Johnson Gretchen A, Krishnamoorthy Raghu R, Nagaraj Ram H, Stankowska Dorota L
North Texas Eye Research Institute, College of Biomedical and Translational Sciences, University of North Texas Health Science Center, Fort Worth, TX 76107, USA.
Department of Microbiology, Immunology, and Genetics, College of Biomedical and Translational Sciences, University of North Texas Health Science Center, Fort Worth, TX 76107, USA.
Biomolecules. 2025 Feb 3;15(2):219. doi: 10.3390/biom15020219.
This study evaluated the neuroprotective potential of peptain-1 conjugated to a cell-penetrating peptide (CPP-P1) in an ocular hypertension model of glaucoma. Brown Norway (BN) rats were subjected to intraocular pressure (IOP) elevation via intracameral injection of silicone oil (SO), with concurrent intravitreal injections of either CPP-P1 or a vehicle. Retinal cross-sections were analyzed for markers of neuroprotection, including cAMP response element-binding protein (CREB), phosphorylated CREB (p-CREB), growth-associated protein-43 (GAP43), synapsin-1 (SYN1), and superoxide dismutase 2 (SOD2). Hematoxylin and eosin staining was used to assess retinal-layer thickness. SO-treated rats exhibited significant reductions in the thickness of the inner nuclear layer (INL, 41%, = 0.016), inner plexiform layer (IPL, 52%, = 0.0002), and ganglion cell layer (GCL, 57%, = 0.001). CPP-P1 treatment mitigated these reductions, preserving INL thickness by 32% ( = 0.059), IPL by 19% ( = 0.119), and GCL by 31% ( = 0.057). Increased levels of CREB ( = 0.17) and p-CREB ( = 0.04) were observed in IOP-elevated, CPP-P1-treated retinas compared to IOP-elevated, vehicle-treated retinas. Although overall GAP43 levels were low, there was a modest increase in expression within the IPL and GCL in SO- and CPP-P1-treated retinas ( = 0.15 and = 0.09, respectively) compared to SO- and vehicle-treated retinas. SO injection reduced SYN1 expression in both IPL and GCL ( = 0.01), whereas CPP-P1 treatment significantly increased SYN1 levels in the IPL ( = 0.03) and GCL ( = 0.002). While SOD2 expression in the GCL was minimal across all groups, a trend toward increased expression was observed in CPP-P1-treated animals ( = 0.16). The SO model was replicated with SO removal after 7 days and monitored for 21 days followed by retinal flat-mount preparation to assess retinal ganglion cell (RGC) survival. A 42% loss in RGCs ( = 0.009) was observed in SO-injected eyes, which were reduced by approximately 37% ( = 0.03) with CPP-P1 treatment. These findings suggest that CPP-P1 is a promising neuroprotective agent that promotes retinal ganglion cell survival and the preservation of other retinal neurons, potentially through enhanced CREB signaling in a rat model of SO-induced ocular hypertension.
本研究评估了与细胞穿透肽（CPP-P1）偶联的肽酶-1在青光眼高眼压模型中的神经保护潜力。通过前房内注射硅油（SO）使棕色挪威（BN）大鼠眼压升高，同时玻璃体内注射CPP-P1或赋形剂。分析视网膜横断面的神经保护标志物，包括环磷酸腺苷反应元件结合蛋白（CREB）、磷酸化CREB（p-CREB）、生长相关蛋白43（GAP43）、突触素-1（SYN1）和超氧化物歧化酶2（SOD2）。苏木精和伊红染色用于评估视网膜层厚度。经SO处理的大鼠内核层（INL，减少41%，P = 0.016）、内网层（IPL，减少52%，P = 0.0002）和神经节细胞层（GCL，减少57%，P = 0.001）的厚度显著降低。CPP-P1治疗减轻了这些降低，使INL厚度保留32%（P = 0.059），IPL保留19%（P = 0.119），GCL保留31%（P = 0.057）。与眼压升高且接受赋形剂治疗的视网膜相比，在眼压升高且接受CPP-P1治疗的视网膜中观察到CREB（P = 0.17）和p-CREB（P = 0.04）水平升高。尽管总体GAP43水平较低，但与经SO和赋形剂治疗的视网膜相比，在经SO和CPP-P1治疗的视网膜中，IPL和GCL内的表达有适度增加（分别为P = 0.15和P = 0.09）。SO注射降低了IPL和GCL中的SYN1表达（P = 0.01），而CPP-P1治疗显著增加了IPL（P = 0.03）和GCL（P = 0.002）中的SYN1水平。虽然所有组中GCL中的SOD2表达均极低，但在接受CPP-P1治疗的动物中观察到表达增加的趋势（P = 0.16）。在7天后去除SO并监测21天，随后制备视网膜平铺片以评估视网膜神经节细胞（RGC）存活情况，复制了SO模型。在注射SO的眼中观察到RGC损失42%（P = 0.009），而CPP-P1治疗使其减少约37%（P = 0.03）。这些发现表明，CPP-P1是一种有前景的神经保护剂，可能通过增强SO诱导的高眼压大鼠模型中的CREB信号传导，促进视网膜神经节细胞存活和其他视网膜神经元的保存。
