mRNA Expression Profiles of Tissue Inhibitors of Matrix Metalloproteinases in Cultured Human Trabecular Meshwork and Ciliary Body in Response to Latanoprost Treatment

Abstract

Abstract: : Purpose: To determine the expression pattern of tissue inhibitors of matrix metalloproteinases (TIMPs) in human trabecular meshwork (TM) and ciliary body (CB) and to assess changes in mRNA expression of these TIMPs in response to incubation with latanoprost. Methods: The presence or absence of TIMPs was examined, using RT–PCR, in both TM and CB tissues and cultures of human TM endothelial and CB smooth muscle (CBSM) cells. The PCR products were sequenced and compared against the Genbank database. Changes in mRNA expression of TIMPs to latanoprost were assessed using quantitative RT–PCR in control and latanoprost–treated cultures of TM endothelial and CBSM cells. The cultures were incubated with 30 ng/µl, 300 ng/µl, and 3 µg/µl of latanoprost for 24 hours. The comparative C_T method was used to quantify target genes relative to GAPDH as a reference. Results: All four known TIMPs were detected in both tissues and cultures of TM endothelial and CBSM cells. Relative levels of expression in cultures in TM were TIMP–1≥TIMP–2=TIMP–3>>TIMP–4, and in CBSM cells were TIMP–2>TIMP–1>TIMP–3 >>TIMP–4. In response to latanoprost incubation at therapeutic level (30 ng/µl), cultures of TM showed that TIMP–2 and TIMP–3 were up–regulated, while TIMP–1 was not changed. In CBSM cultures, TIMP–3 was up–regulated, while TIMP–4 was down–regulated; TIMP–1 and TIMP–2 showed no changes. Variability among four individuals was noted. Conclusions: All TIMPs were detected in both tissues and cultures of TM endothelial and CBSM cells. Latanoprost induced increases in mRNA expressions of TIMP–2 and TIMP–3 in TM endothelial cells, while an increase in mRNA expression of TIMP–3 in CBSM cells. Increases in these TIMPs may be important in the modulation of extracellular matrix by playing a compensatory/inhibitory role against uncontrolled matrix metalloproteinase (MMP) activity. As with MMPs, there was a tissue–specific response to latanoprost incubation for TIMPs induction.

Keywords: outflow: trabecular meshwork • gene/expression • outflow: ciliary muscle