尿病相关并发症，但有关其对糖尿病肾脏足细胞的保护和抗炎作用的研究尚少。糖尿病肾病（diabetic nephropathy，DN）发病机制复杂，现研究认为炎症反应及足细胞损伤参与了DN发生发展的全过程。

Nucleotide-binding Oligomerization Domain-1 Ligand Induces Inflammation and Attenuates Glucose Uptake in Human Adipocytes

Objective To investigate the effects of stimulant for nucleotide-binding oligomerization domain 1 （NOD1） on secretion of proinflammatory chemokine/cytokines and insulin-dependent glucose uptake in human differentiated adipocytes. Methods Adipose tissues were obtained from patients undergoing liposuction. Stromal vascular cells were extracted and differentiated into adipocytes. A specific ligand for NOD1, was administered to human adipocytes in culture. Nuclear factor-κB transcriptional activity and proinflammatory chemokine/cytokines production were determined by reporter plasmid assay and enzyme-linked immunosorbent assay, respectively. Insulin-stimulated glucose uptake was measured by 2-deoxy-D-[ 3 H] glucose uptake assay. Furthermore, chemokine/cytokine secretion and glucose uptake in adipocytes transfected with small interfering RNA （siRNA） targeting NOD1 upon stimulation of NOD1 ligand were analyzed. Results Nuclear factor-κB transcriptional activity and monocyte chemoattractant protein-1 （MCP-1）, interleukin （IL）-6, and IL-8 secretion in human adipocytes were markedly increased stimulated with NOD1 ligand （all P〈0.01）. Insulin-induced glucose uptake was decreased upon the activation of NOD1 （P〈0.05）. NOD1 gene silencing by siRNA reduced NOD1 ligand-induced MCP-1, IL-6, and IL-8 release and increased insulin-induced glucose uptake （all P〈0.05）. Conclusion NOD1 activation in adipocytes might be implicated in the onset of insulin resistance.

Effect of monocyte chemoattractant protein-1 on chemotactic gene expression by macrophage cell line U937

Objective: To study the chemotactic superfamily genes expression profiling of macrophage line U937 treated with monocyte chemoattractant protein-1 (MCP-1) using gene chip technique. Methods: Total RNA from macrophage line U937 (as control) and U937 with MCP-1 was extracted, made reverse transcript to cDNA and tested with gene expression chip HO2 human. Results: Some chemotactic-related gene expressions were changed in all analyzed genes. Regulated upon activation, normal T cell expressed and secreted (RANTES) was up-regulated over 2-fold and 7 chemotactic-related genes (CCR2, CCR5, CCL16, GROβ, GROγ, IL-8 and granulocyte chemotactic protein 2) were down-regulated over 2-fold in MCP-1 treated U937 cells at mRNA level. Conclusion: MCP-1 can influence some chemokines and receptors expression in macrophage in vitro, in which MCP-1 mainly down-regulates the chemotactic genes expression of those influencing neutrophilic granulocyte (GROβ, GROγ, IL-8 and granulocyte chemotactic protein 2). Another novel finding is that it can also down-regulate the mRNA level of CCR5, which plays a critical role in many disorders and illnesses.

Acute pancreatitis in aging animals:Loss of pancreatitis-associated protein protection?

AIM:To investigate the effect of age on severity of acute pancreatitis(AP) using biochemical markers,histology and expression of the protective pancreatitisassociated proteins(PAPs).METHODS:AP was induced via intraductal injection of 4% sodium taurocholate in young and old rats.Sera and pancreata were assayed at 24 h for the parameters listed above;we also employed a novel molecular technique to assess bacterial infiltration using polymerase chain reaction to measure bacterial genomic ribosomal RNA.RESULTS:At 24 h after induction of AP,the pancreata of older animals had less edema(mean ± SE histologic score of young vs old:3.11 ± 0.16 vs 2.50 ±-0.11,P < 0.05),decreased local inflammatory response(histologic score of stromal infiltrate:3.11 ± 0.27 vs 2.00 ± 0.17,P < 0.05) and increased bacterial infiltration(174% ± 52% increase from sham vs 377% ± 4%,P < 0.05).A decreased expression of PAP1 and PAP2 was demonstrated by Western blotting analysis and immunohistochemical staining.There were no differences in serum amylase and lipase activity,or tissue myeloperoxidase or monocyte chemotactic protein-1 levels.However,in the most-aged group,serum C-reactive protein levels were higher(young vs old:0.249 ± 0.04 mg/dL vs 2.45 ± 0.68 mg/dL,P < 0.05).CONCLUSION:In older animals,there is depressed PAP expression related to a blunted inflammatory response in AP which is associated with worsened bacterial infiltration and higher C-reactive protein level;this may explain the more aggressive clinical course.

Expression characteristics of neutrophil and mononuclear-phagocyte related genes mRNA in the stable angina pectoris and acute myocardial infarction stages of coronary artery disease

Objective To investigate expression differences of neutrophil and mononuclear phagocyte related gene mRNAs among acute myocardial infarction （AMI）, stable angina （SA） and control groups, and then discuss their expression characteristics in the stable angina pectoris （SAP） and AMI stages of coronary artery disease （CAD）. Methods Whole Human Genome Oligo Microarrays were applied to assess the differential expression characteristics of neutrophil and mononuclear phagocyte related mRNAs in patients with AMI （n = 20）, SA （n = 20） and controls （n = 20）. Results （1） Almost all colony-stimulating factors （CSF） and their receptors related mRNAs was up-regulated in AMI and SA groups compared with the control group, and the expression of granulocyte-macrophage colony stimulating factor receptor （GM-CSFR） and granulocyte colony stimulating factor receptor （G-CSFR） mRNAs in the AMI group was significantly up-regulated compared with the other two groups （P 〈 0.01）. （2） The expression of mRNAs related to monocyte chemoattractant protein-1 （MCP-1）, CCR2 （MCP-1 receptor） and CXCR2 （IL-8 receptor） was significantly up-regulated （P 〈 0.01） in AMI group compared with SA and control groups IL-8 mRNA expression in the AMI group was clearly higher than the controls （P 〈 0.05）. （3） All mRNAs expression related to opsonic re- ceptors （IgG FoR and C3bR/C4bR） was significantly up-regulated in AMI group compared with SA and control group （P 〈 0.01）, and the SA group showed an upward trend compared with controls. （4） Most pattern recognition receptor （PRR）-related mRNAs expression was up-regulated in AMI group compared with SA and control groups. Most toll-like receptor （TLR） mRNAs expression was significantly up-regulated （P 〈 0.01） than the SA and control groups, macrophage scavenger receptor （MSR） mRNA was significantly up-regulated in AMI group compared with the control group （P 〈 0.01）, and the SA group showed an upward trend compared with the controls. Conclusions The expression of most neutrophil and mononuclear-macrophage function related genes mRNAs was significantly up-regulated by stages during the progression of CAD, suggesting that the adhesive, chemotactic and phagocytic functions of neutrophil and mononudear-macrophage were strengthened in the occurrence and development of coronary atherosclerosis and AMI. This also showed a stepped up- ward trend as the disease progressed.

Agmatine induces gastric protection against ischemic injury by reducing vascular permeability in rats

AIM:To investigate the effect of administration of agmatine(AGM) on gastric protection against ischemia reperfusion(I/R) injury.METHODS:Three groups of rats(6/group);sham,gastric I/R injury,and gastric I/R + AGM(100 mg/kg,i.p.given 15 min prior to gastric ischemia) were recruited.Gastric injury was conducted by ligating celiac artery for 30 min and reperfusion for another 30 min.Gastric tissues were histologically studied and immunostained with angiopoietin 1(Ang-1) and Ang-2.Vascular endothelial growth factor(VEGF) and monocyte chemoattractant protein-1(MCP-1) were measured in gastric tissue homogenate.To assess whether AKt/phosphatidyl inositol-3-kinase(PI3K) mediated the effect of AGM,an additional group was pretreated with Wortmannin(WM)(inhibitor of Akt/PI3K,15 μg/kg,i.p.),prior to ischemic injury and AGM treatment,and examined histologically and immunostained.Another set of experiments was run to study vascular permeability of the stomach using Evan's blue dye.RESULTS:AGM markedly reduced Evan's blue dye extravasation(3.58 ± 0.975 μg/stomach vs 1.175 ± 0.374 μg/stomach,P < 0.05),VEGF(36.87 ± 2.71 pg/100 mg protein vs 48.4 ± 6.53 pg/100 mg protein,P < 0.05) and MCP-1 tissue level(29.5 ± 7 pg/100 mg protein vs 41.17 ± 10.4 pg/100 mg protein,P < 0.01).It preserved gastric histology and reduced congestion.Ang-1 and Ang-2 immunostaining were reduced in stomach sections of AGM-treated animals.The administration of WM abolished the protective effects of AGM and extensive hemorrhage and ulcerations were seen.CONCLUSION:AGM protects the stomach against I/R injury by reducing vascular permeability and inflammation.This protection is possibly mediated by Akt/PI3K.

NANOPARTICLE AS A NEW GENE TRANSFERRING VECTOR IN SPECIFIC EXPRESSION GENE

Objective. To evaluate the possibility and efficiency of nanoparticle as a new vector in specific gene transference.Methods. Nanoparticle-DNA complex was prepared with Poly- dl-lactic-co-glycolic acid (PLGA) bearing anti-sense monocyte chemotactic protein-1 (A-MCP-l), a specific expression gene, and the package efficiency, release progress in vitro, and the size of the complex were determined. The possibility of the new vector was evaluated with genomic DNA PCR by transferring gene into cultured smooth muscle cells (SMC), cationic lipids as a control. For study in vivo, jugular vein-to-artery bypass grafting procedures were performed on 20 New Zealand white rabbits, of which 6 grafts were transferred with nanoparticle-A-MCP-1 (200 μg), 6 with A - MCP-1 (200 μ g) by cationic liposome, 4 with LNCX plasmid, and 4 as control. Fourteen days after the grafts were harvested, the expression of A-MCP-l and its effect on MCP-1 in vein grafts were detected by dot blot, and the morphologic evaluation of grafts was performed.Results. The package efficiency of the nanoparticle-DNA complex was 0. 9% , release progress in vitro lasted 2 weeks, and the size ranged from 150 to 300nm. SMC genomic DNA PCR showed that A-MCP-l gene could be successfully transfected into cells by nanoparticle. The study in vivo indicated that A-MCP-l mRNA was expressed in both local gene delivery groups, nanoparticle and liposome, meanwhile, MCP-1 expression in vein grafts was significantly inhibited and neointimal hyperplasia was notably reduced.Conclusion. Nanoparticle can act as a vector to transfect specific gene.

Effects of Wenxiao Ⅱ Decoction on the expression of MCP-1 and VCAM-1 in atherosclerotic rabbits

OBJECTIVE:To observe the effects of different doses of Wenxiao Ⅱ Decoction on the expression of monocyte chemoattractant protein-1(MCP-1) and vascular cell adhesion molecule-1(VCAM-1) in an experimental model of atherosclerosis in rabbits and to explore the mechanism by which it alleviates atherosclerosis.METHODS:Sixty 3-4 month-old New Zealand rabbits of both sexes were randomly divided into six groups:simvastain;model;blank;and high-dose,mid-dose,and low-dose Wenxiao Ⅱ Decoction groups.Except for those in the blank group,all rabbits were fed a high-cholesterol diet.Carotid atherosclerosis was established by balloon-induced injury to the endothelium of the carotid artery in conjunction with consumption of a high-cholesterol diet.After 8 weeks,all rabbits were killed to evaluate the expression of MCP-1 and VCAM-1 by immunohistochemical staining.RESULTS:Expressions of MCP-1 and VCAM-1 were significantly decreased in all groups except the blank group compared with the model group(P<0.05).When compared with the simvastain group only variation of MCP-1 expression in low-dose group was not appreciable,and the differences were indistinct(P<0.05).When comparing among Wenxiao Ⅱ Decoction groups,MCP-1 expression in the mid-and high-dose groups was significantly lower than that seen in the low-dose group(P<0.01),but there were no differences among three dosage groups with respect to VCAM-1 expression(P>0.05).CONCLUSION:These data suggested that high,mid,and low doses of Wenxiao Ⅱ Decoction can inhibit the expression of MCP-1 and VCAM-1,which may prevent the formation of or stabilize atherosclerotic plaques.There may be a direct relationship between the dosage of Wenxiao Ⅱ Decoction and its therapeutic efficacy.

Differential gene expression profile of Buyanghuanwu decoction in rats with ventricular remodeling post-myocardial infarction

OBJECTIVE: To investigate the effect of Buyanghuanwu decoction(BYHWD) on gene expression in ventricular remodeling post-myocardial infarction in rats.METHODS: Animal models of myocardial infarction were established by permanent ligation of the left anterior descending coronary artery. Echocardiography measurements were performed after the treatment of BYHWD(18 g·kg-1 collagen was observ·d-1) for 90 days.Myocardialed by mallory trichrome staining. Capillary density was quantified by using Factor rentially expⅧre immunohistochemical staining.Diffessed genes were explored by a short-read sequencing technology combined with a tag-based digital gene expression profiling(DGE)system. Real-time quantitative polymerase chain reaction detecting system(q PCR) was used to validate the sequencing results. After assembling the gene information from Sham, model and BYHWD groups, we constructed three DGE libraries based on each group. The sequencing of three libraries generated 66 000-73 000 unique tags, which were mapped to reference sequences for annotation of expressed genes.RESULTS: Among them, 511 and 352 differentially expressed genes were found in comparison with sham/model and model/BYHWD, respectively. Fifty-five genes exhibited reversed direction of gene expression differences between Sham/Model and Model/BYHWD groups. We found that transforming growth factor beta receptor-1, junctophilin-2,monocyte chemotactic protein 1, neuropeptide Y,arachidonate 5-Lipoxygenase, arachidonate 15-Lipoxygenase were significantly modulated, which suggested the involvement of these genes in BYHWD treatment.CONCLUSION: The DGE profiling data provide comprehensive gene expression information at the transcriptional level that could facilitate our understanding of the pharmacological mechanisms of BYHWD in ventricular remodeling post-myocardial infarction.