Regulated Expression of Truncated Short Variants of Caspase-12 mRNA in Human Retinal Pigment Epithelial Cells Suggests Their Immunomodulating Role

¹ Ophthalmology, University of Michigan, ann arbor, Michigan, United States
² Ophthalmology, University of Michigan, Ann Arbor, Michigan, United States
³ Department of Ophthalmology, W. K. Kellogg Eye Center, University of Michigan, 1000 Wall Street, Ann Arbor, Michigan, 48105, United States

^* To whom correspondence should be addressed. E-mail: velner{at}umich.edu.

	Abstract

PURPOSE. To investigate the expression and regulation of the short form of caspase-12, caspase-12S, in human retinal pigment epithelial (hRPE) cells. METHODS. hRPE cells were stimulated by pro-inflammatory agents IL-1 (2 ng/ml) and TNF- (20 ng/ml), LPS (1000 ng/ml), co-culture with monocytes, immunomodulating agent cyclosporine A (CSA, 30 ng/ml), and anti-inflammatory cytokine IL-10 (100 U/ml), and endoplasmic reticulum (ER) stress inducers tunicamycin (3 or 10 µM) and thapsigargin (25 or 100 nM) for 6 hr or longer. The total RNAs were isolated and subjected to semi-quantitative and quantitative real time RT-PCR analysis. Effects of tunicamycin and thapsigargin on IL-1- and TNF--stimulated MCP-1 mRNA expression and protein production were further examined by RT-PCR and ELISA, respectively. RESULTS. RT-PCR results showed that caspase-12S is the predominant form of caspase-12 in the examined hRPE cells of this study with expression at as high levels as in many other human tissues such as pancreas, prostate, small intestine, lung, spleen, and kidney. Treatment with IL-1 and TNF-, as well as LPS and co-culture with monocytes reduced hRPE caspase-12S mRNA expression within 6 hr. In contrast, hRPE exposure to cyclosporine A (CSA) and cytokine IL-10 for 6 hr increased caspase-12S mRNA expression. Compared to CSA and IL-10, ER stress activators tunicamycin and thapsigargin were even more potent enhancers of hRPE caspase-12S gene expression. Tunicamycin and thapsigargin also caused corresponding reductions in IL-1- and TNF--induced MCP-1 mRNA expression and protein production. CONCLUSION. hRPE cells express high level of caspase-12S. The regulated expression of caspase-12S suggests that the caspase recruitment domain (CARD)-only caspase-12S may be an endogenous dominant negative regulator modulating inflammatory responses in hRPE cells.

Key Words: retinal pigment epithelium, cytokine, immunoregulation, apoptosis

This article has been cited by other articles:

				Z.-M. Bian, S. G. Elner, and V. M. Elner Dual Involvement of Caspase-4 in Inflammatory and ER Stress-Induced Apoptotic Responses in Human Retinal Pigment Epithelial Cells Invest. Ophthalmol. Vis. Sci., December 1, 2009; 50(12): 6006 - 6014. [Abstract] [Full Text] [PDF]