A new technique of coronary artery ligation: Experimental myocardial infarction in rats in vivo with reduced mortality

In vivo models of myocardial infarction following coronary artery ligation in the rat still suffer from high early mortality and a low rate of success of myocardial infarction. This study investigated the possibility of reducing early mortality and increasing the rate of myocardial infarction by modifications of surgical techniques. Eighteen rats were divided into two groups: normal control (3 rats) and ligation (15 rats). The major modifications of surgical techniques used in this study include: (1) no exteriorization of the heart, (2) ligation of the origins of the branches rather than the main trunk of the left coronary artery, (3) removal of air from the chest after closure, (4) supplying oxygen immediately after extubation. Following surgery, the rats recovered uneventfully and 11 rats were alive after 16 weeks. One rat, with a large myocardial infarction, died 2 h after surgery. Early mortality (during surgery and 1 week after surgery) was 6.7% with a success rate of myocardial infarction of 85%. The left ventricle in the ligation group showed significant dilation relative to normal and shamoperated control hearts (317% of control hearts, p < 0.001). However, myocardial mass did not increase. The average infarct size was 33%. These results demonstrate that a reduction in early mortality and an increased success rate of myocardial infarction can be achieved by modifications of surgical techniques.     

大鼠脑缺血再灌注后Cathepsin D和caspase．9表达的动态变化

目的：探讨大鼠缺血再灌注后溶酶体组织蛋白酶D（Cathepsin DCD）、caspase-9在不同时段蛋白质及mRNA表达变化。方法：将60只SD大鼠随机分为三组：正常组（10只）,假手术组（10只）,脑缺血再灌注组（40只）,线栓法制备大脑中动脉梗死模型（MCAO）,免疫组化及RT—PCR法分别检测CathepsinD、caspase-9的蛋白和mRNA表达。结果：与正常组和假手术组比较,模型组大鼠脑缺血再灌注损伤后6h Cathepsin D的蛋白和mRNA表达明显增强（P〈0．05）,24h达高峰,48h仍保存高水平。caspase．9蛋白和mRNA6h开始明显升高,12h达高峰,此后缓慢下降,但48h组仍显著高于对照组（P〈0．05）,结论：Cathepsin D、caspase．9在大鼠脑缺血再灌注后表达增强,溶酶体可能参与了脑缺血再灌注损伤后神经细胞凋亡的过程。     

大鼠脑缺血再灌注后组织蛋白酶D蛋白质及mRNA的表达

目的：探讨大鼠缺血再灌注后溶酶体组织蛋白酶D（Cathepsin DCD）在不同时段蛋白质及mRNA表达变化。方法：将60只SD大鼠随机分为三组：正常组（10只）,假手术组（10只）,脑缺血再灌注组（模型组）（40只）,线栓法制备大脑中动脉梗死模型（MCAO）,免疫组化及RT-PCR法分别检测CathepsinD的蛋白和mRNA表达。结果：与正常组和假手术组比较,模型组在大鼠脑缺血再灌注损伤后6hCathepsin D的蛋白和mRNA表达明显增强（P＆lt;0.05）,24h达高峰,48h仍保存高水平。结论：Cathepsin D在大鼠脑缺血再灌注后表达增强,溶酶体CathepsinD可能参与了脑缺血再灌注损伤后神经细胞凋亡。     

Differential Modulation of Arcuate Nucleus and Mesolimbic Gene Expression Levels by Central Leptin in Rats on Short-Term High-Fat High-Sugar Diet

Objective

Leptin resistance is a common hallmark of obesity. Rats on a free-choice high-fat high-sugar (fcHFHS) diet are resistant to peripherally administered leptin. The aim of this study was to investigate feeding responses to central leptin as well as the associated changes in mRNA levels in hypothalamic and mesolimbic brain areas.

Design and Methods

Rats on a CHOW or fcHFHS diet for 8 days received leptin or vehicle intracerebro(lateral)ventricularly (ICV) and food intake was measured 5 h and 24 h later. Four days later, rats were sacrificed after ICV leptin or vehicle and mRNA levels were quantified for hypothalamic pro-opiomelanocortin (POMC) and neuropeptide Y (NPY) and for preproenkephalin (ppENK) in nucleus accumbens and tyrosine hydroxylase (TH) in ventral tegmental area (VTA).

Results

ICV leptin decreased caloric intake both in CHOW and fcHFHS rats. In fcHFHS, leptin preferentially decreased chow and fat intake. Leptin increased POMC and decreased NPY mRNA in CHOW, but not in fcHFHS rats. In CHOW rats, leptin had no effect on ppENK mRNA and decreased TH mRNA. In fcHFHS, leptin decreased ppENK mRNA and increased TH mRNA.

Conclusion

Despite peripheral and arcuate leptin resistance, central leptin suppresses feeding in fcHFHS rats. As the VTA and nucleus accumbens are still responsive to leptin, these brain areas may therefore, at least partly, account for the leptin-induced feeding suppression in rats on a fcHFHS diet.     

Correlations between the endothelium-dependent relaxation of the aorta and arterial pressure in rats with myocardial infarction

The effect of experimental myocardial infarction on endothelium-dependent relaxation was studied on isolated rat aorta and compared with the dynamics of arterial pressure (AP). It was shown that the endothelium-dependent relaxation of aorta was increased 1.8 times 3 h following the myocardial infarction. Simultaneously the drop in AP which had begun immediately following the experimental infarction became maximal. In 24 h both the indices were restored practically to the initial level. There was a significant negative correlation between the extent of endothelium-dependent relaxation and AP. It was suggested that the increase in endothelium-dependent relaxation could influence vascular tone, the drop in AP, and, finally, the development of cardiogenic shock in myocardial infarction in man.     

ZFP580对大鼠心肌缺血/再灌注后心室重塑的影响

目的：探究锌指转录因子（ZFP580）与心肌缺血/再灌注损伤后心室重塑的关系。方法：72只SD大鼠随机分为假手术（sham）组（n=8）和心肌缺血/再灌注（I/R）组（n=64）,其中I/R组分别在再灌注后的0.5 h、1 h、2 h、4h、1 d,7 d,14 d,28 d处死后取材,观察心肌组织中ZFP580的表达。培养大鼠H9C2心肌细胞,每组设3个复孔,分别在转化生长因子β1（TGF-β1）刺激0 h、8 h、16 h、24 h后观察心肌细胞肥大情况,并检测心肌细胞中β-MHC、心房利钠肽（ANP）以及ZFP580 mRNA的表达。利用慢病毒介导的基因转染获得高表达ZFP580的H9C2心肌细胞,转染72h后,检测心肌细胞中基质金属蛋白酶3（MMP-3）的表达。结果：成功建立心肌缺血/再灌注损伤模型,大鼠心肌I/R损伤后第14天,心肌组织大面积梗死,心肌细胞呈嗜酸性变。大鼠心肌组织中ZFP580及TGF-β1表达上调。TGF-β1（5 ng/ml）刺激H9C2心肌细胞后诱导心肌细胞肥大,心肌细胞肥大标志蛋白β-MHC、ANP表达上调,且心肌细胞中ZFP580mRNA表达上调（P < 0.05）。高表达ZFP580的H9c2心肌细胞中MMP-3表达下调（P < 0.05）。结论：锌指转录因子ZFP580可能参与了心肌缺血/再灌注后心室重塑的过程,其作用可能与参与TGF-β1诱导的心肌细胞肥大过程以及抑制心肌细胞产生MMP-3有关。     

Expression of the preproenkephalin A gene in the heart of cardiomyopathic hamsters.

The expression of the preproenkephalin A gene (Enk gene), which codes for the precursor of enkephalins, was investigated in the heart of hamsters with a hereditary cardiomyopathy at four different stages of the disease: the prenecrotic stage (30 days), the necrotic stage (60 days), the hypertrophic stage (120 days), and the final stage (200 days). In control atria and ventricles, the relative abundance of the Enk mRNA, as assessed by Northern blot analysis, did not change upon ageing. In the ventricles of cardiomyopathic hamsters, however, it increased about two- to three-fold at the necrotic stage, but was unaltered at the other time points studied; whereas in the atria, it progressively decreased to reach about half that of control hamsters at the final stage. Enkephalin levels, as measured by radioimmunoassay, decreased at 60 days in both the atria and ventricles of control hamsters, and also in the atria of cardiomyopathic hamsters, and remained stable thereafter, corresponding to one-third to one-half of those at 30 days. However, in the ventricles of cardiomyopathic hamsters, the peptide level decreased only slightly, the consequence being that at 60, 120, and 200 days, it was about two- to three-fold that of control hamsters. The lack of correlation between peptide levels and the relative abundance of the Enk mRNA suggests that translational and (or) post-translational mechanisms are important in the control of the expression of the Enk gene in the heart of hamsters.     

Distribution of mRNAS of fibrinogen polypeptides and albumin in free and membrane-bound polyribosomes and induction of alpha-fetoprotein mRNA synthesis during liver regeneration after partial hepatectomy

To study the effect of regenerative response of the liver following partial hepatectomy on the synthesis of major plasma proteins (secretory proteins), we have determined the sequence contents and the distribution of albumin and fibrinogen polypeptide mRNAs in rat liver at intervals after partial hepatectomy and sham operation. Using a quantitative technique for the isolation of polyribosomes, we demonstrated that the distribution of RNA between free and membrane-bound polyribosomal fraction was unchanged in these experiments. There was no shift in the polyribosomal population to favor free polyribosomes after partial hepatectomy. However, there was a dramatic increase (5-6-fold) of the fibrinogen polypeptide mRNA concentration during the first 24 h after resection. In contrast, the albumin mRNA concentration decreased (2-3-fold). There were no alpha-fetoprotein mRNA sequences detectable in any liver RNA fraction in these experimental animals. In sham-operated rats with intact livers, similar changes of fibrinogen polypeptide and albumin mRNA concentrations as described in regenerating liver after partial hepatectomy, were observed. These results suggest that albumin and fibrinogen synthesis after partial hepatectomy is reciprocally regulated at the mRNA level and represents a nonspecific acute phase response to surgical trauma.     

Differential time course of liver and kidney glucose-6 phosphatase activity during long-term fasting in rat correlates with differential time course of messenger RNA level

We have studied the role of Glc6Pase mRNA abundance in the time course of Glc6Pase activity in liver and kidney during long-term fasting in rat. Refered to the mRNA level in the fed state, Glc6Pase mRNA abundance was increased by 3.5 ± 0.5 and 3.7 ± 0.5 times (mean ± S.E.M., n = 5) in the 24 h and 48 h-fasted liver, respectively. Then, the liver Glc6Pase mRNA was decreased to the level of the fed liver after 72 and 96 h of fasting (1.0 ± 0.3 and 1.4 ± 0.3). In the kidney, Glc6Pase mRNA abundance was increased by 2.7 ± 1.0 and 5 ± 1.2 times at 24 and 48 h of fasting, respectively. Then, it plateaued at the level of the 48 h fasted kidney after 72 h and 96 h of fasting (4.5 ± 1.0 and 4.3 ± 1.0). After 24 and 48 h-refeeding, the abundance of Glc6Pase mRNA in 48 h-fasted rats was decreased to the level found in the liver and kidney of fed rats. The time course of the activity of Glc6Pase catalytic subunit during fasting and refeeding was strikingly parallel to the time course of Glc6Pase mRNA level in respective tissues. These data strongly suggest that the differential expression of Glc6Pase activity in liver and kidney in the course of fasting may be accounted for by the respective time course of mRNA abundance in both organs.Abbreviations Glc6Pase Glucose-6 phosphatase - GNG Gluconeogenesis     

Enhanced early after-myocardial infarction concentration of TNF-alpha subsequently increased circulating and myocardial adrenomedullin in spontaneously hypertensive rats

Both inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) and the cardiac protective peptide adrenomedullin (AM) are increased in cardiac tissues and plasma in patients with myocardial infarction (MI) and chronic heart failure. Recently they have been increasingly recognized as important factors in the pathophysiology of MI and resultant congestive heart failure. Compared with sham-operated spontaneously hypertensive rats (SHR), we investigated myocardial immunoreactivity of TNF-alpha and AM and also their mutual relations in vivo in SHR+MI. Residual myocardial depression after MI was studied also in isolated perfused hearts. In chronic experiments, 24 and 48 h after permanent ligation of the descending anterior branch of the left coronary artery, we examined hemodynamics, plasma and myocardial peptide levels. Left ventricular function was assessed in isolated perfused hearts subjected to "global ischemia and reperfusion" and after induction of "calcium paradox". Circulating and myocardial TNF-alpha concentrations increased early after MI in SHR. Studies with global ischemia and calcium paradox in isolated heart showed early myocardial depression and calcium-dependent gradual increase of left-ventricular end-diastolic pressure. In the SHR+MI myocardial AM concentrations were increased 9- and 49-fold after respective 24 h and culminated 48 h following MI. Circulating and myocardial AM was increased in SHR+MI in association with TNFalpha-induced myocardial depression. The both studied cardiac parameters displayed the beneficial effect of the enhanced myocardial AM concentration.     

Proliferative and nutritional dependent regulation of glyceraldehyde-3-phosphate dehydrogenase expression in the rat liver

Glyceraldehyde-3-phosphate dehydrogenase is a multifunctional protein possessing numerous cytoplasmic and nuclear functions associated with cellular proliferation. Despite the emerging role of glyceraldehyde-3-phosphate dehydrogenase in regulating the proliferative process, there is a paucity of data regarding its expression and intracellular distribution in non-malignant proliferating hepatocytes. Thus the aim of the present study was to document the intracellular distribution of glyceraldehyde-3-phosphate dehydrogenase protein in proliferating hepatocytes derived from regenerating rat livers, and glyceraldehyde-3-phosphate dehydrogenase gene expression in fasted and re-fed rats following partial hepatectomy (PHx). Glyceraldehyde-3-phosphate dehydrogenase mRNA and protein expression were documented by Northern and Western blot analyses, respectively, at various times following 70% PHx in adult Sprague-Dawley rats. At 24 h post-surgery, glyceraldehyde-3-phosphate dehydrogenase mRNA expression was significantly increased in both PHx and sham operated rats (P < 0.001), respectively. Despite the increase in glyceraldehyde-3-phosphate dehydrogenase mRNA expression in both groups, only PHx rats had a significant increase in the nuclear fraction of glyceraldehyde-3-phosphate dehydrogenase protein (threefold increase compared to sham and baseline levels, P < 0.01), cytoplasmic levels of glyceraldehyde-3-phosphate dehydrogenase protein remained unaltered in both groups. In terms of the effects of feeding and fasting on rats there were no significant differences in glyceraldehyde-3-phosphate dehydrogenase mRNA levels, whether fasted or refed, in rats that had undergone PHx, 8 h earlier. On the other hand, glyceraldehyde-3-phosphate dehydrogenase mRNA levels were significantly increased in refed compared to fasted sham operated rats 8 h following surgery. Serum insulin concentrations were higher in the refed PHx and sham groups compared to their fasted counterparts. The results of this study indicate that although glyceraldehyde-3-phosphate dehydrogenase mRNA are altered to the same extent in PHx and sham-operated rats following surgery, increases in the nuclear fraction of glyceraldehyde-3-phosphate dehydrogenase protein only occur in PHx rats. The results also indicate that glyceraldehyde-3-phosphate dehydrogenase expression is affected by the nutritional status of animals undergoing abdominal sham surgery.     

CD4^＋T细胞在大鼠心肌缺血再灌注损伤中的作用

目的通过大鼠心肌缺血再灌注损伤的动物模型,分析CD4^＋T细胞在心肌组织损伤中的作用。方法结扎大鼠冠状动脉左前降支45min,随后恢复再灌的方法,制作缺血再灌损伤的动物模型,随机分为再灌注0、2、6、9、12h组及相应的对照组。II导联心电图及TTC确定模型,组织病理学观察心肌细胞的损伤情况,免疫荧光染色计数浸润的炎性细胞,半定量PCR进一步验证各型T细胞的表达。结果心肌的梗死面积与心肌缺血再灌时间成正相关,至观察结束未出现峰值;组织中浸润的中型粒细胞和T细胞分别在2h和12h有峰值出现,但CD4^＋T/CD3^＋T的比率几乎保持不变;观察所见CD4^＋T细胞是组织中存在最多的T细胞。结论大鼠缺血再灌注损伤中,心肌组织中浸润的CD4^＋T细胞作为主要的效应细胞,参与了持续稳定的心肌损伤过程。     

Distribution of mRNAs of fibrinogen polypeptides and albumin in free and membrane-bound polyribosomes and induction of α-fetoprotein mRNA synthesis during liver regeneration after partial hepatectomy

To study the effect of regenerative response of the liver following partial hepatectomy on the synthesis of major plasma proteins (secretory proteins), we have determined the sequence contents and the distribution of albumin and fibrinogen polypeptide mRNAs in rat liver at intervals after partial hepatectomy and sham operation. Using a quantitative technique for the isolation of polyribosomes, we demonstrated that the distribution of RNA between free and membrane-bound polyribosomal fraction was unchanged in these experiments. There was no shift in the polyribosomal population to favor free polyribosomes after partial hepatectomy. However, there was a dramatic increase (5–6-fold) of the fibrinogen polypeptide mRNA concentration during the first 24 h after resection. In contrast, the albumin mRNA concentration decreased (2–3-fold). There were no α-fetoprotein mRNA sequences detectable in any liver RNA fraction in these experimental animals. In sham-operated rats with intact livers, similar changes of fibrinogen polypeptide and albumin mRNA concentrations as described in regenerating liver after partial hepatectomy, were observed. These results suggest that albumin and fibrinogen synthesis after partial hepatectomy is reciprocally regulated at the mRNA level and represents a nonspecific acute phase response to surgical trauma.     

原花青素对脑缺血再灌损伤大鼠模型的影响

目的研究原花青素对脑缺血/再灌损伤(ischemia/reperfusion,I/R)大鼠神经功能评分(neurologicaldeficit score,NDS)、脑梗死体积、脑含水量等指标的药理作用。方法采用大鼠大脑中动脉阻断(middle cerebralartery occlusion,MCAO)法复制类似人类缺血性卒中的I/R损伤模型。结果该模型各时间点内均有程度不同的神经功能缺失,原花青素给药组神经功能评分明显低于对照组(P0.05),假手术组大鼠均无神经功能缺失,脑水肿情况均较对照组明显改善(P0.05),脑梗死体积与盐水对照组相比差异有显著性(P0.05),而假手术组均未见有梗死灶。结论原花青素具有一定的保护大鼠I/R后受损脑组织的作用,可供后续研究,并可为缺血性卒中使用原花青素治疗提供确凿的理论依据。     

Protein-synthesizing function of the liver of rabbits in experimental myocardial infarct

The content of serum albumin in rabbit blood was found to be lowered within the first day after reproduction of experimental myocardial infarction. The rate and the level of translation of endogenous mRNA were studied in cell-free systems from normal rabbit liver and 6-12-24 h after experimental myocardial infarction. The decrease of the total protein synthesis in the crude cell-free system from the liver of experimental animals was shown to depend on the lack of energy supply rather than on the reduced activity of the protein-synthesizing apparatus. The relative drop of protein synthesis in the cell-free system with saturating concentration of ATP, GTP and creatine phosphate is likely to be connected with a decrease in the proportion of membrane-bound polysomes.     

Levels of mRNAs coding for lipogenic enzymes in rat lung upon fasting and refeeding and during perinatal development

The relative amounts of mRNAs coding for fatty-acid synthase (EC 2.3.1.85), acetyl-CoA carboxylase (EC 6.4.1.2), ATP citrate lyase (EC 4.1.3.8) and malic enzyme (EC 1.1.1.40) were determined in lungs and livers of adult rats that were normally fed, starved for 48 h or starved for 48 h and subsequently refed for 72 h with a carbohydrate-rich, fat-free diet. In the liver, starvation caused a small decrease in the relative abundance of the mRNAs which was not statistically significant. Subsequent refeeding caused a statistically significant increase in mRNAs for all of the enzymes studied. In the lung, no significant changes were found, indicating that the regulation of the abundance of mRNAs encoding the lipogenic enzymes in the lung differs from that in the liver. In the developing rat lung, mRNA for fatty-acid synthase increased 3-fold in abundance between fetal days 18 and 20 and decreased directly after birth (at day 22 of gestation). A similar pattern was observed for ATP citrate lyase mRNA. The level of acetyl-CoA carboxylase mRNA decreased significantly after birth. These observations indicate that in perinatal rat lungs, pretranslational regulation is involved in the control of the synthesis of these enzymes. The abundance of acetyl-CoA carboxylase mRNA did not change in the prenatal period, a time during which the specific activity of this enzyme increases. This lack of correlation between the specific activity of acetyl-CoA carboxylase and the abundance of its mRNA may indicate that translational regulation of the synthesis of the enzyme or post-synthetic regulatory effects on enzyme molecules are involved in the control of this enzyme in the prenatal period. No changes in the abundance of lung malic enzyme mRNAs were observed throughout the perinatal period.