Regulation of Skeletal Growth by Soft Tissue Extracellular Matrix

软组织细胞外基质对骨骼生长的调节

基本信息

批准号：
10437366
负责人：
Dirk Hubmacher
金额：
$ 8.56万
依托单位：
ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-03-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10437366
关键词：
ADAMTS Address Affect Bees Binding Binding Proteins Biochemical Biological Biological Assay Biomechanics Birth Bone Growth Cartilage Cell Culture Techniques Cells Chondrocytes Chondrogenesis Connective Tissue Data Digit structure Disease Distal Dysplasia Epiphysial cartilage Etiology Extracellular Matrix Extracellular Matrix Proteins FBN1 Fibrillin Microfibrils Fibroblasts Functional disorder Gene Dosage Gene Expression Gene Expression Profiling Gene Mutation Genes Genetic Genetic Diseases Genetic Transcription Glycoproteins Growth Histologic Human Human Genetics Impairment Individual Joint Laxity Joints Knockout Mice LTBP2 gene Lead Length Ligaments Limb Bud Limb structure MADH4 gene Maps Marfan Syndrome Mechanics Mediating Mendelian disorder Mesenchyme Messenger RNA Microfibrils Modeling Molecular Mus Musculoskeletal Mutation Pathway interactions Peptide Hydrolases Phenotype Physiological Property Protein Isoforms Proteins Recombinants Regulation Reporting Roentgen Rays Role Signal Transduction Site Skeletal Muscle Structure Syndrome System Tendon structure Testing Tissues Transcriptional Regulation Transforming Growth Factors Translating Weill-Marchesani syndrome Western Blotting X-Ray Computed Tomography achilles tendon bone fibrillin fibrillin-2 geleophysic dysplasia insight joint stiffness microCT mouse model novel novel therapeutic intervention postnatal prenatal protein protein interaction regenerative approach skeletal soft tissue

项目摘要

ABSTRACT While it is undisputed that extracellular matrix and soft tissues influence skeletal growth, few specific pathways explaining this effect have been uncovered. In Marfan syndrome, which affects 1-2 in 5000 individuals, skeletal overgrowth, long digits, poorly developed musculature and lax joints result from fibrillin-1 (FBN1) mutations. Strikingly, specific mutations in FBN1 can also cause the “opposite” of Marfan syndrome, i.e. short stature, disproportionally short digits (brachydactyly), stiff joints, and a “pseudomuscular” build, which are the hallmarks of acromelic dysplasias, comprising a group of Mendelian disorders. Identical acromelic dysplasias can also be caused by genes encoding ADAMTS proteases, ADAMTS-like (ADAMTSL) proteins, latent transforming growth factor- (TGF) binding protein-3 (LTBP3), and SMAD4. The overlapping phenotypes of different gene mutations underlying acromelic dysplasias strongly support my hypothesis that a fibrillin-ADAMTS-TGF axis constitutes a novel extracellular matrix (ECM) network regulating postnatal limb growth. Mutations in ADAMTSL2 or FBN1 lead to geleophysic dysplasia (GD), a severe, frequently lethal human acromelic dysplasia. ADAMTSL2 is a secreted glycoprotein that binds to FBN1 and FBN2, and is implicated in TGF signaling. Intriguingly, ADAMTSL2 mRNA is not expressed in growth plate cartilage or bone, but has its strongest expression in tendon. My studies show that FBN2 microfibrils are increased at sites of Adamtsl2 expression in a mouse knockout model of GD, suggesting a role for ADAMTSL2 in switching from prenatal FBN2-dominated microfibrils to postnatal FBN1-dominated microfibrils. Furthermore, skeletal growth is impaired upon limb-specific ADAMTSL2 deletion (Prx1-Cre) or tendon and ligament specific deletion (Scx- Cre). I observed disproportionate distal limb shortening (i.e. acromelic dysplasia) and a reduction in Achilles tendon length in both conditional deletions. A model for skeletal growth in geleophysic dysplasia provides an opportunity to determine how tissue non-autonomous regulation of skeletal growth occurs via mechanical or regulatory input from tendon ECM. In aim 1, I will test the hypothesis by analyzing postnatal limb growth and ECM alterations in the microfibril system after Adamtsl2 deletion in tendons with Scx-Cre. In aim 2, I will investigate how ADAMTSL2 interacts with FBN1 and FBN2 and how ADAMTSL2 executes its role in the isoform switch from FBN2 to FBN1. I will analyze the genetic interaction of Adamtsl2 with Fbn1 and Fbn2 in mice and I will use protein-protein interaction studies and cell culture assays to gain mechanistic insights in the function of ADAMTSL2 in regulating the fibrillin isoform switch. Impact: The anticipated results will provide novel insights into the pathophysiology of acromelic dysplasias and other fibrillinopathies. These insights could be translated for targeting tendon ECM in regenerative strategies. The proposal addresses fundamental questions of how functional properties of soft tissues are determined by ECM subsequently might regulate postnatal limb growth.

抽象的虽然毫无疑问是细胞外基质和软组织会影响骨骼生长，但很少有特定途径解释了这种效果。在影响5000个人中有1-2位的马凡氏综合症中，骨骼过度生长，数字长，发育不良的肌肉和松弛关节是由纤维蛋白-1（FBN1）突变引起的。令人惊讶的是，FBN1中的特定突变也可能引起Marfan综合征的“相反”，即短统计数据，数字不成比例的短数（腕足），僵硬的接头和“假肌肉”的构建，这是标志雅亲域的发育不良，完成了一群孟德尔疾病。相同的雅亲骨发育不良也可以是由编码ADAMTS蛋白，ADAMTS样（ADAMTSL）蛋白的基因引起的，潜在转化生长因子-（TGF）结合蛋白3（LTBP3）和SMAD4。不同基因的重叠表型雅亲骨异常增生的突变强烈支持我的假设，即纤维蛋白-Adamts-TGF轴构成了一种新型的细胞外基质（ECM）网络调节产后肢体生长。 ADAMTSL2或FBN1突变导致地球物理发育不良（GD），这是一种严重的，经常致命的人雅亲粒发育不良。 ADAMTSL2是与FBN1和FBN2结合的分泌的糖蛋白 TGF信号传导。有趣的是，ADAMTSL2 mRNA在生长板软骨或骨骼中未表达，但具有肌腱中的强烈表达。我的研究表明，ADAMTSL2部位的FBN2微纤维增加在GD的小鼠敲除模型中的表达，这表明ADAMTSL2在从产前切换中的作用 FBN2主导的微纤维到产后FBN1主导的微纤维。此外，骨骼生长是肢体特异性ADAMTSL2缺失（PRX1-CRE）或肌腱和韧带特异性缺失（SCX-）受到损害（SCX- CRE）。我观察到不成比例的远端肢体缩短（即雅亲骨发育不良）和阿喀琉斯的减少两种条件缺失的肌腱长度。地球物理发育不良的骨骼生长模型提供了一种确定组织非自治调节骨骼生长是如何通过机械或肌腱ECM的调节输入。在AIM 1中，我将通过分析的产后肢体生长和 ADAMTSL2缺失后，具有SCX-CRE的肌腱中的微纤维系统的ECM改变。在AIM 2中，我会研究ADAMTSL2如何与FBN1和FBN2相互作用，以及Adamtsl2如何在同工型从FBN2转换为FBN1。我将分析ADAMTSL2与FBN1和FBN2的遗传相互作用小鼠和我将使用蛋白质 - 蛋白质相互作用研究和细胞培养分析来获得机械洞察力 ADAMTSL2在确定纤维蛋白同工型开关中的功能。影响：预期的结果将提供有关雅亲粒肿瘤和杂色生理生理学的新见解。其他纤维纤维。这些见解可以转化为针对再生策略的肌腱ECM。该提案解决了软组织功能特性如何决定的基本问题 ECM随后可能调节产后肢体生长。

项目成果

期刊论文数量（12）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Cell-Based Interaction Analysis of ADAMTS Proteases and ADAMTS-Like Proteins with Fibrillin Microfibrils.

ADAMTS 蛋白酶和 ADAMTS 样蛋白与原纤维蛋白微纤维的基于细胞的相互作用分析。

DOI：
10.1007/978-1-4939-9698-8_16
发表时间：
2020
期刊：
Methods in molecular biology (Clifton, N.J.)
影响因子：
0
作者：
Hubmacher,Dirk
通讯作者：
Hubmacher,Dirk

The quest for substrates and binding partners: A critical barrier for understanding the role of ADAMTS proteases in musculoskeletal development and disease.

DOI：
10.1002/dvdy.248
发表时间：
2021-01
期刊：
Developmental dynamics : an official publication of the American Association of Anatomists
影响因子：
0
作者：
Satz-Jacobowitz B;Hubmacher D
通讯作者：
Hubmacher D

Stable Knockdown of Genes Encoding Extracellular Matrix Proteins in the C2C12 Myoblast Cell Line Using Small-Hairpin (sh)RNA.

DOI：
10.3791/60824
发表时间：
2020-02-12
期刊：
Journal of visualized experiments : JoVE
影响因子：
0
作者：
Taye N;Stanley S;Hubmacher D
通讯作者：
Hubmacher D

Alternative splicing of the metalloprotease ADAMTS17 spacer regulates secretion and modulates autoproteolytic activity.

DOI：
10.1096/fj.202001120rr
发表时间：
2021-03
期刊：
FASEB journal : official publication of the Federation of American Societies for Experimental Biology
影响因子：
0
作者：
Balic Z;Misra S;Willard B;Reinhardt DP;Apte SS;Hubmacher D
通讯作者：
Hubmacher D

Secreted ADAMTS-like 2 promotes myoblast differentiation by potentiating WNT signaling.

分泌的 ADAMTS-like 2 通过增强 WNT 信号传导促进成肌细胞分化。