Natural products as pharmacological modulators of mitochondrial dysfunctions for the treatment of diabetes and its complications: An update since 2010.

Diabetes, characterized as a well-known chronic metabolic syndrome, with its associated complications pose a substantial and escalating health and healthcare challenge on a global scale. Current strategies addressing diabetes are mainly symptomatic and there are fewer available curative pharmaceuticals for diabetic complications. Thus, there is an urgent need to identify novel pharmacological targets and agents. The impaired mitochondria have been associated with the etiology of diabetes and its complications, and the intervention of mitochondrial dysfunction represents an attractive breakthrough point for the treatments of diabetes and its complications. Natural products (NPs), with multicenter characteristics, multi-pharmacological activities and lower toxicity, have been caught attentions as the modulators of mitochondrial functions in the therapeutical filed of diabetes and its complications. This review mainly summarizes the recent progresses on the potential of 39 NPs and 2 plant-extracted mixtures to improve mitochondrial dysfunction against diabetes and its complications. It is expected that this work may be useful to accelerate the development of innovative drugs originated from NPs and improve upcoming therapeutics in diabetes and its complications.

Tanshinone IIA Combined With Cyclosporine A Alleviates Lung Apoptosis Induced by Renal Ischemia-Reperfusion in Obese Rats.

Acute lung injury (ALI), which is induced by renal ischemia-reperfusion (IR), is one of the leading causes of acute renal IR-related death. Obesity raises the frequency and severity of acute kidney injury (AKI) and ALI. Tanshinone IIA (TIIA) combined with cyclosporine A (CsA) was employed to lessen the lung apoptosis led by renal IR and to evaluate whether TIIA combined with CsA could alleviate lung apoptosis by regulating mitochondrial function through the PI3K/Akt/Bad pathway in obese rats. Hematoxylin-eosin (HE) staining was used to assess the histology of the lung injury. Terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) was used to assess apoptosis of the lung. Electron microscopy was used to assess mitochondrial morphology in lung cells. Arterial blood gas and pulmonary function were used to assess the external respiratory function. Mitochondrial function was used to assess the internal respiratory function and mitochondrial dynamics and biogenesis. Western blot (WB) was used to examine the PI3K/Akt/Bad pathway-related proteins. TIIA combined with CsA can alleviate lung apoptosis by regulating mitochondrial function through the PI3K/Akt/Bad pathway in obese rats.

Tanshinone IIA combined with CsA inhibit myocardial cell apoptosis induced by renal ischemia-reperfusion injury in obese rats.

BACKGROUND: Acute myocardial injury (AMI), which is induced by renal ischemia-reperfusion (IR), is a significant cause of acute kidney injury (AKI)-related associated death. Obesity increases the severity and frequency of AMI and AKI. Tanshinone IIA (TIIA) combined with cyclosporine A (CsA) pretreatment was used to alleviate myocardial cell apoptosis induced by renal IR, and to determine whether TIIA combined with CsA would attenuate myocardial cell apoptosis by modulating mitochondrial function through the PI3K/Akt/Bad pathway in obese rats. METHODS: Male rates were fed a high fat diet for 8 weeks to generate obesity. AKI was induced by 30 min of kidney ischemia followed 24 h of reperfusion. Obese rats were given TIIA (10 mg/kg·d) for 2 weeks and CsA (5 mg/kg) 30 min before renal IR. After 24 h of reperfusion, the rats were anaesthetized, the blood were fetched from the abdominal aorta and kidney were fetched from abdominal cavity, then related indicators were examined. RESULTS: TIIA combined with CsA can alleviate the pathohistological injury and apoptosis induced by renal IR in myocardial cells. TIIA combined with CsA improved cardiac function after renal ischemia (30 min)-reperfusion (24 h) in obese rats. At the same time, TIIA combined with CsA improved mitochondrial function. Abnormal function of mitochondria was supported by decreases in respiration controlling rate (RCR), intracellular adenosine triphosphate (ATP), oxygen consumption rate, and mitochondrial membrane potential (MMP), and increases in mitochondrial reactive oxygen species (ROS), opening of the mitochondrial permeability transition pore (mPTP), mitochondrial DNA damage, and mitochondrial respiratory chain complex enzymes. The injury of mitochondrial dynamic function was assessed by decrease in dynamin-related protein 1 (Drp1), and increases in mitofusin1/2 (Mfn1/2), and mitochondrial biogenesis injury was assessed by decreases in PPARγ coactivator-1-α (PGC-1), nucleo respiratory factor1 (Nrf1), and transcription factor A of mitochondrial (TFam). CONCLUSION: We used isolated mitochondria from rat myocardial tissues to demonstrate that myocardial mitochondrial dysfunction occurred along with renal IR to induce myocardial cell apoptosis; obesity aggravated apoptosis. TIIA combined with CsA attenuated myocardial cell apoptosis by modulating mitochondrial function through the PI3K/Akt/Bad pathway in obese rats.

Vascular Endothelial Function as a Valid Predictor of Variations in Pulmonary Function in T2DM Patients Without Related Complications.

To assess the variations in pulmonary function and vascular endothelial function in their early stages (without related complications). A total of 162 type 2 diabetes mellitus (T2DM) patients without diabetes complications and 55 healthy people were selected, comprising the T2DM group and the control group, respectively, to evaluate changes in vascular endothelial function and lung function and determine the correlation between them. In this study, the T2DM group exhibited significantly lower pulmonary function than that of the control group (P < 0.05). The T2DM group also showed significantly lower flow-mediated dilation (FMD) and nitric oxide (NO) (P < 0.05) than those of the control group. Pulmonary functional indexes correlated positively with FMD and NO (P < 0.05) and correlated negatively with endothelin-1 (ET-1) (P < 0.05). FMD and NO correlated negatively with diabetes duration/HbA1c (P < 0.05), whereas ET-1 correlated positively with glycosylated hemoglobinA1c (HbA1c)/diabetes duration (P < 0.05). Pulmonary functional indexes negatively correlated with HbA1c/diabetes duration (P < 0.05). Multiple linear regression was used to analyze the relationship between vascular endothelial function indexes (FMD, ET-1, and NO) and pulmonary functional indexes. The results indicated that each vascular endothelial function index (FMD, ET-1, and NO) was significantly correlated with the pulmonary functional index (P < 0.05). The patients with T2DM presented changes in the subclinical vascular endothelial and pulmonary function. They also had impaired vascular endothelial functions, which were characterized by reduced vascular endothelial function relative to those of healthy people. Regulating glycemia may improve vascular endothelial and pulmonary functions. Moreover, microvascular lesions in preclinical stages, vascular endothelial function indexes (FMD, ET-1, and NO) were valid predictors of alterations in pulmonary function in T2DM patients without related complications. Clinical Trial Registration: ClinicalTrials.gov, identifier NCT03575988.

Enhanced antitumor efficacy using epirubicin and schisandrin B co-delivery liposomes modified with PFV via inhibiting tumor metastasis.

As a malignant tumor, breast cancer is very prone to metastasis. Chemotherapy is one of the most common means for treating breast cancer. However, due to the serious metastasis and the poor targeting effect of traditional chemotherapeutic drugs, even after years of efforts, the therapeutic effect is still unsatisfied. Therefore, in this study, we constructed a kind of PFV modified epirubicin plus schisandrin B liposomes to solve the above disadvantages. In vitro experiments showed that the targeting liposomes with ideal physicochemical property could increase the cytotoxicity of MDA-MB-435S cells, destroy the formation of vasculogenic mimicry (VM), and inhibit tumor invasion and migration. Action mechanisms indicated that the inhibition of targeting liposomes on tumor metastasis was attributed to the regulation of the expression of vascular endothelial growth factor (VEGF), matrix metalloproteinase-9 (MMP-9), vimentin (VIM), and E-cadherin (E-cad). In vivo pharmacodynamic experiments showed that the targeting liposomes could significantly improve the antitumor effect in mice. H&E staining and TUNEL results showed that the targeting liposomes could promote the apoptosis of tumor cells. Hence, the PFV modified epirubicin plus schisandrin B liposomes constructed in this study provided a new therapeutic strategy for breast cancer.

Early changes in pulmonary function and intrarenal haemodynamics and the correlation between these sets of parameters in patients with T2DM.

PURPOSE: The main objectives of this study were to assess the early changes in pulmonary function and intrarenal haemodynamics and to determine the correlation between pulmonary function and intrarenal haemodynamics in patients with type 2 diabetes mellitus (T2DM). METHODS: 96 patients with T2DM (diabetes group) without diabetes kidney disease (DKD) and 33 healthy subjects (control group) were enrolled in studies intended to assess the early changes in pulmonary function and intrarenal haemodynamics associated with diabetes, as well as to determine the correlation between pulmonary function and intrarenal haemodynamics. RESULTS: Pulmonary functional parameters were negatively correlated with HbA1c levels and diabetes duration (P< 0.05). Moreover, renal functional parameters were positively correlated with HbA1c levels and diabetes duration (P<0.05). Additionally, pulmonary functional parameters were negatively correlated with renal functional parameters (P<0.05). Multiple linear regression analysis of the relationship between pulmonary functional parameters and the bilateral kidney arterial resistivity index (RI) showed that all the pulmonary functional parameters were significantly correlated with the arterial RI (P< 0.05). CONCLUSIONS: Patients displayed changes in pulmonary function and intrarenal haemodynamics during the preclinical stages of DKD. Regulating glycaemia may improve intrarenal haemodynamics in the bilateral interlobular renal arteries. Moreover, during the preclinical stages of DKD, the right kidney RI is a effective predictor of early changes in pulmonary function in adult T2DM patients. TRIAL REGISTRATION: ClinicalTrials.gov (NCT02798198); registered 8 June 2016.

Longitudinal follow-up study of the retrobulbar and intrarenal hemodynamics in patients with T2DM.

OBJECTIVE: The primary aim of this study is to examine the hemodynamics of retrobulbar and intrarenal in the changes of early stage of type 2 diabetes mellitus (T2DM) patients from 2000 to 2015 and to assess incidence associated with diabetic kidney disease (DKD) and diabetic retinopathy (DR). METHOD: Our study contained 60 subjects newly diagnosed of T2DM were divided into 2 groups base on the mean resistive index (RI) (≤0.7 and >0.7) of hemodynamic and to compare between-group differences of the early changes in hemodynamics of retrobulbar and intrarenal and also to conclude the incidences of diabetic kidney disease (DKD) and diabetic retinopathy (DR)subsequently with a long follow-up duration(2000-2015). First, to compare the mean RI of central retinal artery (CRA) between 2 groups. Second, to compare the mean RI of intrarenal hemodynamics in the bilateral interlobular renal arteries, renal function parameters (blood urea nitrogen (BUN), creatinine (Cr), blood glucose parameters (glycosylated hemoglobinA1c (HbA1c), fasting plasma glucose (FBG), and 2-hour postprandial blood glucose (2hPBG)), glomerular filtration rate (GFR), albumin excretion rate (AER), and urine albumin-to-creatinine ratio (UACR) between 2 groups. RESULTS: First part of our follow-up studies was to compare hemodynamic RI index of retrobulbar in years of 2000 and 2015, both renal function and blood glucose parameters were fund significantly enhanced in subject group RIs ≤0.7. Incidence of DKD and DR was notably lower in group RIs ≤0.7 than group RIs > 0.7, difference was statistically significant (P < .05). Incidence of HbA1c ≤7% was higher in group RIs ≤0.7 than group RIs >0.7, but difference was not statistically significant (P > .05). Incidence of proliferative diabetic retinopathy (PDR) was notably lower in group RIs ≤0.7 than group RIs >0.7, but the difference was not statistically significant (P > .05). Second part of our follow-up studies was to compare hemodynamic RI index of interlobular renal in years of 2000 and 2015, both renal function and blood glucose parameters were fund significantly enhanced in subject group RIs ≤0.7. Compared data of various incidences from first part of study were coherent with second part. (Incidence of DKD and DR was notably lower in group RIs ≤0.7 than group RIs >0.7, difference was statistically significant (P < .05). Incidence of HbA1c ≤7% was higher in group RIs ≤0.7 than group RIs >0.7, but difference was not statistically significant (P > .05). Incidence of PDR was notably lower in group RIs ≤0.7 than group RIs >0.7, but the difference was not statistically significant (P > .05). CONCLUSIONS: RIs of retrobulbar and interlobular renal which would serve as a good predictors for the hemodynamics changes in retrobulbar and intrarenal would assess incidence of DKD and DR during the preclinical stage in long-term range excluding renal function and HbA1c in T2DM patients.

Valproic acid attenuates global cerebral ischemia/reperfusion injury in gerbils via anti-pyroptosis pathways.

Ischemic stroke is the third most common cause of death and the leading cause of disability worldwide in adults. The antiepileptic drug valproic acid (VPA) was reported to protect cerebral ischemia/reperfusion injury. However, the action mechanism of VPA in cerebral ischemia/reperfusion injury has not been fully understood. We explored the action mechanism of VPA in vivo and in vitro. Gerbils were subjected to transient global cerebral ischemic-reperfusion injury, and hippocampal neuron injury was treated with oxygen-glucose deprivation in vitro. Morris water maze test was performed to evaluate the cognitive dysfunction. Histopathological examinations and western blot were performed to evaluate the pyroptosis of neurons. The results showed that VPA attenuated the cognitive dysfunction, pyroptosis of the gerbils suffer from ischemic-reperfusion injury and decreased hippocampal neurons pyroptosis induced by oxygen-glucose deprivation in vitro. In addition, western blot and real-time PCR analysis revealed that VPA modulated the protein expression of apoptosis repressor with caspase recruitment domain (ARC), caspase-1 and IL-1ß/IL-18. Our results suggested that VPA alleviated ischemic/reperfusion injury-mediated neuronal impairment by anti-pyroptotic effects.

Effect of tanshinone IIA on oxidative stress and apoptosis in a rat model of fatty liver.

Oxidative stress is a crucial factor associated with fatty liver disease, which raises the possibility of using antioxidants to improve liver steatosis. Tanshinone IIA (TSIIA) is a traditional Chinese medicine that has been reported to have antioxidant effects in vitro. The present study aimed to investigate whether TSIIA possesses antioxidant effects in vivo and whether TSIIA was able to improve liver steatosis. Hence, the ability of TSIIA to protect rats from liver disease was explored, particularly in regard to antioxidant activity. Rats were fed a high-lipid diet for 90 days, causing severe liver steatosis, both morphologically and biochemically. An increase in reactive oxygen species (ROS) in the liver was exhibited in addition to significantly elevated serum lipids and malondialdehyde (MDA). Furthermore, hepatocyte apoptosis was measured by Hoechst staining, reverse transcription-quantitative polymerase chain reaction and western blot analysis and an increase in hepatocyte apoptosis rate was indicated in mice on a high-fat diet. Following intraperitoneal injection of TSIIA (10 mg/kg/day), liver steatosis was significantly inhibited. In rats receiving TSIIA treatment, less ROS were indicated in the liver and significantly decreased levels of MDA (P<0.05) in serum were exhibited, whereas significantly increased activities of total superoxide dismutase (T-SOD) and glutathione peroxidase (GSH-PX) were observed (P<0.05 and P<0.01, respectively). In addition, the rate of hepatocyte apoptosis was significantly decreased in the TSIIA group (P<0.01). However, TSIIA elicited no effect on serum lipid profiles. These results suggest that TSIIA attenuates oxidative stress by decreasing ROS and MDA production and enhancing the activity of T-SOD and GSH-PX, which may contribute to the inhibition of apoptosis and amelioration of liver steatosis.

[Correlation between autophagy and polarization of macrophages in atherosclerosis plaque in arteriosclerosis obliterans amputees].

This study was designed to investigate the correlation between autophagy and polarization of macrophages in atherosclerosis (AS) plaque in arteriosclerosis obliterans amputees. Femoral artery specimens from arteriosclerosis obliterans amputees were performed hematoxylin and eosin (HE) staining, oil red O and immunofluorescence staining to observe the morphology of atherosclerotic plaque, phenotype of macrophages and autophagy in plaque; using real-time quantitative RT-PCR technology to detect the mRNA level of M1 and M2 type markers in arterial tissue; to analyze polarized signal pathway and autophagy protein levels in macrophages by Western blotting. Arterial specimens staining showed obvious lipid deposition and obvious infiltration of amount of foam cells and inflammatory cells. Macrophages were mainly expression M1 type in percentage in fibrous plaque. Although both M1 and M2 macrophages were upregulated in atheromatous plaque, the increase was dominant in M2 type in percentage. The level of autophagy was significantly higher in the atheromatous plaque than that of fibrous plaque. The expression of tumor necrosis factor- α (TNF-α), monocyte chemotactic protein-1 (MCP-1), inducible nitric oxide synthase (iNOS), interleukin-6 (IL-6) and interleukin-12 (IL-12) mRNA was significantly higher in fibrous plaque than that of atheromatous plaque (P < 0.01 or 0.05), and arginase-1 (Arg-1), transforming growth factor-ß (TGF-ß), CD163 and interleukin-10 (IL-10) mRNA was significantly lower than that in atheromatous plaque (P < 0.01). The levels of p-STAT1 and NF-κB were significantly increased in fibrous plaque (P < 0.01), while p-STAT6 expression was significantly increased in atheromatous plaque (P < 0.01). The level of LC3-II was significantly higher in atheromatous plaque than that in fibrous plaque (P < 0.01). Macrophages in early atherosclerotic plaque were induced to M1 type through p-STAT1/NF-κB pathway and expressed moderate levels of autophagy; while macrophages in advanced plaques were induced to polarization of M2 type through p-STAT6 pathway. M2 macrophages expressed a higher level of autophagy than M1 macrophages.

[Study on Serum Metabolite Profiling in Pi-deficiency Rats Based on LC-MS Technique].

OBJECTIVE: To explore metabolite profiling changes in serum of rats with pi-qi deficiency syndrome (PQDS) and pi-yang deficiency syndrome (PYDS) based on liquid chromatograph-mass spectrometer (LC-MS) technique, and to explore the essence of Pi-deficiency syndrome (PDS) from small molecule metabolite level. METHODS: Totally 21 male SD rats of SPF grade were randomly divided into three groups, the normal control group, the PQDS group, and the PYDS group, 7 in each group. Rats in the PQDS group overate for 1 day and fasted for 2 days. They drank freely and then swam to be exhausted in water at 35 degrees C - 37 degrees C for 15 successive days. The PYDS model was established by the same method for PQDS rats plus drenching 20% Folium sennae water extract (2 mL/100 g), once in the morning and once in the evening for one successive week. After modeling, models were evaluated according to rat general state, changes in body weight and rectal temperature. Serum metabonomic profiles were detected using LC-MS technique. Difference in inter-group metabolite spectrograms was analyzed using orthogonal partial least squares discriminant analysis (OPLS-DA). Potential biomarkers related to syndrome types in rat serum were selected via the parameter of variable importance in the projection (VIP). RESULTS: The weight of rats in the PQDS group and the PYDS group decreased more significantly after modeling. The difference in prepost weight was statistically significant from that of the normal control group (P < 0.01). It was more obviously lowered in the PYDS group than in the PQDS group (P < 0.05). Compared with the normal control group, the rectal temperature of rats in the PYDS group and the PQDS group decreased (P < 0.05, P < 0.01). It decresed more in the PYDS group than in the PQDS group (P < 0.05, P < 0.01). Compared with the normal control group, levels of PC(19:0)/PE(22:0), PC(17:0)/PE(20:0), capric acid, oleic acid, stearic acid, succinic acid, fumaric acid, malic acid, glucose increased; arachidonic acid, linolenic acid, lauric acid, androsterone, 4-heptanone, dihydroxyacetonephosphate (DHAP) (6:0), and uridine decreased in the PYDS group and the PQDS group. Compared with the PQDS group, levels of PC(22:1), PC (22:6), PE (18:0)/PC (15:0), retinol, and deoxycytidine increased significantly in the PYDS group; PC (18:1), PC(19 :3), PC (20:3), PC (17:0)/PE (20:0), PC (19:1)/PE (22:1), PC (19:0)/PE (22:0), PC (17:1)/PE (20: 1), PC (16:1)/PE (19:1), cholic acid, hippuric acid, furoic acid, undecanedicarboxylic acid, palmitoleic acid, hydroxy stearic acid, eicosatrienoic acid, phenylalanine, tyrosine, glutamic acid, serine, carbamoyl aspartic acid, palmitoyl carnitine, tetradecanoyl carnitine, acetylcarnitine, and linoleylcarnitine decreased more significantly in the PYDS group. CONCLUSIONS: Comparative contents of various serum metabolites changed significantly in PQDS and PYQS model groups. Some potential small molecular biomarkers related to PDS were preliminarily identified. These results might provide some data reference for exploring scientific connotation and pathological mechanisms of PDS.

Tanshinone IIA affects the HDL subfractions distribution not serum lipid levels: Involving in intake and efflux of cholesterol.

AIM OF STUDY: Tanshinone IIA is an active component of the traditional Chinese medicine. This study aimed at investigating the mechanism of tanshinone IIA on anti-atherosclerosis, which may be because of that Tanshinone IIA can affect the HDL subfractions distribution and then regulate reverse cholesterol transport. MATERIALS AND METHODS: A model of hyperlipidaemia in rats was used. Tanshinone IIA was given daily after hyperlipidaemia. In vivo, lipid deposition and morphological changes in liver were analyzed; HDL subfractions and lipid level in serum as well as in liver were measured; the expression of genes related to cholesterol intake in liver and peritoneal macrophage cholesterol efflux were evaluated. In vitro, HepG2 cells and THP-1 cells were pretreated with tanshinone IIA and subsequently with ox-LDL to evaluate the total cholesterol and the expression of related genes. RESULTS: Tanshinone IIA reduced the lipid deposition in liver. Moreover, it did not affect the serum lipid levels but reduced the levels of HDL middle subfractions and increased the levels of HDL large subfractions. Furthermore, tanshinone IIA could regulate the expressions of CYP7A1, LDL-R, SREBP2 and LCAT in the liver as well as the ABCA1 and CD36 in macrophage cells which is involving in the cholesterol intake and efflux respectively. It could reduce lipid accumulation caused by ox-LDL induction, and that also regulate the expressions of LDL-R, HMGCR and SREBP2 in HepG2 and ABCA1, CD36 in THP-1 cells. CONCLUSION: A novel finding that tanshinone IIA was not reduce the serum lipid level but affects the HDL subfractions distribution and thereby regulating the intake and efflux of cholesterol.

[Intervention of Huayu Qutan Recipe on liver SREBP-2 signal pathway of hyperlipidemia rats of pi deficiency syndrome].

OBJECTIVE: To explore the intervention of Huayu Qutan Recipe (HQR) on liver SREBP-2 signal pathway of hyperlipidemia rats of Pi deficiency syndrome (PDS). METHODS: Totally 100 SPF grade SD rats were randomly divided into the blank control group, the hyperlipidemia group, the hyperlipidemia treatment group, the PDS hyperlipidemia group, and the PDS hyperlipidemia treatment group, 20 in each group. Common granular forage was fed to rats in the blank control group. High fat forage was fed to rats in the hyperlipidemia group and the hyperlipidemia treatment group. Rats in the PDS hyperlipidemia group and the PDS hyperlipidemia treatment group were treated with excessive labor and improper diet for modeling. They were administered refined lard by gastrogavage (3 mL each time, twice per day) and fed with high fat forage on the odd days, and fed with wild cabbage freely on even days. The modeling lasted for 30 days. Rats in the hyperlipidemia treatment group and PDS hyperlipidemia treatment group were administered with Huayu Qutan Recipe (20 mL/kg) by gastrogavage, once a day, for 30 successive days. Levels of serum cholesterol (TC), triglyceride (TG), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), and serum amylase (AMY) were detected by automatic biochemical analyzer. D-xylose excretion rate was determined using phloroglucinol method. Morphological changes of liver and the lipid deposition in liver were observed using HE stain and oil red O stain respectively, mRNA and protein expression levels of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), cholesterol 7α-hydroxylase 1 (CYP7A1), LDL-R, and sterol regulatory element binding protein-2 (SREBP-2) were detected using real time RT-PCR and Western blotting. RESULTS: Compared with the blank control group, serum levels of TC (1.84 ± 0.19 mmol/L, 2.23 ± 0.43 mmol/L) and LDL-C (0.99 ± 0.24 mmol/L, 1.13 ± 0.56 mmol/L) were higher in the hyperlipidemia group and the PDS hyperlipidemia group, serum levels of HDL-C (0.41 ± 0.66 mmol/L, 0.41 ± 0.11 mmol/L) and AMY activities (351 ± 45 mmol/L, 153 ± 30 mmol/L) were lower, and urinary D-xylose excretion rates were lower (26.9 ± 2.1 ng/mL, 15.0 ± 1.7 ng/mL) (all P < 0.05). Lipid deposition occurred in liver cells. Much fat vacuoles occurred in the cytoplasm. Expression levels of HMGCR, CYP7A1, LDL-R, and SREBP-2 mRNA and proteins in liver significantly decreased (P < 0.01). Compared with the hyperlipidemia group, serum levels of TC and LDL-C significantly increased (P < 0. 05), AMY activities and urinary D-xylose excre- tion rates significantly decreased in the PDS hyperlipidemia group (P < 0.01). A large amount of lipid deposition occurred in liver. The atrophy of liver cells was obviously seen. Expression levels of CYP7A1, LDL-R, and SREBP-2 mRNA and proteins in liver were significantly lower (P < 0.01, P < 0.05). Serum levels of TC and LDL-C significantly decreased (P < 0.05), AMY activities and urinary D-xylose excretion rates significantly increased in the hyperlipidemia treatment group (P < 0.01). Expression levels of CYP7A1, LDL-R, and SREBP-2 mRNA and proteins in liver were significantly increased (P < 0.01, P < 0.05). Compared with the PDS hyperlipidemia group, serum level of TC significantly decreased (P < 0.05), HDL-C levels, AMY activities and urinary D-xylose excretion rates significantly increased in the PDS hyperlipidemia treatment group (P < 0.01),expression levels of CYP7A1, LDL-R, and SREBP-2 mRNA and proteins in liver were significantly increased (P < 0.01). Similar changes occurred in the two treatment groups. CONCLUSIONS: Pi deficiency exacerbates abnormal serum TC level and the lipid deposition in liver. These might be related to regulating expression levels of LDL-R, HMGCR, and CYP7A1 genes in the SREBP-2 signal pathway. HQR could regulate this pathway to intervene abnormal metabolism of TC.

PEGylated VRB plus quinacrine cationic liposomes for treating non-small cell lung cancer.

BACKGROUND: Non-small cell lung cancer (NSCLC) is the most common form of lung cancer, and the treatment effects are usually unsatisfactory. Vinorelbine (VRB) is extensively used in cancer treatment, but it has some disadvantages when used alone. PEGylated liposomes have been extensively used as a delivery carrier for antitumor drugs via prolonging the circulation time in the blood. PURPOSE: The nanostructured liposomes were designed and prepared for treating NSCLC. METHODS: In the liposomes, PEG was modified on the liposomal surface, DC-Chol was used as cationic materials, and VRB plus quinacrine were encapsulated in an aqueous core of the liposomes as an antitumor drug and an apoptosis-inducing agent, respectively. Evaluations were performed on A549 cells, tubular network formations and xenografts of the A549 cells. RESULTS: The PEGylated drugs-loaded cationic liposomes could significantly enhance cellular uptake and selectively accumulate in A549 cells, thus leading to show strongest antitumor efficacy to tumor cells and to tumor-bearing mice. Action mechanisms showed that the enhanced efficacy in treating NSCLC was related to activate caspase 9 and caspase 3, to activate Bax and P53, and to suppress Bcl-2 and Mcl-1. CONCLUSION: The PEGylated VRB plus quinacrine cationic liposomes showed a potential strategy for treating NSCLC.

[Comparative proteomics analysis of the ileum of Pi-Yang deficiency model rats].

OBJECTIVE: Based on proteomics technology, Pi-yang deficiency syndrome (PYDS) correlated differential proteins were screened, thus providing powerful experiment reliance for exploring the essence of PYDS. METHODS: Totally 36 SD rats of SPF grade were randomly divided into the normal control group (n = 16) and the PYDS group (n = 20). The PYDS model rats were induced by improper diet, overstrain, and administration of yang impairing bitter cold herbs. The total proteins of the ileum were separated and extracted from rats in the PYDS group and the normal control group. The differential protein dots were identified using Delyder 2D 6.5 image analysis software by two-dimensional gel electrophoresis (2-DE) technology. The finger print map of corresponding peptide qualities was obtained by applying MALDI TOF/TOF. The differential proteins were identified using Mascot search library. RESULTS: Judged by statistics and fuzzy mathematics, Pi-yang deficiency rat model was successfully established. Eight proteins with differential expressions involving cell skeleton, energy metabolism, and signal transduction, and so on were obtained. Of them, there were 4 up-regulated proteins, i.e., desmin, cytokeratin8 (CK8), pyruvate kinase (PK), and ezrin. Four down-regulated proteins were glyceraldehyde-3-phosphate dehydrogenase (GAPDH), cytokeratin19 (CK19), cytokeratin1 (CK1), and actin. CONCLUSION: The pathogenesis of PYDS might be slowed energy metabolism rate, reduced energy production, changed structure of ileal villin, and weakened absorbing and digestive functions.

Neuroprotective activities of catalpol against CaMKII-dependent apoptosis induced by LPS in PC12 cells.

BACKGROUND AND PURPOSE: Neurodegenerative diseases present progressive neurological disorder induced by cell death or apoptosis. Catalpol, an iridoid glucoside isolated from the root of Rehmannia glutinosa Libosch, is present in a wide range of plant families. Although catalpol is an effective anti-apoptotic agent in LPS-induced neurodegeneration, the underlying mechanism has not been established. Here we have identified some of the mechanisms involved the prevention by catalpol of apoptosis induced by LPS in an experimental model of neurodegeneration in vitro. EXPERIMENTAL APPROACH: Apoptosis was induced by adding LPS (80 ng·mL(-1)) to pheochromocytoma (PC12) cells, pretreated with catalpol for 12 h. We measured intracellular reactive oxygen species (ROS), apoptosis and intracellular calcium concentration ([Ca(2+)]i) by flow cytometry or laser confocal scanning microscopy. We also analysed the protein expression of Bcl-2, Bax and Ca(2+)-calmodulin-dependent protein kinase II (CaMKII)-dependent apoptosis signal-regulating kinase-1 (ASK-1)/JNK/p38 signalling pathway in PC12 cells by Western blot. KEY RESULTS: Catalpol stimulated expression of Bcl-2 and inhibited the expression of Bax. Catalpol also attenuated the increase in Ca(2+) concentration induced by LPS in PC12 cells and down-regulated CaMK phosphorylation. The CaMKII-dependent ASK-1/JNK/p38 signalling cascade was blocked by catalpol. All these changes were accompanied by a decrease of apoptosis induced by LPS in PC12 cells. CONCLUSIONS AND IMPLICATIONS: The data presented here provide new mechanistic insights into the links between the CaMKII-dependent ASK-1/JNK/p38 signalling pathway and the protective effect of catalpol on apoptosis induced by LPS in PC12 cells.

Tanshinone IIA reduces apoptosis induced by hydrogen peroxide in the human endothelium-derived EA.hy926 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Salvia Miltiorrhiza Bunge (also known as herb Danshen in Chinese) is a widely used Chinese herbal medicine. Tanshinone IIA (TSN IIA) is considered to be the most important bioactive ingredient in Danshen and exhibits an anti-atherosclerotic activity. AIM OF STUDY: To evaluate the protective effect of TSN IIA on the human endothelial EA.hy926 cells injured by hydrogen peroxide in vitro and its possible mechanism. MATERIALS AND METHODS: The EA.hy926 cells were incubated for 24h with different concentrations of TSN IIA (5, 10 and 20 µg/µL ) or DMEM. Subsequently, cells were treated with 300 µmol/L H(2)O(2) for another 4h. Then, the percentage of cell viability was evaluated by 3-(4, 5-di-methylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay. The apoptosis of EA.hy926 cells was detected by flow cytometry with AnnexinV-FITC/PI double staining and laser scanning spectral confocal technique. The generation of intracellular reactive oxygen species (ROS) generation was analyzed by flow cytometry. The mRNA expressions of caspase-3, Bcl-2 and Bax were tested by real time-reverse transcription polymerase chain reaction (real time RT-PCR). The protein expression of Bcl-2 and Bax was determined by Western blotting. MDA levels, NO production, LDH leakage, and SOD as well as caspase-3 activities were also measured using standard methods. RESULTS: Loss of cell viability and excessive cell apoptosis were observed in EA.hy926 cells after 4h of challenge with H(2)O(2) (300 µmol/L). However, cell apoptosis was attenuated in different concentrations of TSN IIA (5, 10 and 20 µg/µL) pretreated cells. Furthermore, TSN IIA markedly inhibited the elevation of ROS evoked by H(2)O(2). Real time RT-PCR and Western blotting analysis showed that TSN IIA significantly decreased the expressions of pro-apoptotic proteins (Bax and caspase-3) while significantly increased the expression of anti-apoptotic protein Bcl-2, and resulted in obvious reduction of Bax/Bcl-2 ratio in EA.hy926 cells induced by H(2)O(2). CONCLUSION: These observations provide preliminary evidence that TSN IIA protects EA.hy926 cells against H(2)O(2) damage, which is mainly associated with the ROS generation, followed by the imbalance of the Bax/Bcl-2 ratio, and caspase-3 activation leading to apoptosis.

[Experimental study on the ileum tissue proteins differentially expressed in Pi-qi deficiency syndrome rats].

OBJECTIVE: To identify the ileum tissue proteins differentially expressed in Pi-qi deficiency syndrome rats using proteomic approach. METHODS: Thirty-seven rats were randomly divided into 3 groups, i. e., the normal control group (Group 1), the Pi-qi deficiency syndrome model group (Group 2), and the reserpine group (Group 3). The Pi-qi deficiency syndrome model was established using excessive exerting combined with irregular diet, and peritoneally injecting Reserpine Injection (1 mg/mL) respectively. The ileum tissues were separated after identified fuzzy method. The differentially expressed proteins were separated with two dimensional electrophoresis (2-DE), analyzed by Mass Spectrometry, and identified by MASCOT Software. RESULTS: Three proteins were differentially expressed in Group 2 and nine proteins were differentially expressed in Group 3 (P < 0.05). CONCLUSION: Pi-qi deficiency syndrome was closely related with decreased expression of albumin as well as increased expressions of trypsin and glucose-regulated protein 78.

[Effect of tanshinone II A on TLR4 and TNF-α of endothelial cells induced by LPS].

AIM: To observe the influence of tanshinone II A on the expression of toll like receptor 4(TLR4) and tumor necrosis factor-α(TNF-α) in cultured human umbilical veins endothelial cells EA.hy926 induced by LPS, and to investigate the molecular mechanism of tanshinone II A against atherosclerosis(AS). METHODS: The EA.hy926 cells were cultured in vitro. The mRNA and protein expression of TLR4 were tested by RT-PCR and immunocytochemistry technique respectively. The mRNA and protein expression of TNF-α were detected by RT-PCR and ELISA technique respectively. RESULTS: The expression of TLR4 and TNF-α were both increased compared to normal group after stimulation of LPS(P<0.05). The expression of TLR4 and TNF-α in tanshinone II A group were obviously inhibited compared to stimulation group(P<0.05). CONCLUSION: The molecular mechanism of tanshinone II A against AS is probably to inhibit the expression of TLR4 and TNF-α.

[Study on the role of Wnt/beta-catenin signaling transduction pathway in hepatocellular carcinoma cell line HepG2 and L02 cell line].

AIM: To explore the possible mechanism of action of Wnt/beta-catenin signaling pathway in hepatocarcinogenesis by investigating the different expressions of the main members on this signaling pathway in hepatocellular carcinoma cell line HepG2 and L02 cell line. METHODS: The mRNAs of Wnt1, Wnt4, beta-catenin, cyclinD1 and c-myc genes were amplified by means of semiquantitative reverse transcription polymerase chain reaction (RT-PCR) in normal liver cell line L02 and hepatocellular carcinoma cell line HepG2, respectively. At the same time, the proteins expression of beta-catenin which was the key member in the Wnt/beta-catenin signaling pathway was examined by immunocytochemical method and Western blot technique. RESULTS: In normal liver cell line L02, the mRNAs of Wnt1, Wnt4, cyclin D1 and c-myc genes were not detected except for the gene of beta-catenin. In hepatocellular carcinoma cell line HepG2, the mRNAs of Wnt1, beta-catenin, cyclin D1 and c-myc genes were detected except for the gene of Wnt4. Meanwhile, found that beta-catenin proteins were accumulated in the cytoplasm and/or nucleus in HepG2 but only in cell membrane in L02.Using Western blot technique, found that beta-catenin proteins expression was higher in HepG2 than in L02. CONCLUSION: The Wnt/beta-catenin signaling transduction pathway is activated with aberrant expression of Wnt1 in hepatocellular carcinoma cell line HepG2.