Kumar Sandeep, Shah Shaily, Tang Hai Michael, Smith Matthew, Borrás Teresa, Danias John
Department of Cell Biology, SUNY Downstate Medical Center, Brooklyn, New York, United States of America.
PLoS One. 2013 Aug 19;8(8):e72447. doi: 10.1371/journal.pone.0072447. eCollection 2013.
Tissue plasminogen activator, a serine protease encoded by the PLAT gene is present in the trabecular meshwork (TM) and other ocular tissues and has been reported to be downregulated by treatment with steroids in vitro. Steroids are known to cause changes in outflow facility of aqueous humor in many species. In the present study, we tested whether overexpression of PLAT can prevent and/or reverse the outflow facility of mouse eyes treated with steroids. Animals received bilateral injection with 20 µl of triamcinolone acetonide (TA) (40 mg/ml) suspension subconjunctivally to induce outflow facility changes. Some animals received unilateral intracameral injection with 2 µl of adenoviral suspension [3-4 x 10(12) virus genomes per milliliter (vg/ml)] carrying sheep PLAT cDNA (AdPLAT) either concurrently with TA injection or one week after TA injection, whereas others received bilateral intracameral injection with 2 µl of adenoviral suspension (9 x 10(12) vg/ml) carrying no transgene (AdNull) concurrently with TA injection. Animals were sacrificed one week after AdPLAT or AdNull treatment. Endogenous mRNA expression levels of mouse PAI-1 and MMP-2, -9 and -13 were also measured using qRT-PCR. Outflow facility one week after AdPLAT administration was increased by 60% and 63% respectively for animals that had not or had been pretreated with steroids. Overexpression of PLAT significantly upregulated expression of PAI-1, MMP-2, -9 and -13 compared to the levels found in TA only treated eyes. These findings suggest that overexpression of PLAT in TM of mouse eyes can both prevent and reverse the decrease in outflow facility caused by steroid treatment and is associated with upregulation of MMPs.
组织型纤溶酶原激活剂是一种由PLAT基因编码的丝氨酸蛋白酶，存在于小梁网（TM）和其他眼组织中，并且据报道在体外经类固醇处理后其表达会下调。已知类固醇会导致许多物种房水流出率发生变化。在本研究中，我们测试了PLAT的过表达是否能够预防和/或逆转经类固醇处理的小鼠眼睛的流出率变化。动物接受双侧结膜下注射20 μl曲安奈德（TA）（40 mg/ml）悬浮液以诱导流出率变化。一些动物在注射TA的同时或在注射TA一周后接受单侧前房内注射2 μl携带绵羊PLAT cDNA的腺病毒悬浮液[每毫升3 - 4×10¹²病毒基因组（vg/ml）]（AdPLAT），而其他动物在注射TA的同时接受双侧前房内注射2 μl不携带转基因的腺病毒悬浮液（9×10¹² vg/ml）（AdNull）。在AdPLAT或AdNull处理一周后处死动物。还使用qRT-PCR测量了小鼠PAI-1和MMP-2、-9及-13的内源性mRNA表达水平。对于未接受类固醇预处理或已接受类固醇预处理的动物，AdPLAT给药一周后的流出率分别增加了60%和63%。与仅接受TA处理的眼睛相比，PLAT的过表达显著上调了PAI-1、MMP-2、-9和-13的表达。这些发现表明，小鼠眼睛小梁网中PLAT的过表达既能预防又能逆转类固醇处理引起的流出率降低，并且与基质金属蛋白酶的上调有关。
