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Objective. In this study, we investigated the mechanism of Qing-Gan Li-Shui formulation (QGLSF) in treating primary open glaucoma (POAG) by network pharmacology and in vitro experiments. Methods. The active pharmaceutical ingredients (APIs) of GLQSF (prepared with Prunella vulgaris, Kudzu root, Plantago asiatica, and Lycium barbarum) were obtained from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) and Yet Another Traditional Chinese Medicine database (YATCM). The targets of POAG were screened out with GeneCards, OMIM, PharmGKB, Therapeutic Target Database (TTD), and DrugBank databases. The Venny platform was used to summarize the core targets. Topological analysis was performed using Cytoscape3.8.0. A protein-protein interaction network was plotted by STRING online. The key targets were subjected to GO and KEGG enrichment analyses. Finally, the effects of APIs were verified by a model of chloride hexahydrate (CoCl2)-induced retinal ganglion cells-5 (RGC-5). Results. The main APIs were selected as quercetin (Que) by network pharmacology. Nine clusters of QGLSF targets were obtained by the PPI network analysis, including AKT-1, TP53, and JUN. KEGG enrichment analysis showed that these targets were mainly involved in the AGE-RAGE signaling pathway. By in vitro experiments, Que promoted cell proliferation. The secretion of AKT-1, TP53, JUN, AGE, and RAGE in the cell culture supernatant decreased, as shown by ELISA. The mRNA levels of AKT-1, TP53, JUN, and RAGE decreased, as shown by RT-PCR. QGLSF may employ the AGE-RAGE signaling pathway to counter POAG. Conclusion. This study preliminarily elucidates the efficacy and mechanism of QGLSF in the treatment of POAG. [ABSTRACT FROM AUTHOR]

Ethnopharmacology relevance He-Ying-Qing-Re Formula (HF) was empirically modified from Si-Miao-Yong-An Decoction (SD), which was recorded in the literature of Divine Doctor's Secret Transmission, and has been utilized for centuries to treat vasculopathy through clearing heat and accelerating bloodstream. HF has been used as an effective holistic treatment of diabetic retinopathy (DR) for decades and experimentally reported to ameliorate retinal condition in diabetic mice. Aim of the study Our study aims to investigate the effect of HF in preventing sustained hyperglycemia and hyperlipidemia-associated retinal ganglion cell (RGC) cell death and its possible mechanism. Materials and methods Chromatographic fingerprint of HF was obtained upon the UPLC-based analytic system; Diabetic retinopathy was established in streptozotocin (STZ) injection-induced hyperglycemic mice; Alterations of retinal structure was assayed by H&E staining. Expression of PSD-95 and CHOP in retinae was assessed by immunofluorescence; RGC cell line (mRGC) was used for in vitro study. Cell death was analyzed by flow cytometry; Intracellular reactive oxygen species (ROS) was measured by 2′,7′-dichlorofluorescin diacetate (DCFDA); Apoptosis-related proteins and signaling were monitored with immunoblotting and colorimetric assay. Results Chlorogenic acid, ferulic acid, and rutin were identified in HF. HF attenuates the loss of RGCs, thinning of inner retinal layers in diabetic mice. Furthermore, expressions of Brn3a and PSD-95 were restored while CHOP level was downregulated upon HF treatment. In vitro study, HF alleviates H 2 O 2 -induced apoptosis of mRGCs and loss of postsynaptic protein via scavenging ROS and suppressing ATF4/CHOP-mediated endoplasmic reticulum stress and mitochondria-related pro-apoptotic factors, probably as cleaved-caspase-3, and phospho-p38 mitogen-activated protein kinase (MARK). Meanwhile, both pro-survival protein levels like Bcl-2/Bcl-xL and postsynaptic protein of PSD-95 were upregulated upon HF treatment. Conclusion HF administration was a valid therapeutic approach for DR treatment, oriented at the blockade of endoplasmic reticulum- and mitochondria-dependent oxidative stress-induced retinal neurodegeneration including RGC apoptosis. [ABSTRACT FROM AUTHOR]

Ethnopharmacology relevance He-Ying-Qing-Re Formula (HF) is a formula modified from “Si-Miao-Yong-An Decoction”, a traditional Chinese medical classic emerged in the Qing dynasty and has been reported for treatment of vascular diseases. HF, containing 8 herbs, has been used in local hospital for decades as a complementary method for diabetic retinopathy (DR) with retinal vascular dysfunction. Clinical reports revealed HF could ameliorate vision defects, microaneurysms, hemorrhages and macular edema. The aim of this study is to investigate the anti-DR action of HF and its underlying mechanism experimentally. Methods Chromatographic fingerprinting of HF and rodent model of DR were established; hypoglycemic effect of HF was measured by fasting, random blood glucose and glucose tolerance test; vascular degeneration was measured by retinal digestion; blood-retina-barrier (BRB) permeability was assessed with Evans Blue leakage assay. Advanced glycation end products (AGEs) were measured in vitro and in vivo level; Migration of retinal vascular endothelial cells were determined by wound healing and transwell chamber assays; permeability of endothelial monolayer was monitored with dextran transport. AGEs-related proteins and signaling were measured with immunoblotting and immunohistochemistry. Results Chlorogenic acid, ferulic acid and arctin were identified as major components in HF; HF suppresses retinal vasculature degeneration and BRB permeability damage without significant inhibition on hyperglycemia; HF reduces in vitro and in vivo formation of AGEs and AGEs-induced migration as well as permeability of retinal vascular endothelial cells. Expression of tight junction proteins Zo-1 and Claudin-1 was increased while activation of AGEs receptor and downstream signaling Akt were suppressed upon HF treatment. Conclusions HF exhibits protective effect against diabetic retinopathy, which may be associated with inhibition on AGEs and recovery on endothelial dysfunction via modulation of tight junction and AGEs downstream signaling. [ABSTRACT FROM AUTHOR]