Buchanan Ocular Therapeutics Unit, Department of Ophthalmology, New Zealand National Eye Centre, University of Auckland, Auckland, New Zealand; Department of Ophthalmology, New Zealand National Eye Centre, University of Auckland, New Zealand.
Department of Ophthalmology, New Zealand National Eye Centre, University of Auckland, New Zealand.
Exp Eye Res. 2021 Jan;202:108384. doi: 10.1016/j.exer.2020.108384. Epub 2020 Dec 5.
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-1 beta (IL-1β) and tumour necrosis factor alpha (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 assembly, 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.
糖尿病性视网膜病变（DR）是糖尿病最常见的眼部并发症，是一种导致视力丧失的慢性血管和炎症性疾病。炎症小体途径是先天免疫系统的关键部分，需要激活 DR 中的慢性炎症。不幸的是，目前用于 DR 的治疗方法针对的是炎症小体途径下游的病理标志，因此在治疗该疾病方面仅部分有效。通过体外和体内 DR 模型发现，连接蛋白 43 半通道阻滞剂可以抑制炎症小体途径的激活。然而，这些研究是在体外细胞培养和体内动物疾病模型中进行的，这些模型具有预测性，但当然不能像任何模型一样完全复制人类的情况。在这里，我们开发了一种有用的模型附加物，即通过将人供体视网膜外植体暴露于高葡萄糖（HG）和促炎细胞因子白细胞介素 1β（IL-1β）和肿瘤坏死因子α（TNF-α）的组合，来建立 DR 的离体人器官型视网膜培养模型。我们假设，在这种模型中，连接蛋白 43 半通道阻断将防止 NLRP3 炎症小体复合物的组装，从而反过来减少 DR 特有的炎症迹象。为了验证我们的假设，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症小体途径中的分子变化。我们的结果表明，人视网膜外植体 DR 模型与炎症和炎症小体途径的激活有关，这是人类疾病的特征。此外，我们表明，通过用半通道调节剂托纳布沙阻断连接蛋白 43 半通道，我们能够防止 NLRP3 炎症小体复合物的组装、Müller 细胞的激活以及促炎细胞因子和 VEGF 的释放。这进一步支持了使用连接蛋白 43 半通道阻滞剂作为 DR 潜在新疗法的可能性。
