ch7_in_vivo

ch7_in_vivo - Harvard-MIT Division of Health Sciences and...

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Unformatted text preview: Harvard-MIT Division of Health Sciences and Technology HST.523J: Cell-Matrix Mechanics Prof. Myron Spector Massachusetts Institute of Technology Harvard Medical School Brigham and Women's Hospital VA Boston Healthcare System 2.785j/3.97J/BEH.411/HST523J IN VIVO EXPRESSION OF -SMOOTH MUSCLE ACTIN AND CELL CONTRACTION M. Spector, Ph.D. TISSUE CLASSIFICATION Connective Tissue Synthesize and maintain a structurally competent ECM (including a supporting and connecting framework for all other tissue types); matrix and cell continuous Muscle Cells Contraction; cell continuous, BM Epithelia Lining and secretory cells; cell continuous, BM Nerve Voltage conduction; cell continuous, BM FORCES GENERATED BY CELLS All Cells Actin Isoforms Migration - and - cytoplasmic Maintain cell shape - and - cytoplasmic Muscle Cells Contraction -smooth muscle (vascular) -smooth muscle (enteric) -skeletal muscle -cardiac muscle TISSUE CLASSIFICATION Connective Tissue Cells Muscle Cells (contractile cells) -skeletal actin skeletal cardiac -cardiac actin smooth muscle - and -smooth muscle actin Epithelial Cells Nerve Cells TISSUE CLASSIFICATION Connective Tissue Cells "myofibroblasts" (-SMA; contractile cells) Muscle Cells (contractile cells) -skeletal actin skeletal cardiac -cardiac actin smooth muscle - and -smooth muscle actin Epithelial Cells Nerve Cells -Smooth Muscle Actin-Containing Fibroblasts Myofibroblasts (day 10) Photo removed for copyright reasons. Ungrafted Photo removed for copyright reasons. Grafted IV Yannas, et al. Summary: mechanism It has been demonstrated by other investigators that wound contraction in connective tissues is caused by the cooperative pulling force offibroblasts that adopt a contractile phenotype and express an isofromof the protein actinfound in smooth muscle cells (alpha-SM actin). These cells have been termed myofibroblasts. The image on the top shows an ungraftedskin wound that has been stained with an antibody for alpha-SM actin, red indicates positive stain. They are oriented parallel across the wound bed. In this particular configuration, the wound edges are moving together in this direction across the screen. MFB form a cell-continuous network and are able to transmit the force across the wound. In the grafted wound at the bottom, MFB are present, but due tothe random pore walls of the matrix, they are not able to form a continuousaligned network across the wound, and contraction does not take place. Once again, this inhibition of contraction does not happen if the pore size and contact surface are not right and if the chemistry for cell attachment and pulling is not right. The interruption of the MFB network is the proposed mechanism ofaction of the ECM analog in preventing contraction. Maybe also a statementabout MFB imparting the alignment of collagen in scar. CONNECTIVE TISSUE CELLS THAT CAN EXPRESS -SMOOTH MUSCLE ACTIN Articular chondrocyte Osteoblast Meniscus fibroblast and fibrochondrocyte Intervertebral disc fibroblast and fibrochondrocyte Ligament fibroblast Tendon fibroblast Synovial cell Mesenchymal stem cell M. Spector, Wound Repair Regen. 9:11-18(2001) CONTRACTILE CONNECTIVE TISSUE CELLS Express SMA in vivo Capable of contracting collagen-GAG matrices in vitro SMA-positive cells retain differentiated phenotype SMA trait derived from the stem cell Amount of contraction correlated with the SMA content SMA and contraction up-regulated by TGF-1 Roles have yet to be determined, but may be both positive and negative Canine Articular Cartilage Photo removed for copyright reasons. Photo removed for copyright reasons. Normal top zone; 50 7% SMA Neg. Control 9-wk repair Photo removed for copyright reasons. Photo removed for copyright reasons. Normal basal zone; 23 5% Q. Wang, et al., Wound Rep. Regen., 2000;8:145-158 Human Articular Cartilage Kim and Spector, JOR 2000;18:749-755 Photos removed for copyright reasons. Neg. control POSSIBLE ROLES FOR SMA-ENABLED CONTRACTION OF MS CELLS Closure of wounds Tensioning of a healing ligament Retraction of the ends of torn ligaments/tendons that do not heal Disease processes Contracture Tissue formation Modeling of ECM architecture and remodeling (e.g., crimp in ligament/tendon?) Tissue engineering Contracture of scaffolds Healing -smooth muscle actin in fibroblasts in the healing rabbit collateral ligament 10 wks Photo and diagram removed for copyright reasons. Faryniarz, Chaponnier, Gabbiani, Yannas, and Spector; JOR, 14:228 (1996) Cells containing SMA (reddishbrown stain) 10% cells SMA+ Myofibroblasts in the Healing Rabbit Medial Collateral Ligament (10 wks post-rupture) Faryniarz, Chaponnier, Gabbiani, Yannas, and Spector; JOR, 14:228 (1996) Photo removed for copyright reasons. Smooth muscle actin Myofibroblasts draw the ruptured ends together and tension the ligament. Photo removed for copyright reasons. SMA-containing cells in the intact human ACL SMA (red) Up to 50% cells SMA+ Photos removed for copyright reasons. Neg. Control; no SMA antibody MM Murray, et al., JOR, 1999;17:18-27 Histologic Changes in the Human ACL after Rupture Diagram removed for copyright reasons. A. Inflammation B. Epiligamentous Regeneration SMA-expressing cells "Retraction" D. Remodeling M. Meaney Murray, et al., J. Bone Jt. Surg., 2000;82-A:1387 C. Proliferation Ruptured Human Anterior Cruciate Ligaments Blood Vessel Evidence supporting the hypothesis that SMA-enabled contraction is responsible for retraction of the ruptured ends. Photo removed for copyright reasons. Crimped morphology of SMA-containing (red) cells consistent with contraction. Imparting crimp to matrix? Photo removed for copyright reasons. M. Meaney Murray, et al. J. Bone Jt. Surg., 2000;82-A:1387 Ruptured Human Rotator Cuff Photos removed for copyright reasons. Is SMA-enabled contraction responsible for retraction of the ruptured ends? J. Premdas, et al. JOR, 2001;19:221-228 Osteoblasts Expressing SMA Canine trabecular bone Photo removed for copyright reasons. C. Menard, et al., Biomat. 2000;21:1867 Human trabecular bone B. Kinner, et al. JOR 2002;20:622 Photo removed for copyright reasons. Graph of %SMA+ vs. Patient Age removed for copyright reasons. B. Kinner, et al. JOR 2002;20:622 Osteoblasts Expressing SMA in Human Bone Explants 6 wks Photo removed for copyright reasons. B. Kinner, et al. JOR 2002;20:622 Osteoblastic cells (MC3T3-E1) contracting a collagen-GAG matrix Pores compressed as specimens decrease in size (no evident dissolution) Photo removed for copyright reasons. 1 wk Loss of SMA Photo removed for copyright reasons. Photo removed for copyright reasons. 2 wk 4 wk C. Menard, et al., Biomat. 2000;21:1867 Mouse Tibia (Closed) Fracture Model B. Kinner, et al., Bone 2002;30:738 Photos removed for copyright reasons. 3 weeks post-fracture Mouse Tibia (Closed) Fracture Model SMA immunohistochemistry 3 weeks post-fracture Photos removed for copyright reasons. Neg. control B. Kinner, et al., Bone 2002;30:738 Distraction Osteogenesis; Rat Model Photo removed for copyright reasons. 2 latent+13 distraction +3 consolidation (days) B. Kinner, et al. JOR 2003;21:20 Distraction Osteogenesis; Rat Model Photo removed for copyright reasons. 2 latent+10 distraction (days) B. Kinner, et al. JOR 2003;21:20 SMA AND CONTRACTION OF MUSCULOSKLETAL CELLS Many Questions to be Answered What are the roles of SMA-enabled contraction in normal and pathological processes? What therapeutic approaches can be taken for its regulation? How does the SMA-enabled contraction impact musculoskeletal tissue engineering? TISSUE CLASSIFICATION Connective Tissue Synthesize and maintain a structurally competent ECM for all tissue types Employ SMA-enabled contraction to model the ECM and to close wounds Muscle Cells Epithelia Nerve ...
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This note was uploaded on 11/11/2011 for the course BIO 20.410j taught by Professor Rogerd.kamm during the Spring '03 term at MIT.

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