Abstract
Purpose:
To evaluate whether direct downregulation of Cx43 expression promotes development of apoptosis in vascular cells of rat retinas.
Methods:
Non-diabetic control rats, diabetic rats, and rats intravitreally injected with Cx43 small interfering RNA (siRNA) or scrambled siRNA were used in this study to determine if acute downregulation of Cx43 expression in the retina contributes to retinal vascular cell death. Western blot (WB) analysis and Cx43 immunostaining were performed to assess Cx43 protein levels and its distribution in the vascular cells of retinal capillaries. Concurrently, retinal networks were subjected to TUNEL assay to assess the number of apoptotic cells.
Results:
WB analysis indicated a significant decrease in Cx43 protein level in retinas of diabetic rats (59 ± 4 % of N, n=4, p<0.01) and those intravitreally injected with Cx43 siRNA (67 ± 7 % of N, n=4, p<0.01) compared to retinas of control rats. Likewise, Cx43 immunostaining was significantly reduced in the vascular cells of the retinal capillaries in diabetic rats (63 ± 7 % of N, n=4, p<0.01) and in those of Cx43 siRNA-treated rats (68 ± 6 % of N, n=4, p<0.01). As expected, increased number of apoptotic cells was observed in retinal capillaries of diabetic rats compared to those of non-diabetic control rats(178 ± 48 % of N, n=4, p<0.05). Importantly, increased number of apoptotic cells was observed in retinal capillaries of rats intravitreally injected with Cx43 siRNA (167 ± 39% of N, n=4, p<0.05) compared to those of the control rats.
Conclusions:
Results indicate that downregulation of Cx43 expression alone is sufficient to induce vascular cell loss characteristic of diabetic retinopathy. These findings suggest that diabetes-induced downregulation of Cx43 participates in promoting retinal vascular lesions associated with diabetic retinopathy.
Keywords: 499 diabetic retinopathy •
688 retina •
426 apoptosis/cell death