Efficacy and Mechanism of a Chinese Classic Prescription of Yueju in Treating Nonalcoholic Steatohepatitis and Protecting Hepatocytes from Apoptosis.

Yueju, a famous classic Chinese prescription, has been extensively used in treating depression syndromes for hundreds of years. Recent studies have reported that Yueju showed good effects in treating metabolic diseases, such as obesity and hyperlipidemia. Nonalcoholic steatohepatitis (NASH), which leads to cirrhosis and severe cardiovascular diseases, is closely linked to obesity and abnormal lipid metabolism. In this study, Yueju could decrease the levels of alanine aminotransferase, aspartate transaminase, triglyceride, cholesterol, and low-density lipoprotein-C but increase the high-density lipoprotein-C in the serum of the NASH rat model induced by high-fat and high-cholesterol diet. Yueju could alleviate hepatosteatosis by increasing the phosphorylation of acetyl-CoA carboxylase and inhibiting the expression of fatty acid synthase and stearoyl-CoA desaturase 1. Yueju downregulated the expression of α-smooth muscle actin and collagen type 1A1, ameliorating the liver fibrilization. Yueju could also protect the hepatocytes from apoptosis by upregulating antiapoptosis protein Bcl-2 and X-linked inhibitor of apoptosis protein and downregulating apoptotic proteins Bax and cleaved poly ADP-ribose polymerase. Thus, Yueju could improve liver function, regulate lipid metabolism, alleviate hepatosteatosis and fibrosis, and protect hepatocytes from apoptosis against NASH. Yueju may be used as an alternative effective medicine for NASH treatment.

Protective effects of a G. lucidum proteoglycan on INS-1 cells against IAPP-induced apoptosis via attenuating endoplasmic reticulum stress and modulating CHOP/JNK pathways.

Fudan-Yueyang-G. lucidum (FYGL) is a water-soluble macromolecular proteoglycan extracted from Ganoderma lucidum which has been used for health promotion for a long time in China. The aim of this study was to investigate the protective effects of FYGL on INS-1 rat insulinoma beta cells against IAPP-induced cell apoptosis, as well as the underlying mechanisms. The results showed that apoptotic cells were significantly increased when incubated with islet amyloid polypeptide (IAPP). However, cytotoxicity of IAPP was significantly attenuated by co-incubation of the cells with FYGL. The results of RT-PCR showed that mRNA expression of caspase-3, caspase-12 and C/EBP homologous protein (CHOP) in IAPP-treated cells were inhibited by FYGL. Moreover, FYGL significantly prevented the IAPP-induced abnormal expression of inositol-requiring protein-1α (IRE1α), protein kinase RNA (PKR)-like ER kinase (PERK), activating transcription factor 6 (ATF6), as well as suppressed the activation of CHOP and c-Jun N-terminal kinase (JNK). Taken together, our results suggest that FYGL protects INS-1 pancreatic beta cells against IAPP-induced apoptosis through attenuating endoplasmic reticulum stress (ERS) and modulating CHOP/JNK pathways.

[Treating Type 2 Diabetes Mellitus Patients Complicated with Metabolic Syndrome by Benefiting Qi Dissolving Method].

Objective To observe clinical efficacy of Yiqi Huaju Recipe (YHR) combined routine Western medical treatment on type 2 diabetes mellitus (T2DM) patients complicated metabolic syndrome (MetS). Methods Totally 96 T2DM patients complicated MetS were assigned to the treatment group (YHR +routine Western drugs) and the control group (placebo +routine Western drugs) according to random digit table, 48 cases in each group. The therapeutic course for all was 12 weeks. Body mass index (BMI) , waistline, waist-hip ratio (WHR) , fasting plasma glucose (FPG) , 2 h postprandial plasma glucose (2 h PPG) , glycosylated hemoglobin A1c (HbAlc) , homeostasis model assessment of insulin resistance (HOMA-IR) , blood lipids, blood pressure, disease transformation of MetS, changes of con- stituent numbers were detected before and after treatment. Results BMI, WHR, waistline obviously decreased in the treatment group after treatment, with statistical difference as compared with the control group (P<0.01 , P <0.05). Post-treatment FPG, 2 h PPG, HbAlc, HOMA-IR, systolic blood pressure (SBP), diastolic blood pressure (DBP) , and mean artery pressure (MAP) obviously decreased in the two groups, but more obviously in the treatment group (P <0. 05). Post-treatment total cholesterol (TC) , low density lipoprotein cholesterol (LDL-C) , and triglycerides (TG) all obviously decreased in the two groups , but TG decreased more obviously in the treatment group (P <0. 05). High density lipoprotein cholesterol (HDL-C) obviously increased in the treatment group (P <0. 05). Patient numbers of central obesity, uncontrolled hypertension, and uncontrolled diabetes obviously decreased and constituent numbers were obviously reduced in the treatment group after treatment, with better efficacy than those of the control group (P <0. 01 , P <0. 05). Conclusions YHR plus routine Western drugs could further reduce blood glucose, and had comprehensive interventional effects on multiple cardiovascular risk factors such as central obesity, blood lipids, and blood pressure in T2DM patients complicated with MetS. Its mechanism might be possibly correlated with improving insulin resistance and elevating insulin sensitivity of peripheral tissues.

Yiqi Huaju formula, a Chinese herbal medicine, reduces arterial pressure in salt­sensitive hypertension by inhibiting renin­angiotensin system activation.

Hypertension is a chronic disease with a high prevalence, and is associated with a high risk of vascular disease and premature death. Traditional Chinese medicine has been administered to treat hypertension for many years. In the present study, the effects of Yiqi Huaju formula (YQ; a compound used in traditional Chinese herbal medicine) were observed in salt­sensitive hypertension, which was induced by a high­salt and high­fat (HSF) diet and the potential mechanism was investigated. YQ was prepared from five plant extracts and was dissolved in normal sodium chloride prior to use. Male Sprague­Dawley rats were randomly divided into three groups, and fed either a normal diet (control), an HSF diet or an HSF diet with YQ. At week eight, blood pressure was measured and 24­h urine samples were collected from all of the rats. The rats were subsequently sacrificed, and their blood was collected for biochemical analyses and kidney tissue samples were dissected for the immunohistochemical assay. YQ was observed to decrease the high arterial pressure and serum total cholesterol level, which had been induced by the HSF diet. It also enhanced the excretion of urinary angiotensinogen, Na+, and decreased the loss of urinary aldosterone, K+ and microalbuminuria. In addition, YQ inhibited the high mRNA expression level of renal renin, angiotensin II (Ang II), and Ang II receptor, type 1 (AT1R), and inhibited the protein expression of renal AT1R and Ang II receptor type 2, which had been induced by the HSF diet. These results indicate that YQ may reduce the arterial pressure in salt­sensitive hypertension via the inhibition of renin­angiotensin system activation.

[Effect of yiqi huaju recipe combined with routine therapy in treating hypertension patients with metabolic syndrome: a clinical study].

OBJECTIVE: To investigate the therapeutic effect of Yiqi Huaju Recipe (YHR) combined with routine therapy on the blood pressure, the blood pressure variability and other cardiovascular risk factors in hypertension patients complicated with metabolic syndrome (MetS). METHODS: Totally 43 hypertension patients complicated with MetS were recruited in this study and randomly assigned to the treatment group (22 cases, treated with basic routine treatment +YHR) and the control group (21 cases, treated with basic routine treatment + placebo). The treatment course was 12 weeks. Detected were parameters such as 24-h ambulatory blood pressure monitoring (ABPM), body mass index (BMI), waist circumference (WC), waist to hip ratio (WHR), homeostatic model assessment for insulin resistance (HOMA-IR), fasting glycosylated hemoglobin A1c (HbA1c), fasting plasma glucose (FPG), 2 h postprandial plasma glucose (2 h PPG), fasting plasma insulin (FPI), serum lipid, etc. RESULTS: The anthropometric parameters and plasma glucose levels (except HbAlc) were obviously lowered after treatment than before treatment in the treatment group (P < 0.01, P < 0.05). Besides, better effects were obtained in the WC, WHR, 2 h PPG, FPI and HOMA-IR (P < 0.05). The average blood pressure amplitude, the blood pressure variability, and blood pressure load at any time point were more obviously improved in the two groups after treatment than before treatment (P < 0.01, P < 0.05). Besides, partial indices were better in the treatment group than in the control group (P < 0.01, P < 0.05). CONCLUSIONS: YHR combined with routine therapy exhibited better effect on reducing the blood pressure amplitude, the blood pressure variability, and the blood pressure load in hypertension patients complicated with MetS. It could also effectively decrease the risk of other vascular disease.

Effects of Chinese herbal medicine Yiqi Huaju Qingli Formula in metabolic syndrome patients with microalbuminuria: a randomized placebo-controlled trial.

BACKGROUND: Microalbuminuria (MAU) is a key component of metabolic syndrome (MetS) and is an early sign of diabetic nephropathy as well. Although routine Western medicine treatments are given to MetS patients to control high blood pressure, hyperglycemia and dyslipidemia, some patients still experience progressive renal lesions and it is necessary to modify and improve the treatment strategy for MetS patients. OBJECTIVE: To investigate the efficacy of Yiqi Huaju Qingli Herb Formula, a compound traditional Chinese herbal medicine, in MetS patients with MAU when it is combined with routine Western medicine treatment. DESIGN, SETTING, PARTICIPANTS AND INTERVENTIONS: Sixty patients with MetS were randomized into the Chinese herbal formula group (CHF, Yiqi Huaju Qingli formula treatment in combination with Western medicine) and control group (placebo in combination with Western medicine). All treatments were administered for 12 weeks. MAIN OUTCOME MEASURES: Urinary microalbumin (MA), urinary albumin-to-creatinine ratio (UACR), 24-hour total urine protein (24-hTP), body mass index (BMI), waist circumference (WC), waist-to-hip ratio (WHR), fasting plasma glucose (FPG), 2-hour postprandial plasma glucose (2-hPPG), glycosylated hemoglobin (HbA1c), homeostasis model assessment for insulin resistance (HOMA-IR), blood lipid profile and blood pressure were observed. RESULTS: Compared with the control group, CHF treatment significantly decreased BMI (P<0.05), WC (P<0.01) and WHR (P<0.01). Both groups had significant decreases in FPG, 2-hPPG, HbA1c, HOMA-IR, MA, and UACR, with CHF treatment showing better effects on these parameters compared with the control treatment (P<0.05). Both treatments significantly reduced the levels of total cholesterol, low-density lipoprotein cholesterol and triacylglycerol (TAG), and a greater reduction in TAG was observed with CHF treatment (P<0.05). The level of high-density lipoprotein cholesterol did not change in the control group after treatment (P>0.05), whereas it significantly increased with CHF treatment (P<0.01). Compared with before the treatment, significant decreases in systolic blood pressure, diastolic blood pressure and mean arterial blood pressure were observed in both groups (P<0.01). However, there was no significant difference between the two groups (P>0.05). CONCLUSION: Combined treatment of Yiqi Huaju Qingli Formula and Western medicine significantly alleviated MAU, which may correlate with the improvement of insulin sensitivity and glucose and lipid metabolism. TRIAL REGISTRATION IDENTIFIER: This trial was registered in the Chinese Clinical Trial Registry with the identifier ChiCTR-TRC-11001633.

Effects of Chinese herbal medicine Yiqi Huaju Formula on hypertensive patients with metabolic syndrome: a randomized, placebo-controlled trial.

BACKGROUND: Patients with hypertension coupled with metabolic syndrome (MetS) are among the high risk population in cardiovascular and cerebrovascular diseases. To reduce the prevalence of cardiovascular and cerebrovascular diseases, it is essential to appropriately control blood pressure together with other cardiovascular risk factors. OBJECTIVE: The current study was designed to investigate the therapeutic effects on blood pressure, blood pressure variability and other cardiovascular risk factors by giving Yiqi Huaju Formula, a compound traditional Chinese herbal medicine, in addition to routine treatment to hypertensive patients coupled with MetS. DESIGN, SETTING, PARTICIPANTS AND INTERVENTIONS: A total of 43 patients with hypertension coupled with MetS were recruited into this study. The enrolled patients were randomly divided into the Chinese herbal formula group (anti-hypertensive drugs plus Yiqi Huaju Formula, CHF) and the control group (anti-hypertensive drugs plus placebo). The CHF group enrolled 22 patients while the control group received 21 cases. Treatments were given for 12 weeks in both groups. MAIN OUTCOME MEASURES: Parameters examined include 24-hour ambulatory blood pressure monitoring, body mass index, waist circumference, waist-to-hip ratio, homeostatic model assessment for insulin resistance (HOMA-IR), fasting glycosylated hemoglobin (HbA1c), fasting plasma glucose, 2-hour postprandial plasma glucose (PPG), fasting plasma insulin, serum lipid, etc. RESULTS: Compared with the control group, the CHF group had significant improvement (P<0.01) in anthropometric parameters, FPG, HOMA-IR, blood pressure amplitude, blood pressure variability and blood pressure load. CONCLUSION: This study showed that integrated traditional Chinese and Western medicine treatment can achieve better results in controlling blood pressure as well as other cardiovascular risk factors. The mechanism of controlling of blood pressure may be associated with the improvement of insulin sensitivity due to the Yiqi Huaju intervention. TRIAL REGISTRATION IDENTIFIER: ChiCTR-TRC-11001633.

Effect of a novel proteoglycan PTP1B inhibitor from Ganoderma lucidum on the amelioration of hyperglycaemia and dyslipidaemia in db/db mice.

Protein tyrosine phosphatase 1B (PTP1B) is implicated in the negative regulation of the insulin signalling pathway by dephosphorylating the insulin receptor (IR) and IR substrates. Ganoderma lucidum has traditionally been used for the treatment of diabetes in Chinese medicine; however, its anti-diabetic potency and mechanism in vivo is still unclear. Our previously published study reported a novel proteoglycan PTP1B inhibitor, named Fudan-Yueyang-Ganoderma lucidum (FYGL) from G. lucidum, with a half-maximal inhibitory concentration (IC50) value of 5·12 (sem 0·05) µg/ml, a protein:polyglycan ratio of 17:77 and 78 % glucose in polysaccharide, and dominant amino acid residues of aspartic acid, glycine, glutamic acid, alanine, serine and threonine in protein. FYGL is capable of decreasing plasma glucose in streptozotocin-induced diabetic mice with a high safety of median lethal dose (LD50) of 6 g/kg. In the present study, C57BL/6 db/db diabetic mice were trialed further using FYGL as well as metformin for comparison. Oral treatment with FYGL in db/db diabetic mice for 4 weeks significantly (P < 0·01 or 0·05) decreased the fasting plasma glucose level, serum insulin concentration and the homeostasis model assessment of insulin resistance. FYGL also controlled the biochemistry indices relative to type 2 diabetes-accompanied lipidaemic disorders. Pharmacology research suggests that FYGL decreases the plasma glucose level by the mechanism of inhibiting PTP1B expression and activity, consequently, regulating the tyrosine phosphorylation level of the IR ß-subunit and the level of hepatic glycogen, thus resulting in the improvement of insulin sensitivity. Therefore, FYGL is promising as an insulin sensitiser for the therapy of type 2 diabetes and accompanied dyslipidaemia.

[Effects of Pollen Typhae total flavones on mRNA expressions of peroxisome proliferator-activated receptors in 3T3-L1 adipocytes].

OBJECTIVE: To explore the effects of Pollen Typhae total flavones (PTF) on the mRNA expressions of peroxisome proliferator-activated receptors (PPARs) alpha,gamma, beta/delta so as to analyze its possible mechanism in improving the insulin sensitivity of 3T3-L1 adipocytes. METHODS: Adipocytes were treated with PTF, and expressions of PPARalpha, PPARgamma, PPARbeta/delta mRNAs relating to the adipocyte glucose and lipid metabolism were determined by reverse transcription polymerase chain reaction. RESULTS: PTF obviously up-regulated the expressions of PPARalpha and PPARgamma mRNAs, all showing significant differences as compared with those in the normal control group (P<0.01). CONCLUSION: With its function as an insulin sensitizer, PTF may enhance the PPARalpha and PPARgamma mRNA expressions in 3T3-L1 adipocytes.

[Effects of Yiqi Sanju Formula on non-alcoholic fatty liver disease: a randomized controlled trial].

OBJECTIVE: To observe the therapeutic effects of Yiqi Sanju Formula (YQSJF), a compound traditional Chinese herbal medicine, in treatment of non-alcoholic fatty liver disease (NAFLD). METHODS: Sixty-seven patients diagnosed with NAFLD were randomly divided into two groups: YQSJF-treated group (39 cases) and placebo group (28 cases). The NAFLD patients in the two groups were treated with YQSJF and placebo respectively for 3 months. Clinical symptoms, the CT ratio of liver-spleen, body mass index (BMI), waist circumference, homeostatic model assessment for insulin resistance (HOMA2-IR) and the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumor necrosis factor-alpha (TNF-alpha), high sensitivity C-reactive protein (hs-CRP), triglyceride (TG), total cholesterol (TC), high-density lipoprotein (HDL) and low-density lipoprotein (LDL) were evaluated before and after treatment. RESULTS: After treatment, the clinical symptoms were improved and the levels of BMI, waist circumference, HOMA2-IR, ALT, AST, TG and TC were decreased significantly in the YQSJF-treated group (P<0.05). The CT ratio of liver-spleen in the YQSJF-treated group was increased significantly as compared with the placebo group (P<0.01).

[Pollen Typhae total flavones inhibit expression of interleukin-6 in C2C12 skeletal muscle cells cultured with palmitate].

OBJECTIVE: To observe the effects of Pollen Typhae total flavones (PTF) on expression of interleukin-6 (IL-6) mRNA and protein secretion in C2C12 cell strain of skeletal muscle cells cultured with palmitate, and to explore the mechanism of PTF in relieving insulin resistance (IR). METHODS: The IR of C2C12 cells was induced by co-culturing with palmitate. The C2C12 cells were divided into normal control group, untreated group, PDTC (a nuclear factor-kappaB inhibitor) treated group, rosiglitazone (ROS)-treated group, ROS+ PDTC-treated group, PTF-treated group and PTF+PDTC-treated group. Sixteen hours after culture, the transportation rate of glucose was observed by (3)H-deoxyglucose uptake method; IL-6 mRNA expression in C2C12 cells was assayed by real time polymerase chain reaction (Real-time PCR), and level of IL-6 protein secretion in culture supernatant was detected by enzyme linked immunosorbent assay. RESULTS: Compared with the normal control group, the transportation rate of glucose of cells in untreated group was decreased 30.43% after 16-hour palmitate culture, and was increased 32.39% in the PTF-treated group. Compared with the untreated group, the levels of IL-6 mRNA expression in cells and IL-6 protein secretion in supernatant were significantly decreased in the PTF-treated group (P<0.05). The levels of IL-6 mRNA expression in cells and IL-6 protein secretion in supernatant were increased in PTF+PDTC-treated group as compared with PFT-treated group (P<0.05). CONCLUSION: PTF can inhibit the IL-6 mRNA expression and IL-6 protein secretion via nuclear factor-kappaB pathway in C2C12 skeletal muscle cells, which may be one of its mechanisms in relieving inflammation conditions and insulin resistance in C2C12 skeletal muscle cells.

[Effects of berberine on cell proliferation, peroxisome proliferation activated receptor gamma, CAAT/enhancer binding protein mRNA and protein expression in 3T3-L1 pre-adipocytes].

OBJECTIVE: To study the effect and mechanism of berberine (BER) on the proliferation and differentiation of adipocytes. METHODS: The proliferation of 3T3-L1 pre-adipocytes was detected by XTT method. Lipid droplets accumulated in the cytoplasm of adipocytes in the differentiating process were observed by oil red O staining and quantified by colorimetry. The expressions of peroxisome proliferation activated receptor gamma (PPARgamma), CAAT/enhancer binding protein alpha (C/EBPalpha) mRNA and protein were detected by Real-time PCR and Western blotting respectively. RESULTS: Intervention with BER in concentration below 10 micromol/L for 24 h showed insignificant effect on the proliferation of adipocytes, as compared with that in the control group (P > 0.05); but that in concentrations 20, 40 and 80 micromol/L revealed significant suppressive effect; that in different concentrations acting for 48 h and 72 h could affect the proliferation and the effect displayed a dose-dependent manner, i. e. the higher the concentration of BER, the more apparent the suppression, showing significant difference as compared with those in the control group (P <0.05 or P <0.01). The pre-adipocyte treated with 10 micromol/L BER showed that the lipid droplets in the cytoplasm significantly lessened, so did the expression of differentiation related factor PPAR gamma mRNA as well as the expressions of C/EBPalpha mRNA and protein, as compared with those in the blank control group and the group intervened with rosiglitazone, the difference was significant (P < 0.05 or P < 0.01). CONCLUSIONS: BER can suppress the proliferation and differentiation of 3T3-L1 pre-adipocytes, reduce the accumulation of lipid drops in the adipocyte differentiating process, which may be associated with its effects in decreasing the expressions of adipocyte differentiation related gene PPARgamma, C/EBPalpha mRNA and protein. The study provides a basis for applying BER on the prevention and treatment of such metabolic related diseases as obesity.

[Clinical research of Yiqi Sanju Formula in treating central obese men at high risk of metabolic syndrome].

OBJECTIVE: To observe the therapeutic effects of Yiqi Sanju Formula (YQSJF), a compound Chinese herbal medicine, on central obese men at high risk of metabolic syndrome (MS). METHODS: Compared with 30 healthy male volunteers, 45 central obese men were separated randomly into two groups and received the interventions with YQSJF and placebo respectively for 10 weeks. Baseline characteristics, insulin resistance, inflammation cytokines and plasminogen activator inhibitor-1 (PAI-1) and tissue plasminogen activator (t-PA) were evaluated before and after treatment. RESULTS: The score of homeostatic model assessment for insulin resistance (HOMA-IR) and the levels of C reactive protein (CRP), free fatty acid (FFA) and PAI-1 in obese men were higher than those in the control group, while t-PA was lower. After treatment, compared with placebo group, body mass index, waist, and waist-to-hip ratio were decreased significantly in subjects who received YQSJF (P<0.01). The score of HOMA-IR and the levels of CRP, FFA and PAI-1 were decreased significantly, and the level of t-PA was increased significantly (P<0.01). CONCLUSION: YQSJF can reduce obesity and insulin resistance in central obese men at high risk of MS and improve inflammation and fibrinolysis, which indicates that it can reduce the risk of atherosclerosis.

[Effects of Pollen Typhae total flavone on glucose and lipid metabolism in 3T3-L1 adipocytes].

OBJECTIVE: To observe the effects of Pollen Typhae total flavone (PTF) on glucose and lipid metabolism in 3T3-L1 adipocytes. METHODS: The content of glucose which disappeared from the culture medium after incubation with drugs for 24 hours was determined as glucose consumption of the cells. The activity of cells was detected by XTT method. The transport of glucose was observed by (3)H-glucose uptake method. The efflux of free fatty acid (FFA) from adipocytes was observed by the concentration of FFA in the culture medium. RESULTS: The glucose concentration in culture medium was significantly decreased with a concentration-dependent effect, when PTF concentrations were from 0.025 g/L to 0.4 g/L. The toxic effect on cells appeared while PTF concentration was 0.4 g/L, and the MTT value decreased. PTF also significantly increased glucose transportation in the 3T3-L1 adipocytes as rosiglitazone (ROS) did. At the same time, FFA concentration in culture medium was significantly decreased as compared to the normal control group, while ROS-treated group did not show any difference. CONCLUSION: PTF can increase insulin sensitivity by increasing glucose transportation and consumption in the 3T3-L1 adipocytes as well as decreasing the FFA efflux from the cells.

Effects of Pinus massoniana bark extract on cell proliferation and apoptosis of human hepatoma BEL-7402 cells.

AIM: To study the effects of Pinus massoniana bark extract (PMBE) on cell proliferation and apoptosis of human hepatoma BEL-7402 cells and to elucidate its molecular mechanism. METHODS: BEL-7402 cells were incubated with various concentrations (20-200 microg/mL) of PMBE for different periods of time. After 48 h, cell proliferation was determined by 3-(4,5-dimethyl-thiazolyl-2)-2,5-diphenyl tetrazolium bromide (MTT) assay. Apoptosis was evaluated by morphological observation, agarose gel electrophoresis, and flow cytometry analysis. Possible molecular mechanisms were primarily explored through immunohistochemical staining. RESULTS: PMBE (20-200 microg/mL) significantly suppressed BEL-7402 cell proliferation in a time- and dose-dependent manner. After treatment of BEL-7402 cells with 160 microg/mL PMBE for 24, 48, or 72 h, a typical apoptotic "DNA ladder" was observed using agarose gel electrophoresis. Nuclear condensation and boundary aggregation or split, apoptotic bodies were seen by fluorescence and electron microscopy. Sub-G1 curves were displayed by flow cytometry analysis. PMBE decreased the expression levels of Bcl-2 protein in a time-dependent manner after treatment of cells with 160 microg/mL PMBE. CONCLUSION: PMBE suppresses proliferation of BEL-7402 cells in a time- and dose-dependent manner and induces cell apoptosis by possibly downregulating the expression of the bcl-2 gene.