Changes in Gelatinase Activity in the Gastrointestinal Tract After Anastomotic Construction in the Ileum or Colon

PURPOSE The strength of the uninjured and anastomosed intestinal wall is determined by its submucosal connective tissue. Matrix degradation by matrix metalloproteinases may result in loss of strength. It is known that anastomotic construction leads to up-regulation of matrix metalloproteinase activity in the wound area, but no quantitative data are available as to the extent of this effect throughout the intestinal wall. This study was designed to quantitate changes in gelatinolytic activity in the intestine after anastomotic construction in the ileum or colon. METHODS An anastomosis was constructed in the distal ileum or distal colon of rats, and animals were killed after one or three days. Tissue samples (5 mm) were collected containing the suture line, its adjacent segments (2- × 5-mm in both directions) and at nine other, more distant, sites throughout the gastrointestinal tract. Similar samples were collected from nonoperated control rats. All samples were analyzed by quantitative gelatin zymography. RESULTS In control rats, the most prominent gelatinolytic activities were found at 80 kDa, thought to represent a nonspecific proteolytic activity, 60 kDa and 50 kDa, representing the proform and active form of matrix metalloproteinase-2, respectively. Activities were higher in the small bowel than in the large bowel. Anastomotic construction led to massive up-regulation of an activity at 105 kDa, and its dimer, believed to represent promatrix metalloproteinase-9. Matrix metalloproteinase-2 remained unaffected, whereas the activity of the 80 kDa protein was significantly (P < 0.05) reduced. Significantly increased matrix metalloproteinase-9 activity was found in the actual anastomotic segments and in the immediately adjacent tissue. Matrix metalloproteinase-9 activities in the anastomotic segments were highest at Day 1 in the ileum and at Day 3 in the colon. Anastomotic construction in the ileum or colon did not lead to any significant changes of any gelatinolytic activity at the more distant sites in the bowel wall. CONCLUSIONS Up-regulation of gelatinase activity after anastomotic construction in the intestine is caused by matrix metalloproteinase-9. Because the effect is local and not systemic, unwanted matrix degradation at distant sites seems unlikely. Supported by the Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands. Reprints are not available.     

Transient Profound Mesenteric Ischemia Strongly Affects the Strength of Intestinal Anastomoses in the Rat

Purpose Experimental data suggest that transient preoperative ischemia and reperfusion may compromise anastomotic strength. However, data on this subject are equivocal, in particular as to the onset and duration of this effect. This study was designed to comprehensively characterize the effects of profound transient intestinal ischemia on anastomotic healing during the first postoperative week. Methods Ischemia was induced in rats by clamping both the superior mesenteric artery and ileal branches for 30 minutes. Immediately after declamping, anastomoses were constructed in both terminal ileum and descending colon. After three, five, or seven days, both bursting pressure and breaking strength were measured. Anastomotic collagen content, gelatinase activity, and histology were analyzed. Results Anastomotic leakage rate was 13 percent in ischemia-reperfusion group and 0 percent (P = 0.02) in controls. The breaking strength in ileum remained significantly (P < 0.05) lower in the ischemic groups than in the control groups at all time points. Bursting pressure in the ileum was not significantly different between ischemic and control groups at either of the time points measured. However, at Day 7 the bursting site was significantly more frequent within the suture line in the ischemic groups. In the colon, at Day 3 the bursting pressure was 35 percent lower in the ischemic group than in the control group (P < 0.05). Anastomotic collagen content and gelatinase activity were similar in ischemic and control groups. Conclusions Transient profound splanchnic ischemia compromises anastomotic strength throughout the entire first postoperative week. This effect does not seem to be caused by impaired accumulation of wound collagen. Presented at the meeting of the European Society for Surgical Research, Konya, Turkey, May 25 to 28, 2005.     

广谱基质金属蛋白酶抑制剂对高血压性脑卒中影响的实验研究

目的研究广谱基质金属蛋白酶抑制剂多西环素对卒中易感型自发性高血压大鼠(SHR-SP)脑血管系统基质金属蛋白酶的作用,探讨多西环素对脑卒中的发病率及病死率的影响。方法80只7周龄雄性SHR-SP大鼠随机分为药物干预组和对照组,每组再随机分为两个亚组,一个亚组用于观察脑卒中体征的发生情况,并比较两组脑卒中的发病率和病死率;采用明胶酶谱分析存活大鼠脑组织MMP-2和MMP-9活性,另一亚组进行血流动力学分析,且经氯化三苯基四唑氮(TTC)染色后用于分析脑卒中病灶的范围、数量和体积。结果(1)血流动力学分析显示多西环素组左心室 dp/dtmax、-dp/dtmin值较对照组增大[(15410.1±1679.2比对照组13684.1±1882.2和-10976.8±1372.6比对照组-9429.6±1840.9)mmHg/s,P<0.05]。(2)脑组织明胶酶谱分析显示多西环素组脑组织MMP-2活性较对照组降低。(3)大脑TTC染色结果显示多西环素组脑卒中病灶体积与对照组比较无显著性差异[(205.4±244.4比对照组167.1±274.3)mm3,P>0.05]。而多西环素组脑出血体积有大于对照组的趋势,但无统计学差异[(19.6±42.1比对照组3.4±5.0)mm3,P=0.176]。(4)多西环素组与对照组比较,脑卒中的病灶率(χ2=0.042)与病死率(χ2=0.135)无显著性差异(P>0.05)。结论多西环素可以抑制SHR-SP大脑中MMP-2活性,但在血压未控制时不能有效降低SHR-SP脑卒中发病率及脑卒中病灶体积。     

氨氯地平对糖尿病大鼠肾脏的保护作用

目的探讨苯磺酸氨氯地平对糖尿病大鼠肾脏细胞凋亡的影响。方法 21只 SD 雄性大鼠分为3组:正常对照组(对照组,n=6)喂以普通饲料;余15只大鼠高糖高脂膳食喂养6周后,一次性腹腔内注射小剂量链脲佐菌素(streptozotocin,STZ)30 mg/kg;2周后血糖升高≥16 mmol/L 者成模,共12只,再随机分为糖尿病组(糖尿病组,n=6)和糖尿病大鼠应用苯磺酸氨氯地平治疗组(治疗组,1 mg/kg·d,n=6)。12周后光镜观察各组。肾脏病理改变;流式细胞术检测大鼠。肾组织细胞凋亡率;检测各组内生肌酐清除率(Ccr)和24 h 尿白蛋白的排泄率(Uaer)。结果 12周后,糖尿病组大鼠肾小球细胞外基质增生,部分。肾小管轻度萎缩或管腔扩张,上皮细胞肿胀,胞浆内可见脂肪空泡变性,但治疗组较糖尿病组病变减轻;糖尿病大鼠较正常组的肾细胞凋亡率明显升高,氨氯地平能明显降低肾细胞凋亡率[对照组(13.8±1.3)% vs 糖尿病组(26.5±2.7)%vs 治疗组(20.0±2.0)%,P<0.05],增加肌酐清除率[对照组(0.110±0.020)vs 糖尿病组(0.021±0.001)vs 治疗组(0.0...     

葛根素对糖尿病大鼠心功能及心肌细胞内钙离子的影响

目的探讨葛根素对糖尿病大鼠心功能及心肌细胞内钙离子的影响。方法 40只SD大鼠随机分成空白对照组、糖尿病模型组、葛根素低剂量组和葛根素高剂量组,每组10只。测定大鼠心脏功能、心室重量指数、心脏重量指数和心肌细胞内钙离子瞬间变化。结果与糖尿病模型组比较,葛根素低剂量组和葛根素高剂量组心脏功能明显改善,心室重量指数、心脏重量指数及钙瞬变峰幅度下降,钙瞬变时程缩短,且高剂量组改善更加明显。结论葛根素可以改善糖尿病大鼠心功能、心室重量指数和心脏重量指数,其机制可能与调节心肌细胞内钙离子水平有关。     

糖尿病大鼠血、睾丸和脑组织内皮素水平的初步研究

目的　探讨糖尿病大鼠以及用银杏叶提取物 (EGb)治疗后血浆、睾丸和脑组织中内皮素的变化及意义。方法　将SD大鼠分为正常对照组、糖尿病模型组、EGb治疗组三组 ,在EGb治疗 5周后 ,用放射免疫法同时测定各组大鼠血浆、睾丸和脑组织中内皮素 (ET)的含量。结果　脑组织及血浆中ET含量正常对照组与糖尿病组比较无显著性差异 (P >0 0 5 ) ,睾丸组织中ET含量糖尿病组明显高于正常对照组 ,两者有显著性差异 (P <0 0 5 ) ;糖尿病组与EGb治疗组比较各种组织ET含量有下降倾向 ,但无显著性差异 (P >0 0 5 )。结论　糖尿病大鼠睾丸组织中ET含量增高 ,可能造成睾丸组织损害 ;EGb治疗可能有利于降低大鼠血液、睾丸和脑组织中的ET含量 ,但需进一步试验证实。     

Increased blood–brain barrier permeability and altered tight junctions in experimental diabetes in the rat: contribution of hyperglycaemia and matrix metalloproteinases

Aims/hypothesis Although diabetes mellitus is associated with peripheral microvascular complications and increased risk of neurological events, the mechanisms by which diabetes disrupts the blood–brain barrier (BBB) are not known. Matrix metalloproteinase (MMP) activity is increased in diabetic patients, is associated with degradation of tight junction proteins, and is a known mediator of BBB compromise. We hypothesise that diabetes leads to compromise of BBB tight junctions via stimulation of MMP activity. Materials and methods Diabetes was induced in the rat with streptozotocin. At 14 days after injection, BBB function was assessed by in situ brain perfusion. Tight junction proteins were assessed by immunoblot and immunofluorescence. Plasma MMP activity was quantified by fluorometric gelatinase assay and gel zymography. Results In streptozotocin-treated animals, permeability to [¹⁴C]sucrose increased concurrently with decreased production of BBB tight junction proteins occludin (also known as OCLN) and zona occludens 1 (ZO-1, also known as tight junction protein 1 or TJP1). Insulin treatment, begun on day 7, normalised blood glucose levels and attenuated BBB hyperpermeability to [¹⁴C]sucrose. Neither acute hyperglycaemia in naive animals nor acute normalisation of blood glucose in streptozotocin-treated animals altered BBB permeability to [¹⁴C]sucrose. Plasma MMP activity was increased in streptozotocin-treated animals. Conclusions/interpretation These data indicate that diabetes increases BBB permeability via a loss of tight junction proteins, and that increased BBB permeability in diabetes does not result from hyperglycaemia alone. Increased plasma MMP activity is implicated in degradation of BBB tight junction proteins and increased BBB permeability in diabetes. Peripheral MMP activity may present a novel target for protection of the BBB and prevention of neurological complications in diabetes. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible to authorised users.     

L-精氨酸和氨基胍对糖尿病大鼠一氧化氮、一氧化氮合酶及肾功能的影响

目的探讨L-精氨酸和氨基胍对早期糖尿病大鼠一氧化氮、一氧化氮合酶活性及肾功能的影响。方法Wistar大鼠60只,检测其24 h尿蛋白排泄量、血清一氧化氮水平、总一氧化氮合酶和诱导型一氧化氮合酶及结构型一氧化氮合酶活性等5项指标。然后用链脲佐菌素60 mg/kg制备糖尿病大鼠模型,将糖尿病鼠随机分为糖尿病对照组、L-精氨酸组和氨基胍组。于8周末时再检测大鼠上述5项指标并进行统计分析。结果与造模前比较,糖尿病对照组在8周末时24 h尿蛋白排泄量(43.92±7.38 mg)、一氧化氮水平(42.2±6.92μmol/L)和诱导型一氧化氮合酶活性(19.75±3.85 kU/L)升高(P<0.01,P<0.05);L-精氨酸组24 h尿蛋白排泄量(100.47±43.42 mg)和一氧化氮水平(67.34±18.87μmol/L)显著升高(P<0.01);氨基胍组24 h尿蛋白排泄量(22.33±3.47 mg)增加(P<0.01),总一氧化氮合酶(23.34±3.10 kU/L)、诱导型一氧化氮合酶(14.84±1.98 kU/L)和结构型一氧化氮合酶(8.50±2.25 kU/L)降低(P<0.01,P<0.05)。与糖尿病对照组比较,8周末时L-精氨酸组24 h尿蛋白排泄量和一氧化氮均升高(P<0.05),总一氧化氮合酶、诱导型一氧化氮合酶和结构型一氧化氮合酶活性差异无显著性;氨基胍组与L-精氨酸组比较,上述5项指标均下降(P<0.05)。结论在糖尿病肾病早期应用L-精氨酸可增加血一氧化氮的合成,使24 h尿蛋白排泄量增加,损害肾功能;而早期应用氨基胍可降低血一氧化氮、一氧化氮合酶及24 h尿蛋白排泄量,保护肾功能。     

钾流在糖尿病大鼠心室肌细胞的改变机制

目的研究糖尿病对大鼠心室肌细胞瞬间外向钾流(I_(to))的影响及其分子机制,探讨糖尿病引起的心脏损害与心律失常的关系。方法取体质量150～200 g 的雄性 Sprague-Dawley 大鼠,单次腹腔注射链脲菌素(STZ,65 mg/kg,pH=4.5)建立糖尿病大鼠模型,采用酶解法获得单个心室肌细胞,应用膜片钳全细胞方法记录I_(to);并用反转录聚合酶链式反应技术进一步半定量编码该电流通道α亚单位基因(Kv4.2、Kv4.3和 Kv1.4)mR-NA 的表达水平。结果与对照组比较,+70 mV 时,糖尿病大鼠心室肌细胞 I_(to)密度显著降低[对照组:(30.6±3.8)比糖尿病组:(18.9±3.3)pA/pF,P<0.01);半定量分析法显示糖尿病大鼠心室肌细胞 I_(to)通道α亚单位编码基因 Kv4.2、Kv4.3 mRNA 表达水平分别下调56.9%和46.6%;而 Kv1.4 mRNA 表达则上调约48.0%,3组基因表达水平的改变差异均有统计学意义(P<0.05)。结论糖尿病大鼠心室肌细胞 I_(to)密度显著降低主要与编码该通道α亚单位的基因表达下调有关。     

硒、锰对大鼠全血及组织中硒、锰水平以及谷胱甘肽过氧化物酶活性的影响

本文以干酪素及蔗糖作主要成分构成合成饲料,观察饲料中不同水平的硒、锰含量对大鼠体内硒、锰水平及谷胱甘肽过氧化物酶活性的影响。实验结果表明,低硒组的全血、心肌、肝脏、肾及毛中的硒含量均显著低于常硒组。全血、心肌及肝脏的GSH-Px活性也明显受饲料硒含量的影响。值得注意的是喂富锰饲料的大鼠其全血、肝脏及心肌锰含量与常锰组相当,因此,饲料锰含量对大鼠组织硒含量及GSH-Px活性也无明显的影响。提示锰对大鼠体内硒含量及GSH-Px活性的影响是复杂的,它可能受到饲料组成及饲料中蛋白质含量的明显影响。     

Effects of Maternal Alcohol Consumption on Milk and Blood Lymphocytes and IgG Antibody Levels from Trichinella spiralis-Infected Rats

Immunity to the intestinal parasite Trichinella spiralis can be transferred from the mother to the neonate during lactation. Previous studies in our laboratory showed that the passage of immunity to pups from ethanol-treated dams was depressed. This study examined the effect of ethanol consumption during pregnancy and lactation on the T. spiralis-associated immune components in milk and blood. Groups of female rats were fed either ethanol-containing or isocaloric liquid diets for 30 days before T. spiralis infection, mated and maintained on corresponding diets through pregnancy and lactation. Two-color flow cytometric analysis was performed for lymphocyte populations, enzyme-linked immunoabsorbent assay for specific IgG, and radial immunodiffusion assay for total IgG. The percentage of total T cells and their subsets, T helper cells and T cytotoxic/suppressor cells in milk and those in blood were similar between pair-fed and ethanol-treated animals. However, the percentage of natural killer cells in milk from ethanol animals was significantly reduced compared with the pair-fed group (33% vs. 54%). The percentage of activated or memory type T helper cell subset (OX22^?W3/25⁺) was significantly increased in the blood of the ethanol-treated group. Pair-fed animals showed higher T. spiralis-specific IgG antibody levels both in milk and blood compared with ethanol-treated animals. In ethanol-treated animals, specific IgG levels and total IgG concentration in milk were significantly lower than those in blood, whereas in pair-fed animals, only total IgG concentration in milk was lower than that in blood. This study indicates that ethanol consumption during pregnancy and lactation alters the maternal immune system. Decreased milk natural killer cells and depressed specific antibody levels in milk of ethanol-treated animals are possible contributing factors to the previously observed depressed lactational immune transfer to the pups.     

胰岛素样生长因子-1对糖尿病大鼠结肠平滑肌细胞表达干细胞因子的影响

背景：糖尿病胃肠动力障碍与Cajal间质细胞（ICC）数量和超微结构异常有关。前期实验发现胰岛素样生长因子-1（IGF-1）可诱导正常大鼠胃肠平滑肌细胞（SMC）表达干细胞因子（SCF）,从而有利于ICC的生存。目的：探讨IGF-1对糖尿病大鼠结肠SMC表达SCF的影响及其信号转导通路。方法：以链脲霉素建立糖尿病大鼠模型。分离、培养正常和糖尿病大鼠结肠SMC,设不同浓度（0、50、100、150μg/L）、不同时间（0、8、16、24、48h）IGF-1十预组和MEK抑制剂PD98059＋IGF-1、P13K抑制剂LY294002＋IGF-1干预组．以RT—PCR和蛋白质印迹法检测SMC中的SCF表达。结果：糖尿病大鼠结肠SMC的生长速度较正常大鼠减慢。无血清培养条件下．正常和糖尿病结肠SMC中SCFmRNA和蛋白表达较低,IGF-1可诱导SCF表达增加,最大有效浓度为100μg／L,诱导高峰时间正常对照组为16h,糖尿病组为24h。经PD98059预处理的SMC,IGF-1诱导的SCF表达部分受抑．IN294002预处理对IGF-1的作用无明显影响。结论：0～100μg／L IGF-1在24h内能以剂量和时间依赖性方式诱导糖尿病大鼠结肠SMC表达SCF．但效应弱于其对正常大鼠结肠SMC的作用,该作用的发挥可能部分依赖于ERKMAPK信号通路。     

Green Synthesis,Characterizations of Zinc Oxide Nanoparticles from Aqueous Leaf Extract of Tridax procumbens Linn. and Assessment of their Anti-Hyperglycemic Activity in Streptozoticin-Induced Diabetic Rats

Herein, zinc oxide nanoparticles (ZnO NPs) were greenly synthesized from Tridax procumbens aqueous leaf extract (TPE) and characterized physically (e.g., Fourier-transform infrared (FTIR) spectroscopy and scanning electron microscopy (SEM)) and biologically (test of their anti-diabetic activity). Anti-diabetic activities of TPE and TPE-derived ZnO NPs have been carried out in a streptozotocin (STZ)—induced diabetic rat model. Diabetes mellitus (DM) was induced with a single intraperitoneal dosage of the glucose analogue STZ (55 mg/Kg) known to be particularly toxic to pancreatic insulin-producing beta-cells. TPE and TPE-derived ZnO NPs were administered orally, once every day for 21 days in diabetic rats, at 100 and 200 mg/Kg, respectively. The standard antidiabetic medication, glibenclamide, was used as a control at a dose of 10 mg/Kg. Various parameters were investigated, including bodyweight (bw) variations, glycemia, lipidaemia, glycated hemoglobin (HbA1c), and histopathological alterations in the rat’s liver and pancreas. The TPE-mediated NPs were small, spherical, stable, and uniform. Compared to TPE and, to a lesser extent, glibenclamide, TPE-derived ZnO NPs lowered blood glucose levels considerably (p < 0.05) and in a dose-dependent manner while preventing body weight loss. Further, positive benefits for both the lipid profile and glycated hemoglobin were also noticed with TPE-derived ZnO NPs. The histopathological assessment revealed that synthesized TPE-derived ZnO NPs are safe, non-toxic, and biocompatible. At 200 mg/Kg/day, TPE-derived ZnO NPs had a more substantial hypoglycemic response than at 100 mg/Kg/day. Thus, in this first reported experimental setting, ZnO NPs biosynthesized from the leaf extract of Tridax procumbens exert more potent anti-diabetic activity than TPE and glibenclamide. We conclude that such a greenly prepared nanomaterial may be a promising alternative or complementary (adjuvant) therapy, at least to the current Indian’s traditional medicine system. Translational findings are prompted in human populations to determine the efficacy of these NPs.