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Connexin43 hemichannel block inhibits NLRP3 inflammasome activation in a human retinal explant model of diabetic retinopathy
Experimental Eye Research ( IF 3.4 ) Pub Date : 2020-12-05 , DOI: 10.1016/j.exer.2020.108384
Henry H. Louie , Avik Shome , Charisse YJ. Kuo , Ilva D. Rupenthal , Colin R. Green , Odunayo O. Mugisho

Diabetic retinopathy (DR), the most common ocular complication associated with diabetes, is a chronic vascular and inflammatory disease that leads to vision loss. The inflammasome pathway, a key part of the innate immune system, is required to activate chronic inflammation in DR. Unfortunately, current therapies for DR target pathological signs that are downstream of the inflammasome pathway, making them only partly effective in treating the disease. Using in vitro and in vivo DR models, it was discovered that connexin43 hemichannel blockers can inhibit activation of the inflammasome pathway. However, those studies were conducted using in vitro cell culture and in vivo animal disease models that are predictive but do not, of course, like any model, completely replicate the human condition. Here, we have developed an addition to our armamentarium of useful models, an ex vivo human organotypic retinal culture model of DR by exposing human donor retinal explants to a combination of high glucose (HG) and pro-inflammatory cytokines, interleukin-1beta (IL-1β) and tumour necrosis factor (TNF-α). We hypothesized that in this model, connexin43 hemichannel block would protect against NLRP3 inflammasome complex assembly which would in turn decrease signs of inflammation characteristic of DR. To test our hypothesis, molecular changes in the inflammatory and inflammasome pathway were assessed using immunohistochemistry and a Luminex cytokine release assay. Our results showed that the human retinal explant DR model was associated with increased inflammation and activation of the inflammasome pathway, characteristic of the human condition. Furthermore, we showed that by blocking connexin43 hemichannels with the hemichannel modulator, tonabersat we were able to prevent NLRP3 inflammasome complex aggregation, Müller cell activation, as well as release of pro-inflammatory cytokines and VEGF. This further supports the possible use of connexin43 hemichannel blockers as potential new therapies for DR.



中文翻译:

连接蛋白43半通道阻滞在糖尿病性视网膜病变的人类视网膜外植模型中抑制NLRP3炎性体激活

糖尿病性视网膜病(DR)是与糖尿病相关的最常见的眼部并发症,是导致视力丧失的慢性血管和炎症性疾病。炎性体途径是先天免疫系统的关键部分,是激活DR中慢性炎症所必需的。不幸的是,目前针对DR的疗法靶向炎性体途径下游的病理征候,使其仅部分有效地治疗该疾病。使用体外体内DR模型,发现连接蛋白43半通道阻滞剂可以抑制炎性体途径的激活。但是,这些研究是使用体外细胞培养和体内进行的具有预测性的动物疾病模型,但是当然不能像任何模型一样完全复制人类的状况。在这里,我们开发了一种有用模型的补充品,一种离体通过将人类供体视网膜外植体暴露于高葡萄糖(HG)和促炎性细胞因子,白介素-1β(IL-1β)和肿瘤坏死因子(TNF-α)的组合下,建立人类DR的人类器官型视网膜培养模型。我们假设在该模型中,连接蛋白43半通道阻滞可防止NLRP3炎性小体复合物装配,从而减少DR的炎症特征。为了检验我们的假设,使用免疫组织化学和Luminex细胞因子释放测定法评估了炎症和炎性体途径中的分子变化。我们的结果表明,人类视网膜外植体DR模型与炎症的增加和炎性体途径的激活有关,这是人类疾病的特征。此外,我们证明,通过用半通道调节剂tonabersat阻断connexin43半通道,我们能够预防NLRP3炎性体复合物的聚集,Müller细胞活化以及促炎性细胞因子和VEGF的释放。这进一步支持将连接蛋白43半通道阻滞剂用作DR的潜在新疗法。

更新日期:2020-12-05
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