CELL-SELECTIVE EXPRESSION OF FIBROTIC GENE PATHWAYS

纤维化基因途径的细胞选择性表达

• 批准号: 7480441
• 负责人: Nestor F Gonzalez-Cadavid
• 金额: $ 18.17万
• 依托单位: LUNDQUIST INSTITUTE FOR BIOMEDICAL INNOVATION AT HARBOR-UCLA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-07 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7480441
• 关键词: Abbreviations; Actins; Age; Aging; Animal Model; Arterial Medias; Arteries; Arteriosclerosis; Biological Assay; Blood Vessels; Body of uterus; Cell Communication; Cell Nucleus; Cells; Collagen; Collagen Fiber; Condition; Confocal Microscopy; Control Groups; Corpora Cavernosa; Cultured Cells; DAPI; DNA Microarray Chip; DNA Microarray format; Data; Deposition; Detection; Development; Diabetes Mellitus; Disease; Dorsal; End Point; Erectile dysfunction; Fibroblasts; Fibrosis; Fluorescence; Functional disorder; Gene Expression; Gene Expression Alteration; Genes; Goals; Green Fluorescent Proteins; Hydroxyproline; Image Analysis; Immunofluorescence Immunologic; Immunohistochemistry; Implant; Injury; Label; Lead; Microspheres; Molecular; Molecular Profiling; Molecular Target; Myofibroblast; Names; Nitric Oxide Synthase; Numbers; Organ; Pathway interactions; Pattern; Peyronie Disease; Polymerase Chain Reaction; Process; Public Health; RNA; Rattus; Reactive Oxygen Species; Role; Saline; Smooth Muscle Myocytes; Specificity; Stem cells; Time; Tissues; Transforming Growth Factors; Transplantation; Tunica Adventitia; Tunica Albuginea; Vascular Diseases; Vascular System; Veno-occlusive; Vimentin; Week; Western Blotting; adult stem cell; age related; aged; arterial stiffness; cell type; day; implantation; in vivo; laser capture microdissection; male; novel; paracrine; penis; promoter; research study; selective expression

DESCRIPTION (provided by applicant): This project aims to clarify some aspects that would help to define a common molecular profile of vascular fibrosis, that in the penis causes Peyronie's disease (PD) and aging-related vasculogenic erectile dysfunction, and that in the arteries leads to aging-related arteriosclerosis and arterial stiffness, in order to detect cellular and molecular targets of antifibrotic therapy. We will investigate: a) the cell specificity of the alterations of gene expression occurring in penile and vascular fibrosis, clarifying the role of smooth muscle cells (SMC) and myofibroblasts in these processes; b) whether the SMC in the penile trabecular tissue are responsible for excessive collagen synthesis; c) the ontogenic relationship of myofibroblasts and SMC, and d) the effect of cell to cell interactions on their differentiation leading to excessive collagen deposition. In Aim 1, we will determine gene expression patterns in SMC and myofibroblasts in penile and vascular fibrosis. Male rats in groups: 1) "Young"; 2) "Aged"; 3) "PD-like plaque"; and 4) "Normal tunica", will receive in the corpora cavernosa or tunica albuginea an adenoviral collagen I promoter-green fluorescent protein (ColP-gfp) construct. Gfp be detected by dual fluorescence at 6 days in: a) SMC, and possibly myofibroblasts in the corpora and PDA, and: b) fibroblasts and myofibroblasts in the PDA adventitia or the tunica. Some of these cells will be dissected by laser capture microdissection (LCM). In Aim 2 we will determine the in vivo differentiation of penile fibroblasts and stem cells into SMC and myofibroblasts, and of myofibroblasts into SMC, active in collagen synthesis. Penile cultures from the rat tunica (fibroblasts) and the PD plaque (myofibroblasts) will be used as such, or after Sca1+ selection for stem cells, transfected with the ColP-gfp construct, and labeled with DAPI. Implantation will be for 1 week into: A: corpora cavernosa of old rats, with tunical: 1) fibroblasts; or 2) stem cells; and B: TGF¿1-induced PD-like plaque in the tunica, with: 3) tunical fibroblasts; 4) PD-myofibroblasts; or 5) tunical stem cells. End-points will be: A) LCM in tissue sections, with immunofluorescence for gfp, fibroblasts (vimentin), myofibroblasts/SMC (ASMA) and SMC (smoothelin), followed by DNA microarrays for gene expression profiles; B) fibrosis by immunohistochemistry/QIA (quantitative image analysis), RT/real time PCR, and western blots, and assays for hydroxyproline; B) for implanted DAPI+ cells: by QIA fluorescence/dual confocal microscopy for cell markers. Tissue fibrosis is an excessive deposition of collagen fibers, often accompanied by the loss of cellular mass, that occurs in most organs with aging, diabetes, injury, toxic insults, and other conditions, and that severely impairs the function of those organs and is a key factor in various diseases. Using rat animal models and cell cultures, we aim to clarify aspects that would help to define whether there is a common molecular and cellular profile of fibrosis, that in penile tissues causes Peyronie's disease (PD) and erectile dysfunction, and that in the arterial wall leads to arteriosclerosis. We will focus on studying the role of cells named fibroblasts, smooth muscle cells, and myofibroblasts, as well as adult stem cells, in these processes, and whether some of these cells can interconvert into each other. We expect that our studies would lead to the detection of novel targets of antifibrotic therapy for penile and vascular diseases of considerable public health impact.

DESCRIPTION (provided by applicable): This project aims to clarify some aspects that would help to define a common molecular profile of vascular fibrosis, that in the penis causes Peyronie's disease (PD) and aging-related vasculologic erectile dysfunction, and that in the arteries leads to aging-related arterosclerosis and arterial stiffness, in order to detect cellular and molecular targets of抗纤维化疗法。我们将研究：a）在阴茎和血管纤维化中发生的基因表达改变的细胞特异性，阐明了平滑肌细胞（SMC）和肌纤维细胞在这些过程中的作用； b）阴茎小梁组织中的SMC是否负责过度胶原蛋白合成； c）肌纤维细胞和SMC的个体关系，d）细胞相互作用对它们的分化的影响，导致过度胶原蛋白沉积。在AIM 1中，我们将确定SMC中的基因表达模式和阴茎和血管纤维化中的肌纤维细胞。小鼠组为：1）“年轻”； 2）“老化”； 3）“ PD状牌匾”； 4）“普通tunica”将在Corpora Cavernosa或Tinica Albuginea中获得腺病毒胶原I启动子绿色荧光蛋白（COLP-GFP）构建体。在6天内通过双荧光检测到GFP：a）SMC，以及Corpora和PDA中可能的成纤维细胞，以及：B）PDA Adventitia或Tunica中的成纤维细胞和肌纤维细胞。这些细胞中的一些将通过激光捕获显微解剖（LCM）解剖。在AIM 2中，我们将确定阴茎成纤维细胞和干细胞在SMC和肌纤维细胞中的体内分化，以及在胶原蛋白合成中活跃的SMC的肌纤维细胞。来自大鼠TUNICA（成纤维细胞）和PD斑块（肌纤维细胞）的阴茎培养物将其使用，或在SCA1+选择后用于干细胞，用COLP-GFP构建体转染，并用DAPI标记。植入将持续1周：A：老鼠的Corpora Cavernosa，带有孔：1）成纤维细胞；或2）干细胞；和b：TGF¿1诱导的Tunica中的PD样斑块，其中：3）纯成纤维细胞； 4）PD肌纤维细胞；或5）干细胞。终点将是：a）组织切片中的LCM，具有GFP的免疫荧光，成纤维细胞（Vimentin），肌纤维细胞/SMC/SMC（ASMA）和SMC（Smoolelin），然后是基因表达谱的DNA微阵列； b）免疫组织化学/QIA（定量图像分析），RT/实时PCR和Western印迹以及羟基丙烯的测定法； b）用于植入的DAPI+细胞：通过QIA荧光/双共聚焦显微镜用于细胞标记。组织纤维化是胶原蛋白纤维的过量沉积，通常伴随着细胞质量的损失，在大多数具有衰老，糖尿病，损伤，有毒感染和其他疾病的器官中发生，并且严重损害了这些器官的功能，并且是各种疾病中的关键因素。我们使用大鼠动物模型和细胞培养物，旨在阐明有助于定义纤维化的共同分子和细胞谱的方面，在阴茎时机中会导致Peyronie病（PD）和勃起功能障碍，并且在动脉壁中导致动脉粥样硬化。我们将专注于研究所谓的成纤维细胞，平滑肌细胞和肌纤维细胞以及成年干细胞的细胞在这些过程中的作用，以及其中一些细胞是否可以相互互连。我们预计我们的研究将导致发现对公共卫生影响的阴茎和血管疾病的新型抗纤维化治疗靶标。

项目成果

Myostatin genetic inactivation inhibits myogenesis by muscle-derived stem cells in vitro but not when implanted in the mdx mouse muscle.

• DOI: 10.1186/scrt152
• 发表时间: 2013-01-07
• 期刊: Stem cell research & therapy
• 影响因子: 7.5
• 作者: Tsao J;Vernet DA;Gelfand R;Kovanecz I;Nolazco G;Bruhn KW;Gonzalez-Cadavid NF
• 通讯作者: Gonzalez-Cadavid NF

Separate or combined treatments with daily sildenafil, molsidomine, or muscle-derived stem cells prevent erectile dysfunction in a rat model of cavernosal nerve damage.

每日使用西地那非、吗多明或肌肉来源的干细胞单独或联合治疗可预防海绵体神经损伤大鼠模型的勃起功能障碍。

• DOI: 10.1111/j.1743-6109.2012.02913.x
• 发表时间: 2012-11
• 期刊: The journal of sexual medicine
• 作者: Kovanecz I;Rivera S;Nolazco G;Vernet D;Segura D;Gharib S;Rajfer J;Gonzalez-Cadavid NF
• 通讯作者: Gonzalez-Cadavid NF

Prevalence of kidney stones in the United States.

• DOI: 10.1016/j.eururo.2012.03.052
• 发表时间: 2012-07
• 期刊: EUROPEAN UROLOGY
• 影响因子: 23.4
• 作者: Scales, Charles D., Jr.;Smith, Alexandria C.;Hanley, Janet M.;Saigal, Christopher S.
• 通讯作者: Saigal, Christopher S.