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ETHNOPHARMACOLOGICAL RELEVANCE: Euonymus alatus (Thunb.) Siebold. (EA), a traditional Chinese medicine, is widely used in the treatment of diabetes. Our group has previously found that EA could treat diabetic retinopathy (DR) and stigmast-4-en-3-one (Numbered E6) is the active substance responsible for inhibiting angiogenesis in vitro by EA. However, the effects and mechanisms of E6 in the treatment of DR is still unknown. AIM OF THE STUDY: The aim of this study was to investigate the effects and mechanisms of E6 in EA on DR. Additionally, a comparison was made between the effects of E6 and triamcinolone acetonide (TA), as well as the side effects of E6 and dexamethasone. MATERIALS AND METHODS: Ocular affinity assessment and pharmacokinetic parameter prediction were conducted to evaluate the potential of E6 to treat DR. Retinal endothelial cells were used to investigate the in vitro inhibitory effect of E6 on vascular proliferation. Additionally, chicken embryos, zebrafish, and mice were used to investigate the in vivo anti-vascular proliferation effect of E6. Finally, diabetic mice were used to investigate whether E6 improves diabetic retinopathy and to compare its efficacy with that of TA. We then used network pharmacology to study the targets of E6 and performed molecular docking; followed by immunofluorescence experiments, ELISA, Western blot, and tube formation experiments to further investigate its mechanism. Finally, we compared the side effects of E6 with those of dexamethasone. RESULTS: E6 was found to have an affinity for the eye and to inhibit vascular proliferation both in vivo and in vitro. Moreover, E6 was found to be more efficacious than TA in the treatment of DR. Molecular docking experiments predicted that the glucocorticoid receptor (GR) is a potential target of E6, and immunofluorescence analyses confirmed that E6 upregulated the expression of the GR in the retina of hyperglycemic mice. In addition, western blotting results and tube formation experiments showed that E6 also attenuated angiogenesis by inhibiting the Hippo and VEGF pathways. Finally, by comparing the effects of E6 and dexamethasone on glucose regulation and osteoporosis, E6 was found to have fewer side effects. CONCLUSIONS: E6 is a highly effective drug for the treatment of DR, superior to TA and with fewer side effects than dexamethasone. Its mechanism involves the activation of glucocorticoid receptor and inhibition of Hippo and VEGF pathways to alleviate angiogenesis and inflammation. This study is the first to investigate the role and mechanism of E6 in improving DR. The findings suggest that E6 has unique advantages in the treatment of DR.
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Diabetes Mellitus Experimental , Retinopatía Diabética , Euonymus , Receptores de Glucocorticoides , Pez Cebra , Animales , Retinopatía Diabética/tratamiento farmacológico , Ratones , Receptores de Glucocorticoides/metabolismo , Embrión de Pollo , Diabetes Mellitus Experimental/tratamiento farmacológico , Euonymus/química , Masculino , Simulación del Acoplamiento Molecular , Dexametasona/farmacología , Ratones Endogámicos C57BL , Proliferación Celular/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Inhibidores de la Angiogénesis/farmacología , Triamcinolona Acetonida/farmacología , AngiogénesisRESUMEN
Diabetic cardiomyopathy (DCM) is one of the main complications of diabetic patients and the major reason for the high prevalence of heart failure in diabetic patients. Fufang Xueshuantong (FXST) is a traditional Chinese medicine formula commonly used in the treatment of diabetic retinopathy and stable angina pectoris. However, the role of FXST in DCM has not yet been clarified. This study was conducted to investigate the effects of FXST on diabetic myocardial lesions and reveal its molecular mechanism. The rats were intraperitoneally injected with 65 mg/kg streptozotocin (STZ) to induce diabetes mellitus (DM). DM rats were given saline or FXST. The rats in the control group were intraperitoneally injected with an equal amount of sodium citrate buffer and gavaged with saline. After 12 weeks, echocardiography, heart weight index (HWI), and myocardial pathological changes were determined. The expression of transforming growth factor-beta1 (TGF-ß1), collagen I, and collagen III was examined using immunofluorescence staining and western blot. The expressions of Wnt/ß-catenin signaling pathway-related proteins and mRNA were detected by western blot and real-time PCR. The results showed that FXST significantly improved cardiac function, ameliorated histopathological changes, and decreased HWI in the DM rats. FXST significantly inhibited the expression of myocardial TGF-ß1, collagen I, and collagen III in DM rats. Furthermore, FXST significantly inhibited the Wnt/ß-catenin pathway. Taken together, FXST has a protective effect on DCM, which might be mediated by suppressing the Wnt/ß-catenin pathway.
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Purpose: In this study, we investigated the mechanism of Tongluo Yishen (TLYS) decoction in more detail, from the perspective of pyroptosis in the unilateral ureteral ligation (UUO) model and the hypoxia-induced renal tubular epithelial (NRK-52E) cell. Method: The UUO model was used, and after 14 days of TLYS intervention, rats were tested for blood creatinine and urea nitrogen, HE staining was used to observe the pathological changes in the kidney, Masson staining was used to assess the degree of interstitial fibrosis, western blot was used to detect the changes of α-smooth muscle actin (α-SMA) protein expression level, immunohistochemistry and western blot detected the changes in protein expression levels of NOD-like receptor protein 3 inflammasome (NLRP3), gasdermin D (GSDMD), cysteinyl aspartate specific proteinase (caspase-1), interleukin 18 (IL-18) and interleukin 1ß (I L-1ß). A hypoxia model was created using NRK-52E cell, and after different concentrations of TLYS decoction intervention, the changes in the expression levels of pyroptosis were used with immunofluorescence and western blot methods. Results: TLYS decoction improved renal function, delayed the advancement of renal interstitial fibrosis, and inhibited pyroptosis in UUO rats. Furthermore, we observed that TLYS can mitigate hypoxia-induced NRK-52E cell damage via the suppression of the NLRP3-mediated pyroptosis. Conclusion: TLYS decoction exert renoprotective effects by inhibiting NLRP3-mediated pyroptosis.
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ETHNOPHARMACOLOGICAL RELEVANCE: Euonymus alatus (Thunb.) Siebold (family Celastraceae) is a deciduous woody shrub that is recorded in ShenNong BenCaoJing. It has been widely used for diabetes in traditional Chinese medicine. AIM OF THE STUDY: This study aimed to identify the most effective extract of Euonymus alatus (EA) against high glucose-induced endothelial cells in vitro, evaluate its pharmacological effect on retinopathy in diabetic mice and explore its underlying mechanism by RNA sequencing. METHODS: Retinal vascular endothelial cells (RF/6A) were treated with normal glucose (5.5 mmol/L glucose), high glucose (25 mmol/L glucose) or high glucose plus methanol extracts of EA (MEA), ethyl acetate extracts of EA (EEA) or water extracts of EA (WEA). The cytotoxicity and cell viability were determined by Cell Counting Kit-8 (CCK-8) assay. Cell migration was examined using the Transwell assay, and tube formation ability was measured using the Matrigel assay. Then, the KK-Ay mice were administered WEA or water for 12 weeks. The velocities of ocular blood flow were determined by Doppler ultrasound. RNA sequencing and reverse transcription quantitative PCR (RT-qPCR) were performed on WEA-stimulated RF/6A cells to reveal the underlying mechanism. RESULTS: The cytotoxicity assay found that 30 µg/mL MEA, 20 µg/mL EEA and 30 µg/mL WEA had no toxic effect on RF/6A cells. The cell viability results showed that MEA, EEA and WEA all decreased cell viability. Compared with the high-glucose group, both MEA and WEA decreased the number of migrated cells, while the inhibition rate of WEA was higher. The Matrigel results showed that 30 µg/mL WEA effectively reduced the total tube length. Moreover, WEA improved the haemodynamics of the central retinal artery. RNA sequencing coupled with RT-qPCR verified that WEA regulated angiogenesis-related factors in high glucose-stimulated RF/6A cells. CONCLUSIONS: WEA inhibits the migration and tube formation of RF/6A cells and improves diabetic retinopathy (DR) by mediating angiogenesis.
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Supervivencia Celular/efectos de los fármacos , Medicamentos Herbarios Chinos/uso terapéutico , Euonymus/química , Fitoterapia , Animales , Glucemia/efectos de los fármacos , Línea Celular , Movimiento Celular/efectos de los fármacos , Diabetes Mellitus , Medicamentos Herbarios Chinos/química , Glucosa/toxicidad , Haplorrinos , Masculino , Ratones , Ratones Endogámicos ARESUMEN
Tongluo Yishen (TLYS) decoction is an herb that is extensively applied for the treatment of chronic kidney disease (CKD) in traditional Chinese medicine. In this study, 37 different dominant chemical constituents of TLYS were identified. Rats with unilateral ureteral obstruction (UUO) were used as animal models, and TLYS decoction was administered orally for 14 days. TLYS decoction reduced the levels of renal function indicators, serum creatinine levels and blood urea nitrogen levels and alleviated renal pathological changes. Gene Ontology (GO) and KEGG pathway analyses of RNA sequencing data showed that TLYS decoction had significant effects on biological processes, cellular components and molecular functions in UUO rats and that the phagosome (a membrane source in the early stages of autophagy), lysosome (an important component of autolysosome), and oxidation pathways (which contribute to mitochondrial function) might be related to the antifibrotic effects of TLYS decoction. Moreover, we found significant mitochondrial function impairment, including a decreased mitochondrial membrane potential (MMP) and an imbalance in mitochondrial dynamics, excessive oxidative stress, and activation of Pink1/Parkin-mediated mitophagy in UUO rats. Treatment with TLYS decoction significantly increased the MMP, normalized mitochondrial dynamics and ameliorated renal injury. Moreover, TLYS alleviated the mitophagy clearance deficiency. In conclusion, our study showed that TLYS decoction can ameliorate mitochondrial dynamics by reducing oxidative stress and regulating mitophagy, thereby relieving renal injury, protecting renal function, and reducing renal fibrosis. This study provides support for the application of and further research on TLYS decoction.
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Diabetes mellitus (DM), a metabolic disorder, is the causes of oxidative stress leading to complications in micro- and macro-vascular system. The present study investigated sophocarpine for anti-diabetic potential in vivo in mice model. Sophocarpine administration to diabetic mice significantly (p < 0.05) attenuated glucose content in the plasma. The diabetes mediated lowering of GSH, ceruloplasmin and vitamin E was prevented in mice plasma by sophocarpine administration. Sophocarpine significantly (p < 0.05) reversed diabetes mediated suppression of insulin level and total Hb content in the mice plasma. In sophocarpine administrated diabetic mice C-peptide level was elevated and glycosylated hemoglobin content was suppressed significantly (p < 0.05) relative to diabetic group. Administration of sophocarpine significantly (p < 0.05) repressed diabetes mediated increase in TG and TC levels in dose-based manner. Administration of sophocarpine exhibited preventive role against diabetes mediated pathological damage to pancreas in the mice. Sophocarpine administration to diabetic mice repressed PPARγ recruitment significantly (p < 0.05) in dose-dependent manner. Sophocarpine prevents oxidative stress mediated pancreatic damage through increase in vitamin E, GSH and C-peptide levels, Moreover, the PPARγ activity was down-regulated, LDL-c content lowered and HDL-c level elevated in diabetic mice by sophocarpine. Therefore, sophocarpine may be developed for treatment of diabetes, however, further in vivo studies need to confirm the same.
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Alcaloides/farmacología , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Regulación de la Expresión Génica/efectos de los fármacos , Hipoglucemiantes/farmacología , Receptores Activados del Proliferador del Peroxisoma/metabolismo , Alcaloides/uso terapéutico , Animales , Glucemia/metabolismo , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patología , Progresión de la Enfermedad , Relación Dosis-Respuesta a Droga , Hipoglucemiantes/uso terapéutico , Masculino , Ratones , Estrés Oxidativo/efectos de los fármacosRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Fufang Xueshuantong (FXST) is a traditional Chinese patent medicine composed of Panax notoginseng (Burkill) F.H.Chen (Araliaceae), Salvia miltiorrhiza Bunge (Lamiaceae), Astragalus propinquus Schischkin (Leguminosae), and Scrophularia ningpoensis Hemsl. (Scrophulariaceae). It has been widely used for the treatment of diabetic retinopathy (DR) and exerts a positive clinical therapeutic effect. AIM OF THE STUDY: The aim of this study was to observe the effect of FXST on diabetic rat retinas and investigate its pharmacological mechanism for improving DR. METHODS: The diabetic rat model was established by intraperitoneal injection of streptozotocin. The rats were divided into a normal group, diabetic group, and FXST group. The rats in the FXST group were treated with FXST by intragastric administration for 12 weeks while other rats were given the same volume of normal saline. The haemodynamic parameters of the central retinal artery in the rats were measured by ultrasound. Haematoxylin-eosin staining was utilised to observe the pathological structural changes in the retina. The apoptosis of retinal nerve cells was detected by terminal deoxynucleotidyl transferase dUTP nick end labelling. RNA sequencing was used to screen the differentially expressed genes (DEGs), and enrichment analyses were performed. The DEGs were validated through real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). RESULTS: The peak systolic velocity, end diastolic velocity, and mean velocity decreased while the resistance index and pulsatility index increased in the diabetic rat retinas. FXST also improved haemodynamics. In contrast with the diabetic group, FXST allayed the disorder and oedema of the retinal structure in addition to reversing the reductions in retinal thickness and retinal ganglion cell number. It also decreased the apoptosis index of retinal cells. A total of 1134 DEGs were identified by RNA sequencing in the FXST group compared to the diabetic group, including 814 upregulated genes and 320 downregulated genes. These genes were enriched in the complement and coagulation cascades as well as the peroxisome proliferator-activated receptor (PPAR) signalling pathway. Several DEGs, including PPAR gamma, perilipin 4, acyl-CoA dehydrogenase long chain, CD55 molecule, and plasminogen activator urokinase, were identified by qRT-PCR, and the results were consistent with the RNA sequencing data. CONCLUSIONS: FXST alleviates DR by improving the haemodynamics and morphological alterations of diabetic rat retinas, which are mediated by complement and coagulation cascades and the PPAR signalling pathway.
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Diabetes Mellitus Experimental/tratamiento farmacológico , Retinopatía Diabética/tratamiento farmacológico , Medicamentos Herbarios Chinos/farmacología , Receptores Activados del Proliferador del Peroxisoma/efectos de los fármacos , Animales , Coagulación Sanguínea/efectos de los fármacos , Activación de Complemento/efectos de los fármacos , Diabetes Mellitus Experimental/complicaciones , Retinopatía Diabética/patología , Masculino , Receptores Activados del Proliferador del Peroxisoma/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , EstreptozocinaRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: HuoXue JieDu Formula (HXJDF) originates from classical formulas and was formed based on clinical experience. It is composed of Euonymus alatus (Thunb.) Siebold, Panax notoginseng (Burkill) F.H. Chen, the roots of Anguina kirilowii (Maxim.) Kuntze, and Coptis omeiensis (C. Chen) C.Y.Cheng. HXJDF prevents the deterioration of diabetic retinopathy. AIM OF THE STUDY: To evaluate the effects of HXJDF on diabetic retinopathy in rats and investigate the roles of miRNAs in the effects of HXJDF. MATERIALS AND METHODS: A single intraperitoneal injection of streptozotocin (STZ) (65 mg/kg) was used to induce diabetes in rats. Rats were divided into three groups: normal, diabetic, and diabetic + HXJDF. Rats were treated with HXJDF (15.4 g/kg) or water by oral gavage for twelve weeks. At the end of the treatment, rats were anaesthetized, and retinal haemodynamic changes were measured. Then, the retinas were removed and examined by haematoxylin and eosin (HE) staining and TUNEL assays. In addition, miRNA expression profiling was performed using miRNA microarrays and further validated by quantitative real-time PCR (qRT-PCR). RESULTS: Diabetes reduced peak systolic velocity (PSV), end-diastolic velocity (EDV), mean velocity (MV) and central retinal vein velocity (CRV) but increased the resistance index (RI) and pulsatility index (PI). In addition, in the diabetic group, retinal cell arrangement was disordered and loosely arranged, the retinal thickness and retinal ganglion cell (RGC) number decreased, and retinal cell apoptosis increased. In addition, 11 miRNAs were upregulated and 4 miRNAs were downregulated. After treatment, HXJDF improved retinal haemodynamics and morphologic changes, restored retinal thickness and RGC number and decreased retinal cell apoptosis. Furthermore, the changes in miRNA expression were significantly abolished by HXJDF. CONCLUSION: HXJDF may prevent DR by regulating the expression of miRNAs.
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Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , Retinopatía Diabética/tratamiento farmacológico , Retinopatía Diabética/metabolismo , Medicamentos Herbarios Chinos/uso terapéutico , MicroARNs/metabolismo , Animales , Diabetes Mellitus Experimental/genética , Retinopatía Diabética/genética , Composición de Medicamentos/métodos , Medicamentos Herbarios Chinos/síntesis química , Medicamentos Herbarios Chinos/farmacología , Masculino , MicroARNs/genética , Distribución Aleatoria , Ratas , Ratas Sprague-DawleyRESUMEN
The aim of the present study was to investigate the effect of a Traditional Chinese Herbal Medicine (TCHM), named Jinwei Tang on histone deacetylase 2 (HDAC2) and its role in the regulation of corticosteroid resistance in a rat model of chronic obstructive pulmonary disease (COPD). Male Wistar rats were divided into five groups (each n=10): COPD group, established by the intratracheal instillation of lipopolysaccharide and passive smoke exposure, and control, budesonide, theophylline + budesonide and Jinwei Tang + budesonide groups. Lung function was measured, lung tissue histopathology was examined and HDAC2 expression in the lung was assessed by immunohistochemistry. In addition, protein levels of interleukin-8 (IL-8), tumor necrosis factor (TNF)-α and HDAC2 in lung homogenate were quantified by ELISA. The rat COPD model exhibited alterations of the ratio of forced expiratory volume in 0.2 sec (FEV0.2) to the forced vital capacity, FEV0.2, dynamic compliance and airway resistance. HDAC2 expression was markedly reduced in the lung tissue of the COPD group compared with the control group, and treatment with Jinwei Tang + budesonide or theophylline + budesonide resulted in significant attenuation of the reduction of HDAC2 expression in the lungs (P<0.05). However, treatment with budesonide alone did not significantly alter HDAC2 expression. In the Jinwei Tang + budesonide and theophylline + budesonide groups, IL-8 and TNF-α expression was significantly decreased (P<0.05) and the HDAC2 level increased (P<0.05) compared with that in the COPD group. In conclusion, Jinwei Tang modulates airway inflammation and may enhance the anti-inflammatory effect of glucocorticoid through the upregulation of HADC2 expression in a rat model of COPD.
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BACKGROUND: The Hippo signaling pathway is reported to be involved in angiogenesis, but the roles of the Hippo pathway in diabetic retinopathy have not been addressed. Fufang Xueshuantong Capsule has been used to treat diabetic retinopathy in China; however, the effect of Fufang Xueshuantong Capsule on the Hippo pathway has not been investigated. METHODS: In this study, diabetes was induced in Sprague-Dawley rats with intraperitoneal injection of streptozotocin. Twenty weeks later, Fufang Xueshuantong Capsule was administered for 12 weeks. When the administration ended, the eyes were isolated for western blot and immunohistochemistry analyses. The levels of P- mammalian sterile 20-like (MST), large tumor suppressor homolog (Lats), P- yes-associated protein (YAP), transcriptional co-activator with PDZ binding motif (TAZ) and TEA domain family members (TEAD) were measured. RESULTS: Diabetic rats had a decreased P-MST level in the inner plexiform layer and reduced expression of P-YAP in the photoreceptor layers of their eyes. In addition, diabetic rats displayed remarkable increases in Lats, TAZ and TEAD in their retinas. Furthermore, Fufang Xueshuantong Capsule restored the changes in the Hippo pathway. CONCLUSIONS: The Hippo signaling pathway is important for the progression of diabetic retinopathy and will hopefully be a targeted therapeutic approach for the prevention of diabetic retinopathy.
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Retinopatía Diabética/tratamiento farmacológico , Medicamentos Herbarios Chinos/administración & dosificación , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal/efectos de los fármacos , Animales , China , Retinopatía Diabética/genética , Retinopatía Diabética/metabolismo , Humanos , Masculino , Patentes como Asunto , Proteínas Serina-Treonina Quinasas/genética , Ratas , Ratas Sprague-DawleyRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Euonymus alatus, Radix trichosanthis, Panax notoginseng and Coptis chinensis are popular plants used in traditional Chinese medicine to treat diabetes. AIM OF THE STUDY: The aim of the study is to investigate the therapeutic effect of the active components of Euonymus alatus, Radix trichosanthis, Panax notoginseng and Coptis chinensis (cERPC) on diabetic peripheral neuropathy in the rats and explore the underlying mechanism involved. METHODS: After diabetes was induced in rats for 20 weeks, cERPC or water was administered for 12 weeks. After a hot plate test, motor nerve conduction velocity and sciatic nerve blood flow were determined; the sciatic nerves were isolated for toluidine blue staining; and the fibre area, fibre diameter, axon area, axon diameter and myelin thickness were evaluated. The levels of the myelin basic protein, myelin protein zero, Oct6 and Krox20 were measured by western blot or immunofluorescence. RESULTS: cERPC was efficient in reducing the response latency, increasing motor nerve conduction velocity, enhancing sciatic nerve blood flow and ameliorating the pathological changes in diabetic rats. cERPC also had a role in increasing the levels of myelin basic protein and myelin protein zero and improving the expression of Oct6 and Krox20 in sciatic nerves of diabetic rats. CONCLUSIONS: cERPC ameliorates diabetic peripheral neuropathy by attenuating electrophysiological, circulatory and morphological alterations, which is mediated by the Oct6-Krox20 pathway.
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Neuropatías Diabéticas/prevención & control , Medicamentos Herbarios Chinos/uso terapéutico , Sustancias Protectoras/uso terapéutico , Animales , Axones/efectos de los fármacos , Axones/patología , Axones/ultraestructura , Diabetes Mellitus Experimental/tratamiento farmacológico , Neuropatías Diabéticas/patología , Masculino , Neuronas Motoras/efectos de los fármacos , Proteínas de la Mielina/metabolismo , Vaina de Mielina/efectos de los fármacos , Vaina de Mielina/patología , Vaina de Mielina/ultraestructura , Conducción Nerviosa/efectos de los fármacos , Dimensión del Dolor/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Flujo Sanguíneo Regional/efectos de los fármacos , Nervio Ciático/irrigación sanguíneaRESUMEN
CGRP is reported to be implicated in the process of diabetes and neuronal disease. However, the role and underlying mechanism of CGRP involved in diabetic neuropathy is unknown. Schwann cells play a central role in diabetic neuropathy, therefore the protective effect of CGRP on Schwann cells exposed to high glucose is determined. In the present study, full-length CGRP cDNA was isolated and then transferred to gateway adapted lentivirus expression vector by LR recombination reaction. Afterwards, the CGRP bearing recombinant virus was prepared in 293 FT cells and used to infect Schwann cells. The viability and superoxide anions of Schwann cells were evaluated following stimulation with high glucose, and levels of SOD, MDA and NOX1 were assessed. The results suggested that CGRP expression was up-regulated following lentivirus transfection. Lenti-CGRP increased cell viability in high glucose, but the effect was transient. Further lenti-CGRP protected against oxidative stress in Schwann cells triggered by high glucose and lenti-CGRP was effective in increasing SOD and decreasing MDA level. Meanwhile, the increased level of NOX1 caused by high glucose was reversed by lenti-CGRP overexpression. We therefore, suggest that lenti-CGRP may play a role in inhibiting oxidative stress in Schwann cell lines following hyperglycemic stimulation.