Huangdi Anxiao Capsules-containing serum protects cell model from cognitive dysfunction in diabetes via inhibiting NLRP3-mediated pyroptosis / 中国中药杂志

This study aims to investigate the effects and the molecular mechanism of Huangdi Anxiao Capsules(HDAX)-containing serum in protecting the rat adrenal pheochromocytoma(PC12) cells from diabetes-associated cognitive dysfunction induced by high glucose and whether the mechanism is related to the regulation of NOD-like receptor thermal protein domain associated protein 3(NLRP3)-mediated pyroptosis. The PC12 cell model of diabetes-associated cognitive dysfunction induced by high glucose was established and mcc950 was used to inhibit NLRP3. PC12 cells were randomized into control, model, HDAX-containing serum, mcc950, and HDAX-containing serum+mcc950 groups. Methyl thiazolyl tetrazolium(MTT) assay was employed to determine the viability, and Hoechst 33258/PI staining to detect pyroptosis of PC12 cells. Enzyme-linked immunosorbent assay(ELISA) was employed to measure the levels of interleukin-1 beta(IL-1β) and IL-18. Western blot was employed to determine the protein levels of postsynaptic density protein 95(PSD-95), NLRP3, apoptosis-associated speck-like protein containing a CARD(ASC), gasdermin D(GSDMD), GSDMD-N, and cleaved cysteinyl aspartate specific proteinase-1(caspase-1), and RT-PCR to determine the mRNA levels of NLRP3, ASC, GSDMD, and caspase-1. The immunofluorescence assay was adopted to measure the levels and distribution of NLRP3 and GSDMD-N in PC12 cells. Compared with the control group, the model group showed decreased cell proliferation, increased PI positive rate, down-regulated protein level of PSD-95, up-regulated protein levels of NLRP3, ASC, GSDMD-N, GSDMD, and cleaved caspase-1, up-regulated mRNA levels of NLRP3, ASC, GSDMD, and caspase-1, and elevated levels of IL-1β and IL-18. Compared with the model group, HDAX-containing serum, mcc950, and the combination of them improved cell survival rate and morphology, decreased the PI positive rate, down-regulated the protein levels of NLRP3, ASC, GSDMD-N, GSDMD, and cleaved caspase-1 and the mRNA levels of NLRP3, ASC, GSDMD, and caspase-1, and promoted the secretion of IL-1β and IL-18. The findings demonstrated that HDAX-containing serum can inhibit the pyroptosis-mediated by NLRP3 and protect PC12 cells from the cognitive dysfunction induced by high glucose.

Identification of chemical constituents in Huangdi Anxiao Capsules by UPLC-Q-TOF-MS~E combined with UNIFI software / 中国中药杂志

In order to clarify the main chemical constituents of Huangdi Anxiao Capsules, an ultra-high performance liquid coupled with quadrupole time-of-flight mass spectrometry(UPLC-Q-TOF-MS~E) combined with Waters UNIFI software were successfully used to rapidly identify the chemical constituents in Huangdi Anxiao Capsules. The mass spectrometry data of chemical constituents from Huangdi Anxiao Capsules were collected by UPLC-Q-TOF-MS~E, and their structures were identified by the results of UNIFI software according to relative retention time of reference standards, MS feature fragments and literature data of each compound. A total of 100 compounds in Huangdi Anxiao Capsules were identified, including 25 compounds from Pueraria Lobate Radix, 22 compounds from Coptis Rhizoma, 6 compounds from Ophiopogonis Radix, 14 compounds from Eriobotryae Folium, 22 compounds from Rehmanniae Radix, and 15 compounds from Notoginseng Radix et Rhizoma. Among them, 3 compounds were common components. These 100 compounds included flavonoids, alkaloids, saponins and organic acids. This study systematically analyzed the chemical composition of Huangdi Anxiao Capsules, so as to provide evidences for defining the chemical material basis of Huangdi Anxiao Capsules.

Effect of Huangdi Anxiao Capsules on zebrafish vascular lesions induced by high glucose and high fat / 中国中药杂志

Zebrafish of different strains with 5 dpf (5 days post-fertilization) were selected and fed with 0.2% high-fat diet for 8 h and 3% glucose solution for 16 halternatively during the day and night for 4 consecutive days. The zebrafish model was established and randomly divided into model group, Huangdi Anxiao Capsules (260 mg·L⁻¹) group and pioglitazone (32 mg·L⁻¹) group. The drug treatment groups were given the water-soluble drugs, with a volume of 25 mL, and incubated in a 28 °C incubator for 4 days. To detect the exposure to the corresponding drugs, the normal control group was set up. Thirty zebrafish were included in each group. The effect of Huangdi Anxiao Capsules on vascular wall thickness, fluorescence intensity of islet beta cells, fluorescence intensity of macrophages, and blood flow velocity of zebrafish were detected. The expressions of vascular endothelial growth factor (vegfaa) and angiotensin converting enzyme (ACE) were detected by RT-PCR. The results showed that compared with the model group, Huangdi Anxiao Capsules can significantly reduce the thickness of the blood vessel wall, increase the fluorescence intensity of islet β cells and macrophages, increase the blood flow velocity in vivo, and decrease the ACE and vegfaa expressions in zebrafish. It is suggested that Huangdi Anxiao Capsules may alleviate zebrafish vascular lesions by regulating the expressions of ACE and vegfaa.

Improving effects of Huangdi Anxiao Capsules on insulin resistance in type 2 diabetic GK rats / 中成药

AIM To observe the effects of Huangdi Anxiao Capsules (Puerariae lobatae Radix,Eriobotryae Folium,Notoginseng Radix et Rhizoma,etc.) on blood glucose and insulin resistance in GK rats with type 2 diabetes mellitus (T2DM) and its possible mechanism of action.METHODS GK rats with T2DM included in the experiment were randomly divided into model group,rosiglitazone (1.44 mg/kg) group,Shenqi Jiangtang Granules (1.08 g/kg) group,Huangdi Anxiao Capsules high,medium and low (12,6,3 g/kg) group and normal control group of Wistar rats.After six weeks of consecutive administration,fasting blood glucose (FBG),serum insulin (FINS),fasting plasma glucose (FPG),and insulin resistance index (HOMA-IR) were measured in all groups.Serum biochemical indexes of (TG,TC,HDL-C,LDL-C),glycosylated hemoglobin (HbA1c),glucagon-like peptide (GLP-1) and insulin-like growth factor (IGF-1) were measured.The pancreatic tissue was obtained by routine paraffin embedding and HE staining to observe the pathological changes.RESULTS Compared with the normal group,FBG,FPG,FINS,HOMA-IR,TG,TC,HDL-C,LDL-C and HbA1c of the model group were significantly higher (P ＜0.01),and GLP-1 and IGF-1 expressions were markedly decreased (P ＜ 0.01).Compared with the model group,the levels of FBG,FPG,FINS,HOMA-IR,TG,TC,HDL-C,LDL-C and HbA1 c in Huangdi Anxiao Capsules high-,medium-and low-dose group were significantly decreased (P ＜ 0.05,P ＜0.01);GLP-1 and IGF-1 expressions were markedly increased (P ＜0.05,P ＜0.01),compared with the model group,the structure changes of pancreatic tissue in Huangdi Anxiao Capsules high-,medium-and low-dose groups largely reduced.CONCLUSION Huangdi Anxiao Capsules can reduce GK rats fasting blood glucose,insulin resistance,and its mechanism may be related to promoting the emergence and proliferation of pancreatic islet cells,improving the function of islet beta cells,and increasing insulin secretion.