Proteins Implicated In Cardiac Senescence: Results From

心血管科学实验室致力于识别和研究心肌的衰老过程。为了鉴定心脏中可能与衰老有关的基因产物，用来自Fisher 344大鼠左心室的mRNA分析了cDNA微阵列，CBA小鼠用生物肽治疗，认为这在老年保护中很重要，以及来自衰竭和非衰竭心肌的人类活检。这些项目的目的是确定哪些基因产物作为年龄或疾病的函数受到调节，然后使用独立的方法来确定导致基因表达改变的潜在机制。 为了鉴定由生物肽（合成胸腺二肽Vilon（Lys-Glu）或松果体四肽Epitalon（Ala-Glu-Asp-Gly））调节的早期分子事件，与位于彼得堡的俄罗斯医学科学院合作，在CBA小鼠中研究了基因表达谱。与来自对照和实验小鼠（Vilon和Epitalon）的心脏RNA样品杂交的cDNA基因表达阵列的比较分析显示，对于Vilon和Epitalon组，300个克隆（克隆总数的1.97%）的表达变化>2倍。其中，36个转录本的表达被Vilon独特地改变，105个被Epitalon独特地改变。Vilon和Epitalon导致159个转录本的保守和显著的表达变化。该分析鉴定了参与细胞分裂（Vilon或Epitalon为14个）、细胞信号传导/通信（分别为15和14个）、细胞结构/运动性（7和6个）、细胞/生物体防御（16和13个）、基因/蛋白质表达（24和17个）、代谢（19和11个）和由线粒体DNA编码的基因（5和5个）的多个基因。对比分析表明，Vilon和Epitalon对心脏基因表达的影响具有普遍性和独特性。约22%的成绩单表现出改变基因表达的两个实验组。对褪黑激素也进行了类似的研究。这些基因产物可以解释这些肽的老年保护作用的分子基础。 我们目前正在完成老化Fisher 344老化大鼠模型的分析。只有4.3%的转录本显示出年龄依赖性改变，其中大部分发生在生命后期（64%，249/388发生在30个月）。我们正在完成一项广泛的定量PCR分析，以验证这些数据，并通过独立的技术，以确定基因表达的变化与衰老过程的意义。 我们还采用cDNA微阵列来检查来自衰竭（n=8）和非衰竭人类心肌（n=7）的LV的转录组。在通过微阵列和统计学方法鉴定了一组HF应答候选基因后，我们在较大的样本群体（n=34）上采用Q-PCR来验证和检查贡献生物变量（年龄和性别）的作用。我们发现，大多数HF候选基因（包括转录因子，修饰酶，ECM蛋白和代谢酶）表现出基因表达的显着变化，然而，大多数假定的变化取决于变量，如性别和年龄，而不是单独的HF。此外，一些推定的HF反应基因产物表现出高度显着的变化，表达作为年龄和/或性别的函数，但独立于HF。 这些项目的数据正在进行比较，以确定不仅与衰老有关，而且与疾病有关的候选基因。

The Laboratory of Cardiovascular Science has a strong commitment to identifying and studying the process of aging in the myocardium. To identify gene products in heart potentially involved in aging, cDNA microarrays have been analyzed with mRNA from left ventricles of Fisher 344 rats, CBA mice treated with biopeptides thought to be important in gerontoprotection, and human biopsies from failing and non-failing myocardium. The aim of these projects are to determine which gene products are regulated as a function of age or disease, and then use independent methods to determine the underlying mechanisms responsible for the altered changes in gene expression. To identify early molecular events regulated by biopeptides (synthetic thymic dipeptide Vilon (Lys-Glu) or with pineal tetrapeptide Epitalon (Ala-Glu-Asp-Gly)), gene expression profiles have been studied in CBA mice, in collaboration with the Russian Academy of Medical Science, in St. Petersburg. Comparative analysis of cDNA gene expression arrays hybridized with heart RNA samples from control and experimental mice (vilon and epitalon) revealed 300 clones (1,97% of the total number of clones) with >2-fold change in expression for Vilon and Epitalon groups. Of these, expression of 36 transcripts was uniquely altered by Vilon and 105 by Epitalon. Vilon and Epitalon led to conserved and significant changes in expression in 159 transcripts. The analysis identified multiple genes involved at cell division (14 for Vilon or Epitalon), cell signaling/communication (15 and 14, respectively), cell structure/motility (7 and 6), cell/organism defense (16 and 13), gene/protein expression (24 and 17), metabolism (19 and 11), and genes encoded by mitochondrial DNA (5 and 5). Comparative analysis demonstrated both general and unique features of Vilon and Epitalon effects on cardiac gene expression. About 22% of transcripts demonstrated altered gene expression for both experimental groups. Similar studies have been performed with melatonin. These gene products that can explain the molecular basis for the geroprotective effects of these peptides. We are currently completing the analysis of an aging Fisher 344 aging rat model. Only 4.3% of the transcripts showed age-dependent alterations, most of which occurred late in life (64%, 249/388 occurred at 30 months). We are completing an extensive quantitative-PCR analysis to validate these data, and by independent techniques, to determine the significance of the changes in gene expression with the aging process. We have also employed cDNA microarrays to examine the transcriptomes of LVs from failing (n=8) and non-failing human myocardium (n=7). Following identification of a pool of HF-responsive candidate genes by microarrays and statistical methods, we employed Q-PCR on a larger sample population (n=34) to validate and examine the role of contributing biological variables (age and gender). We find that most of the HF-candidate genes (including transcription factors, modifying enzymes, ECM proteins and metabolic enzymes) demonstrated significant changes in gene expression; however, the majority of the putative changes depended on variables such as sex and age, and not on HF alone. Additionally, some putative HF-responsive gene products demonstrated highly significant changes in expression as a function of age and/or sex, but independent of HF. The data from each of these projects are being compared in an effort to identify candidate genes implicated not only in aging, but also in disease.