Renyi Wu

Reduction of nitrite production by endothelin-1 in isolated porcine ciliary processes.

It has been postulated that endothelin-1 and nitric oxide (NO) could participate in the modulation of aqueous humor dynamic in the eye. This study investigates whether endothelin-1 can reduce the production of nitrite (a stable metabolite of NO) in isolated porcine ciliary processes. Nitrite production was measured (Griess reaction) in the medium surrounding isolated ciliary processes before and after exposure to different drugs. Results are expressed in percent of basal nitrite production. In a concentration-dependent manner (0.1 nM to 1 microM), endothelin-1 significantly decreased basal nitrite production (1 microM: 81.7+/-3.5%; P<0.001). This effect was prevented by the endothelin type A (ETA)-receptor antagonist BQ123 (1 microM: 95.6+/-4.3%; P<0.05), but not by the endothelin type B (ETB)-receptor antagonist BQ788 (1 microM: 83.3+/-4.4%; P=0.86) or by the prostaglandin F2alpha analog unoprostone (30 microM: 86.7+/-3.7%; P=0.74). The adenylcyclase activator forskolin significantly increased basal nitrite production (1 microM: 143.4+/-3.9%, P<0.001). This effect was prevented both by endothelin-1 (0.1 microM; 113.9+/-6.7%: P<0.01) or the nitric oxide synthase inhibitor L-NAME (0.5 mM: 103.3+/-8.4%: P<0.001). The inhibitory effect of endothelin-1 on forskolin-induced nitrite production was significantly reversed by BQ123 (1 microM: 132.4+/-5.1%; P<0.01), but not by BQ788 (1 microM: 112.7+/-4.1%; P=0.64) or unoprostone (30 microM: 109.3+/-4.8%; P=0.98). These results suggest that endothelin-1, through an ETA receptor activation, can reduce both basal and forskolin-induced nitrite production in isolated porcine ciliary processes.

Inhibition of calpain expression by E-64d in the rat retina subjected to ischemia/reperfusion injury

To investigate the effect of E-64d, a selective inhibitor of calpain, on the expression of calpain and calpastatin in rat retina was subjected to ischemia/reperfusion injury (IRI). An animal model of retinal IRI was set up by increasing the intraocular pressure (110 mm Hg) of a rat eye for 1 h. The retinal thickness and morphologic changes were detected by histology. The protein expression of m-calpain (a calpain isoform) in the retina was assessed by immunohistochemistry and Western blot assay. The mRNA of m-calpain, as well as calpastatin (an endogenous protein inhibitor of calpain), in the retina was assessed by RT-PCR, and the ratio of m-calpain/calpastatin was then calculated. To evaluate the effect of E-64d on the expression of calpain, the drug (5 μl of 100 μM) was injected intravitreously immediately after IRI. There were retinal edematous changes, particularly in the inner plexiform layer after IRI. The protein expression of m-calpain in the retina was increased 24 h after IRI, an effect that was inhibited by E-64d (P < 0.05). The mRNA expression of m-calpain and calpastatin was also increased 24 h and 3 h after IRI, respectively. Neither m-calpain nor calpastatin mRNA expression was influenced by E-64d (P > 0.05). The mRNA ratio of m-calpain to calpastatin was increased at the 6 h, 24 h and 72 h after IRI, and only at 24 h the increase of the ratio of m-calpain to calpastatin was inhibited by E-64d (P < 0.05). In the rat retina of IRI, E-64d inhibits the increase of m-calpain protein expression, as well as the mRNA ratio increase of m-calpain to calpastatin. E-64d also inhibited the retinal damage induced by IRI, suggesting a role for E-64d in the protection of the retinal apoptosis induced by IRI.