Entactin promotes adhesion and long-term maintenance of cultured regenerated skeletal myotubes.
The basal lamina protein, laminin, has been shown to promote migration and proliferation of cultured skeletal myoblasts, resulting in increased myotube formation. However, skeletal myotubes adhere poorly to a laminin substrate, and long-term cultures of skeletal myotubes on laminin have not been achieved. We have found that cultured satellite cells from bupivacaine-damaged rat skeletal muscle actively proliferate and differentiate on a diluted Matrigel substrate composed of laminin, type IV collagen, heparan sulfate proteoglycan, and entactin. Myotubes cultured on diluted Matrigel are contractile and have never been observed to detach from the culture dish; rather, myotubes generally atrophy after 2-3 weeks in culture. Antibodies directed against the various protein components of Matrigel were used to determine the role of each component in enhancing muscle differentiation. Anti-laminin impaired satellite cell adhesion, whereas antibodies against either type IV collagen or heparan sulfate proteoglycan had no effect. Anti-entactin did not inhibit attachment, proliferation, or fusion of cultured satellite cells; however, myotubes exposed to anti-entactin failed to adhere to the culture dish after spontaneous myotube contractions began. We conclude that entactin is responsible for long-term maintenance and maturation of contractile skeletal myotubes on a diluted Matrigel substrate. This is the first study to assign a biological function for entactin in myogenesis.
Funanage
V L
VL
Research Department, Alfred I. duPont Institute, Wilmington, Delaware 19899.
Smith
S M
SM
Minnich
M A
MA
D000818:Animals; D002448:Cell Adhesion; D002478:Cells, Cultured; D003094:Collagen; D003402:Creatine Kinase / Q000378:metabolism; D004338:Drug Combinations; D005109:Extracellular Matrix / Q000502:physiology; D005353:Fibronectins / Q000502:physiology; D007158:Immunologic Techniques; D066298:In Vitro Techniques; D007797:Laminin / Q000502:physiology; D008297:Male; D008562:Membrane Glycoproteins / Q000502:physiology*; D009132:Muscles / Q000166:cytology* / Q000502:physiology; D011509:Proteoglycans; D051381:Rats; D012038:Regeneration; D013997:Time Factors
D004338:Drug Combinations; D005353:Fibronectins; D007797:Laminin; D008562:Membrane Glycoproteins; D011509:Proteoglycans; C029967:nidogen; C059485:matrigel; D003094:Collagen; D003402:Creatine Kinase