Protection of Taohong Siwu Decoction on PC12 cells injured by oxygen glucose deprivation/reperfusion via mitophagy-NLRP3 inflammasome pathway in vitro.

ETHNOPHARMACOLOGICAL RELEVANCE: Taohong Siwu Decoction (THSWD) is a traditional Chinese medicine formula used to invigorate blood circulation and resolve blood stasis. It consists of Paeonia lactiflora Pall., Conioselinum anthriscoides (H.Boissieu) Pimenov & Kljuykov, Rehmannia glutinosa (Gaertn.) DC., Prunus persica (L.) Batsch, Angelica sinensis (Oliv.) Diels, and Carthamus creticus L. in the ratio of 3:2:4:3:3:2. THSWD is a common prescription for the treatment of ischemic stroke. AIM OF THE STUDY: To study the protective effect and mechanism of Taohong Siwu Decoction (THSWD) on PC12 cells damaged by oxygen glucose deprivation/reperfusion (OGD/R). MATERIALS AND METHODS: OGD/R model of PC12 cells was used to simulate ischemia-reperfusion (I/R) injury of nerve cells in vitro. The experiment was grouped as follows: control, OGD/R and OGD/R + THSWD (5%, 10% and 15%) group. Oxygen and glucose was restored for 24 h after 4-6 h of deprivation. The severity of damage to PC12 cells was evaluated by CCK8, flow cytometry and lactate dehydrogenase (LDH). Mitochondrial morphology and function were examined by transmission electron microscopy (TEM), ATP and mitochondrial membrane potential (MMP) assay kits. Cellular autophagy and NLRP3 inflammasome-associated proteins were detected by Western blot and immunofluorescence staining. RESULTS: THSWD treatment improved the survival rate of PC12 cells injured by OGD/R, reduced cell damage and apoptosis. Moreover, ATP, MMP and the expression of autophagy marker proteins (LC3-II/LC3-I, Beclin1, Atg5) and mitophagy marker proteins (Parkin and PINK-1) was significantly elevated. The reactive oxygen species (ROS), NLRP3 inflammasome and pro-inflammatory cytokines induced by OGD/R were distinctly reduced. In contrast, these above beneficial effects of THSWD on mitochondrial autophagy and NLRP3 inflammasome were reversed by mitochondrial division inhibitory factor 1 (Mdivi-1). CONCLUSION: THSWD protects PC12 cells against OGD/R injury by heightening mitophagy and suppressing the activation of NLRP3 inflammasome.

Neuroprotective Effect of Taohong Siwu Decoction on Cerebral Ischemia/Reperfusion Injury via Mitophagy-NLRP3 Inflammasome Pathway.

Objective: Globally, cerebral ischemia has been shown to be the second leading cause of death. Our previous studies have shown that Taohong Siwu Decoction (THSWD) exhibits obvious neuroprotective effects on cerebral ischemia/reperfusion (I/R) injury (CIRI). In this study, we further explored the modulatory effect of THSWD on mitochondrial autophagy in CIRI and the relationship between modulatory effect and NLRP3 inflammatory vesicle activation, so as to further explain the mechanism of neuroprotective effect of THSWD. Methods: Middle cerebral artery occlusion reperfusion (MCAO/R) model in rats was built to simulate I/R. Adult male SD rats (220-270 g) were randomly divided into the following four groups: the sham group, the MCAO/R group, the MCAO/R + THSWD group, and the MCAO/R + THSWD + Mitochondrial division inhibitor 1 (Mdivi-1) group. Neurological defect scores were used to evaluate neurological function. 2,3,5-Triphenyltetrazolium chloride (TTC) staining was conducted to measure cerebral infarct volume. Nissl staining, H&E staining and TUNEL staining were executed to detect ischemic cortical neuronal cell viability and apoptosis. Electron microscopy was used to observe the ultrastructural changes of mitochondria. Total Reactive Oxygen Species (ROS) in tissue were measured by fluorescence spectrophotometry, and the activation status of microglia was evaluated by Iba-1/CD16 immunofluorescence staining. The levels of mitophagy-related proteins (LC3, Parkin, PINK1), NLRP3 inflammasome-related proteins (NLRP3, ASC, Pro-caspase-1, Cleaved-caspase-1), and inflammatory cytokines (Pro-IL-18, Pro-IL-1ß, IL-18, IL-1ß) were evaluated by western blotting. Results: The studies showed that THSWD treatment alleviated cerebral infarction and neurological deficiencies. THSWD upregulated the expressions of autophagy markers (LC3-II/LC3-I and Beclin1) mitochondrial autophagy markers (Parkin and PINK1) after CIRI. Furthermore, THSWD treatment attenuated microglia activation and damage to mitochondrial structures, thereby reducing ROS production and NLRP3 inflammasome activation. In contrast, the mitochondrial autophagy inhibitor Mdivi-1 inhibited the above beneficial effects of THSWD. Conclusions: THSWD exhibits neuroprotective effects against MCAO/R in rats by enhancing mitochondrial autophagy and reducing NLRP3 inflammasome activation.

Novel Polysaccharide H-1-2 from Pseudostellaria Heterophylla Alleviates Type 2 Diabetes Mellitus.

BACKGROUND/AIMS: To investigate the potential therapeutic effect of novel polysaccharide H-1-2 from pseudostellaria heterophylla against type 2 Diabetes Mellitus (T2DM) and elucidate the underling molecular mechanisms. METHODS: Relative expression of HIF1α and Sirt1 in T2DM patients was determined via real-time PCR. The direct binding of HIF1α on Sirt1 promoter was validated by ChIP assay. The inhibitory regulation of Sirt1 by HIF1α was analyzed using luciferase reporter assay. The endogenous protein of HIF1α and Sirt1 in response to H-1-2 treatment was quantified by western blotting. The blood glucose, secreted insulin and serous lipid profiles were measured with ELISA kits. RESULTS: We consolidated that HIF1α and Sirt1 was dysregulated in T2DM patients and subjected to H-1-2 modulation. H-1-2 significantly inhibited hypoxia and up-regulated Sirt1 expression in EndoC-ßH1 cells. Accordingly, H-1-2 enhanced glucose-stimulation insulin secretion and improved blood glucose and lipid profiles in T2DM cells, and elevated the glucose and insulin tolerance simultaneously. Furthermore, we demonstrated that H-1-2 alleviated T2DM via inhibition of hypoxia and up-regulation of Sirt1 in isolated pancreatic ß-cells from T2DM rats. CONCLUSION: Our data unambiguously demonstrated H-1-2 administration alleviated T2DM by enhancing Sirt1 expression through inhibition of hypoxia.

[Protective effects and mechanism of congguiyishen capsules on diabetic nephropathy rats].

OBJECTIVE: To study the curative and protective effects of Congguiyishen Capsules on the diabetic nephropathy (DN) model rats. METHODS: Established the DN model rats by intraperitoneal injection of urea bacteria element (Streptozotocin, STZ). The rats were divided into six groups including normal control group, model group, positive control group, high-dosage group, medium-dosage group and low-dosage group. After oral administration for 4 weeks, determined the 24 h urinary protein, Cr, kidney mass/body mass, FBG, Ang II, AT1R, AGTRAP and CTGF in the kidney. Observed the pathological damage of kidney tissue with Masson staining. RESULTS: After treatment, Cr, kidney mass index, 24 h urine protein, FBG and Ang II were decreased signicantly (P < 0.05). And the treatment also alleviated the pathological damage of kidney tissue. CONCLUSION: Congguiyishen Capsules have protective effect for DN model rats. The mechanism may be related to the suppression of inflammatory response and down-regulating the expression of AT1R, AGTRAP and CTGF.

[Protective effects of Qizhen Jiangtang granules in diabetic nephropathy rats].

OBJECTIVE: To study the curative and protective effects of Qizhen Jiangtang Granules in the diabetic nephropathy (DN) model rats. METHODS: Healthy SD rats were fed a high-sucrose and high-fat diet and intraperitoneal injection of streptozotocin (STZ, 30 mg/kg) to establish the DN model. The rats were divided into six groups including normal control group,model group, positive control group, high-dosage group(200 mg/kg), medium-dosage group (100 mg/kg), and low-dosage group(50 mg/kg). After oral administration of Qizhen Jiangtang Granules for eight weeks, FBG,TG,TC, LDL-c, HDL-c, SCr and BUN levels in rats serum were determined, while the pathological damage of kidney tissue with PAS and HE staining were observed under microscope. RESULTS: After treatment, TG, TC, LDL-c,SCr and BUN levels were significantly decreased(P <0. 05), and HDL-c level was significantly increased(P <0. 05). The treatment also alleviated the pathological damage of kidney tissue. CONCLUSION: Qizhen Jiangtang Granules have a protective effect against kidney damage in DN model rats. The mechanism may be related to the regulation of lipid and sugar levels in serum.

[Study on the dynamic model with supercritical CO2 fluid extracting the lipophilic components in Panax notoginseng].

OBJECTIVE: To establish a dynamics model for extracting the lipophilic components in Panax notoginseng with supercritical carbon dioxide (CO2). METHODS: Based on the theory of counter-flow mass transfer and the molecular mass transfer between the material and the supercritical CO2 fluid under differential mass-conservation equation, a dynamics model was established and computed to compare forecasting result with the experiment process. RESULTS: A dynamics model has been established for supercritical CO2 to extract the lipophilic components in Panax notoginseng, the computed result of this model was consistent with the experiment process basically. CONCLUSION: The supercritical fluid extract dynamics model established in this research can expound the mechanism in the extract process of which lipophilic components of Panax notoginseng dissolve the mass transfer and is tallied with the actual extract process. This provides certain instruction for the supercritical CO2 fluid extract' s industrialization enlargement.