Altered Matrix-Cells and Intermolecular Interactions

改变的基质细胞和分子间相互作用

• 批准号: 7807035
• 负责人: ANDRZEJ FERTALA
• 金额: $ 32.35万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-20 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7807035
• 关键词: Address; Affect; Biological; Biomechanics; Cartilage; Cell Culture Techniques; Cell Therapy; Cells; Characteristics; Chondrocytes; Collagen; Collagen Gene; Complementary DNA; Connective Tissue; Disease; Dominant-Negative Mutation; Engineering; Experimental Models; Extracellular Matrix; Fibrillar Collagen; Genes; In Vitro; Modeling; Mus; Mutation; Natural regeneration; Nude Mice; Patients; Phenotype; Procollagen; Public Health; Publishing; Research; Simulate; Structure; System; Testing; Therapeutic; Therapeutic Agents; Tissues; Type II Procollagen; Variant; base; cell behavior; cellular engineering; chondrodysplasia; design; disease phenotype; disease-causing mutation; gene therapy; intermolecular interaction; macromolecule; meetings; mutant; repaired; research study; response; skeletal tissue; therapy development

DESCRIPTION (provided by applicant): Research we propose here is a continuation of our current study on the effects of mutations in fibrillar collagens on the structure of extracellular matrices and behavior of cells with special emphasis on mutations in collagen II and their effects on cartilage. The broad, long-term objective of the proposed study is to employ three-dimensional cartilage-like constructs to determine effects of the presence of mutant collagen molecules on the structure of ECM, cell behavior, and to define target parameters for cell and gene therapies that should be reached to change abnormal phenotypes developed in the presence of collagen mutants. Our hypotheses are that mutations in fibrillar collagens alter not only the structure of extracellular matrices, but also impact the behavior of cells, and that requirements for therapy approaches to be successful vary for different collagen mutants. To meet our long-term objectives and test stated hypotheses we formulated the following Specific Aims: (1) To create a biologically relevant experimental model to study effects of collagen mutants on degeneration and repair of affected tissues, (2) To determine effects of suppression of expression of collagen mutants on remodeling of abnormal connective tissue, (3) To define conditions for cell therapies needed to override pathological changes caused by collagen mutants. A fundamental barrier to move forward development of therapies for dominant negative effects of mutations in collagen genes is a lack of information about minimal conditions needed to be achieved to drive affected tissues toward their remodeling into normal structures in response to cell or gene therapies. In our studies we will address this problem by creating a biologically relevant model, which will resemble the complexity of cartilage. This model will consist of engineered cells that, in addition to endogenous wild type procollagen II will conditionally express cDNA constructs encoding procollagen II mutants found in patients with various forms of chondrodysplasias. These cells will be employed to create cartilage-like constructs in cell culture conditions and in athymic nude mice. Subsequently, changes in morphological, biological, and biomechanical characteristics of the cartilage-like constructs formed in the presence of mutant collagen II variants will be studied after switching off mutant cDNAs or after experiments simulating "delivery" of cells expressing wild type procollagen II. By employing models for gene and cell therapies, studies we propose will determine intrinsic capacity of cartilaginous tissues to repair and regenerate. Moreover, these studies will provide important information for designing therapeutic approaches to counterbalance not only dominant negative effects of mutations in collagen II but also, most likely, effects of mutations in other collagenous and noncollagenous extracellular matrix macromolecules that are associated with heritable diseases of skeletal tissues. Thus, the relevance of the proposed study to public health is high.

描述（由申请人提供）：我们在此提出的研究是我们目前研究的继续，该研究涉及纤维胶原突变对细胞外基质结构和细胞行为的影响，特别强调胶原II突变及其对软骨的影响。拟议研究的广泛，长期目标是采用三维软骨样结构来确定突变胶原分子的存在对ECM结构，细胞行为的影响，并定义细胞和基因治疗的目标参数，这些参数应达到改变胶原突变体存在下产生的异常表型。我们的假设是，纤维状胶原蛋白的突变不仅改变细胞外基质的结构，而且影响细胞的行为，并且对于不同的胶原蛋白突变体，成功的治疗方法的要求不同。为了实现我们的长期目标和测试所陈述的假设，我们制定了以下具体目标：（1）建立生物学相关的实验模型以研究胶原突变体对受影响组织的变性和修复的影响，（2）确定抑制胶原突变体的表达对异常结缔组织的重塑的影响，（3）确定细胞治疗所需的条件，以克服由胶原突变体引起的病理变化。推进开发针对胶原基因突变的显性负效应的疗法的基本障碍是缺乏关于需要实现的最小条件的信息，以驱动受影响的组织响应于细胞或基因疗法而重塑成正常结构。在我们的研究中，我们将通过创建一个生物学相关模型来解决这个问题，该模型类似于软骨的复杂性。该模型将由工程化细胞组成，除了内源性野生型II型前胶原外，所述工程化细胞将条件性表达编码在患有各种形式软骨发育不良的患者中发现的II型前胶原突变体的cDNA构建体。这些细胞将用于在细胞培养条件下和在无胸腺裸鼠中产生软骨样构建体。随后，在关闭突变cDNA后或在模拟表达野生型II型前胶原的细胞的“递送”实验后，将研究在存在突变型II型胶原变体的情况下形成的软骨样结构的形态学、生物学和生物力学特性的变化。通过采用基因和细胞治疗模型，我们提出的研究将确定软骨组织修复和再生的内在能力。此外，这些研究将提供重要的信息，设计治疗方法，以抵消不仅在II型胶原蛋白的突变，而且，最有可能的是，在其他胶原和非胶原细胞外基质大分子与骨骼组织的遗传性疾病相关的突变的影响，显性的负面影响。因此，拟议的研究与公共卫生的相关性很高。

项目成果

Distinct Cell Stress Responses Induced by ATP Restriction in Quiescent Human Fibroblasts.

静止人类成纤维细胞中 ATP 限制诱导的独特细胞应激反应。

• DOI: 10.3389/fgene.2016.00171
• 发表时间: 2016
• 期刊: Frontiers in genetics
• 影响因子: 3.7
• 作者: Yalamanchili,Nirupama;Kriete,Andres;Alfego,David;Danowski,KelliM;Kari,Csaba;Rodeck,Ulrich
• 通讯作者: Rodeck,Ulrich

Reducing the effects of intracellular accumulation of thermolabile collagen II mutants by increasing their thermostability in cell culture conditions.

• DOI: 10.1016/j.bbrc.2010.04.056
• 发表时间: 2010-05-28
• 期刊: BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
• 影响因子: 3.1
• 作者: Gawron, Katarzyna;Jensen, Deborah A.;Steplewski, Andrzej;Fertala, Andrzej
• 通讯作者: Fertala, Andrzej

Endostatin binds nerve growth factor and thereby inhibits neurite outgrowth and neuronal migration in-vitro.

• DOI: 10.1016/j.brainres.2010.09.023
• 发表时间: 2010-11-11
• 期刊: Brain research
• 影响因子: 2.9
• 作者: Al Ahmad A;Lee B;Stack J;Parham C;Campbell J;Clarke D;Fertala A;Bix GJ
• 通讯作者: Bix GJ

Persistence of intracellular and extracellular changes after incompletely suppressing expression of the R789C (p.R989C) and R992C (p.R1192C) collagen II mutants.

• DOI: 10.1002/humu.21506
• 发表时间: 2011-07
• 期刊: HUMAN MUTATION
• 影响因子: 3.9
• 作者: Jensen, Deborah A.;Steplewski, Andrzej;Gawron, Katarzyna;Fertala, Andrzej
• 通讯作者: Fertala, Andrzej

R992C (p.R1192C) Substitution in collagen II alters the structure of mutant molecules and induces the unfolded protein response.

• DOI: 10.1016/j.jmb.2009.05.004
• 发表时间: 2009-07-10
• 期刊: JOURNAL OF MOLECULAR BIOLOGY
• 影响因子: 5.6
• 作者: Chung, Hye Jin;Jensen, Deborah A.;Gawron, Katarzyna;Steplewski, Andrzej;Fertala, Andrzej
• 通讯作者: Fertala, Andrzej