Thanuja M Y, Ranganath Sudhir H, Srinivas Sangly P
Bio-INvENT Lab, Department of Chemical Engineering, Siddaganga Institute of Technology, Tumakuru, India.
School of Optometry, Indiana University, Bloomington, Indiana, USA.
J Ocul Pharmacol Ther. 2022 Dec;38(10):664-681. doi: 10.1089/jop.2022.0082. Epub 2022 Oct 17.
To characterize the impact of corneal cold storage (CS) on the endothelial apical junctional complex (AJC). Porcine corneas were held in CS (4°C; 1-7 days) with Cornisol™ preservation medium supplemented with epothilone B (EpoB; microtubule stabilizer; 100 nM), SB-203580 (p38 mitogen-activated protein [MAP] kinase inhibitor; 20 μM), or antioxidants (quercetin, 100 μM; vitamin E, 1 mM; deferoxamine, an iron chelator, 10 mM). After CS termination, the damage to endothelial AJC was characterized by imaging perijunctional actomyosin ring (PAMR) and zonula occludens (ZO-1). The effects of EpoB and SB-203580 were characterized by imaging microtubules. The loss in the barrier function was assessed in cultured cells grown on biotin-coated gelatin by permeability to fluorescein isothiocyanate (FITC)-avidin. The accumulation of reactive oxygen species (ROS), altered mitochondrial membrane potential (MMP), lipid peroxidation, and lactate dehydrogenase (LDH) release were also determined in response to CS. CS led to the loss of microtubules, destruction of PAMR, and breakdown of ZO-1 in the endothelium. The severity of damage increased when CS was prolonged. Although rewarming of the tissue increased the damage, the effect was marginal. CS also induced accumulation of ROS, alteration in MMP, lipid peroxidation, enhanced LDH release, and increased permeability to FITC-avidin. These changes were opposed by EpoB, SB-203580, and antioxidants. Corneal CS destroys AJC of the endothelium, leading to loss of its barrier function. The effects were surmounted by microtubule stabilization, p38 MAP kinase inhibition, and antioxidants. Thus, there is potential for reformulation of the preservation medium to maintain the health of the donor corneal endothelium before transplantation.
为了描述角膜冷藏（CS）对内皮顶端连接复合体（AJC）的影响。将猪角膜置于添加了埃坡霉素B（EpoB；微管稳定剂；100 nM）、SB - 203580（p38丝裂原活化蛋白[MAP]激酶抑制剂；20 μM）或抗氧化剂（槲皮素，100 μM；维生素E，1 mM；去铁胺，一种铁螯合剂，10 mM）的Cornisol™保存培养基中进行冷藏（4°C；1 - 7天）。CS结束后，通过成像连接周肌动球蛋白环（PAMR）和紧密连接（ZO - 1）来描述对内皮AJC的损伤。通过成像微管来描述EpoB和SB - 203580的作用。通过异硫氰酸荧光素（FITC） - 抗生物素蛋白的通透性，评估在生物素包被的明胶上生长的培养细胞中的屏障功能丧失情况。还测定了对CS反应时活性氧（ROS）的积累、线粒体膜电位（MMP）的改变、脂质过氧化和乳酸脱氢酶（LDH）释放。CS导致内皮中微管丢失、PAMR破坏和ZO - 1解体。当CS延长时，损伤的严重程度增加。尽管组织复温会增加损伤，但影响较小。CS还诱导ROS积累、MMP改变、脂质过氧化、LDH释放增强以及对FITC - 抗生物素蛋白的通透性增加。这些变化被EpoB、SB - 203580和抗氧化剂所对抗。角膜CS破坏内皮的AJC，导致其屏障功能丧失。微管稳定、p38 MAP激酶抑制和抗氧化剂可克服这些影响。因此，在移植前重新配制保存培养基以维持供体角膜内皮健康具有潜力。
