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Mary M. Feng, Hong Liu, Cindy Hutnik; Protective Effect of Connexin43 in Human Trabecular Meshwork Cells Exposed to Mechanical Stretch. Invest. Ophthalmol. Vis. Sci. 2012;53(14):3246.
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The purpose of this study was to determine the effect of mechanical stress on human trabecular meshwork cells (HTMC) and to study the role of connexin43 (Cx43) in HTMC exposed to mechanical strain.
Primary HTMC were cultured on collagen I coated Flexcell plates for 5 days in 10% FBS and 1%P/S until 90% confluent, then switched to serum free medium immediately prior to mechanical stress. In addition, HTMC were infected with retrovirus encoding empty vector (V) and vectors encoding Cx43 and a disease-linked dominant negative Cx43 mutant (G138R). HTMC were exposed to 15% stretch for 48 hours at 1 Hz using the FX-5000 Tension system. LDH release was measured using the Cytotoxicity Detection Kit and levels of cytosolic DNA fragments were determined to assess apoptosis. Cx43 mRNA and protein levels were measured using real-time PCR and Western Blot respectively.
Mechanical stretch increased LDH release from HTMC by 42% (p<0.05). In addition, stretching increased HTMC apoptosis by 19% (p<0.05). Cx43 mRNA (p<0.01) and protein expression (p<0.01) decreased significantly after mechanical strain. Cx43 overexpression by retroviral infection reversed the cell injury caused by mechanical stretch, as seen by a significant reduction in LDH release (p<0.05).
Mechanical stretch induces cell injury and apoptosis, and decreases Cx43 expression in HTMC. Overexpression of Cx43 protects HTMC from mechanical stress.
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