The effect of 2,3,4',5-tetrahydroxystilbene-2-O-ß-D-glucoside on pressure overload-induced cardiac remodeling in rats and its possible mechanism.

The aim of the present study was to investigate the effects of 2,3,4',5-tetrahydroxystilbene-2-O-beta-D-glucoside, an active component extracted from Polygonum multiflorum, on pressure overload-induced cardiac remodeling in rats. A rat model with cardiac remodeling was induced by abdominal aortic banding. 2,3,4',5-Tetrahydroxystilbene-2-O-beta-D-glucoside (30, 60, 120 mg/kg/day) was administered 3 days after abdominal aortic banding and continued for 30 days. The abdominal aortic banding-treated rats had significantly elevated blood pressure, left ventricular hypertrophy, and myocardial fibrosis. Left ventricular hypertrophy was characterized by an increase in the ratios of the heart and left ventricular weights to body weight, and increased myocyte cross-sectional areas, hypertrophic ventricular septum, and left ventricular posterior wall. The accumulation of myocardial interstitial perivascular collagen and elevated cardiac hydroxyproline content indicated myocardial fibrosis. The pathological changes above were attenuated by 2,3,4',5-tetrahydroxystilbene-2-O-beta-D-glucoside. Additionally, it markedly reduced collagen I and III expressions and regulated matrix metalloproteinase-2,9 and inhibitors of metalloproteinase expressions, as markers of myocardial fibrosis. Furthermore, we explored the underlying mechanisms for such effects of 2,3,4',5-tetrahydroxystilbene-2-O-beta-D-glucoside. The results showed that it significantly reduced myocardium angiotensin II, enhanced the activities of superoxide dismutase and glutathione peroxidase in serum and myocardial tissue, as well as inhibited protein expression of transforming growth factor-ß1 and phosphorylation of extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase in the myocardial tissue. Our results suggest that 2,3,4',5-tetrahydroxystilbene-2-O-beta-D-glucoside could prevent cardiac remodeling induced by pressure overload in rats. The underlying mechanisms may be related to a decreasing angiotensin II level, an antioxidant effect of the tested compound, suppression of transforming growth factor-ß1 expression, and inhibition of extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase activation.

The role of Toll-like receptor 4 on inflammation and Aß formation in cortex astrocytes.

To investigate the role and possible molecular mechanism of astrocytes in inflammation and amyloid ß-protein (Aß) formation, in this research, by using LPS to stimulate cultured rat astrocytes in vitro with or without anti-Toll-like receptor 4 (TLR4) antibody pretreatment, we first detected the TLR4, TNF-α, IL-1ß, ß-amyloid precursor protein (ß-APP) and ß-site APP clearing enzyme 1 (BACE1) mRNA with real-time PCR, and TLR4, NF-κB/P65 protein in cultured astrocytes by Western blot, and then further probed the translocation of NF-κB/P65 using immunofluorescence and the contents of TNF-α, IL-1ß and Aß in culture supernatant through ELISA. We found that all of these indexes increased at different degrees after LPS-stimulation. However, if pretreatment with anti- TLR4 antibody, such stimulating effects of LPS on the nuclear translocation of NF-κB/P65 and TNF-α, IL-1ß, Aß contents in astrocytic culture supernatant were reduced significantly or disappeared in comparison with the group with only LPS-administration. Our results suggest that TLR4 in astrocytes might play an important role in the inflammation and Aß formation through the TLR4/NF-κB signaling pathway, thus providing new knowledge and understanding of the inflammatory hypothesis of AD pathogenesis.

[Inhibitory effects of anisodamine and pentoxifylline on the expression of lipopolysaccharide -induced intercellular adhesion molecule-1 in rat cardiac muscle].

OBJECTIVE: To investigate the inhibitory effects of anisodamine (654-2) and pentoxifylline (PTX) on the expression of lipopolysaccharide (LPS)-induced intercellular adhesion molecule-1 (ICAM-1) in rat cardiac muscle in vivo. METHODS: The animals were randomly divided into five groups (each n=6): (1) normal control group, (2) model group, (3) 654-2 treated group, (4) PTX treated group and (5) 654-2+PTX treated group. The endotoxemia model was reproduced by intravenous injection LPS 5 mg/kg. The expression of ICAM-1 protein in rat cardiac muscle was assayed by Western blotting at 0, 2, 4, 6, 8, 10 hours after intravenous LPS injection. Then the expression of ICAM-1 protein in different groups was assayed at different time points. RESULTS: The changes in expression of ICAM-1 in rat cardiac muscle after LPS injection were in a time-dependent pattern, gradually elevating to approach the peak at 6 th, then it lowered, but it still appeared at 10 hours (P<0.05). Western blotting also showed that ICAM-1 protein with decreased with pre-treatment of 654-2 or PTX respectively (both P<0.01). It was reduced to a much lower level when the animals were pretreated with a combination of 654-2 and PTX, compared with the group of 654-2 alone or PTX alone (both P<0.05). CONCLUSION: The combination of 654-2 and PTX may play a protective role in rat against injury to cardiac muscle induced by LPS in vivo via inhibiting the production of ICAM-1 protein.

[Expression of tumor suppressor PTEN in hypertrophy cardiomyocyte].

OBJECTIVE: To investigate the role of tumor suppressor PTEN in cardiac hypertrophy, the expression of PTEN mRNA and protein was analyzed in the tissue of left ventricle in abdominal aorta constricted-induced cardiac hypertrophic rats which treated with and without captopril. The expression of PTEN mRNA and protein in cultured neonatal rat cardiomyocyte treated with AngII was studied. METHODS: SD rats were divided into control group, hypertrophy group and captopril group. The expression of PTEN in different groups at 2 and 4 weeks after operation as well as in cultured neonatal rat cardiomyocyte treated with AngII was detected by RT-PCR and Western blot. The localization of PTEN in left ventricle and cultured cardiomyocyte was determined by immunohistochemistry. RESULTS: (1) Compared with control group, the expressions of PTEN mRNA and protein in left ventricle of hypertrophy group as well as in cultured cardiomyocyte treated with AngII were reduced. (2) Compared with hypertrophy group, the expressions of PTEN mRNA and protein in left ventricle of captopril group were upregulated, which were similar to those of control group. (3) Positive immunohistochemical staining of PTEN was located in the nucleus of cardiomyocytes. CONCLUSION: PTEN may play a negative regulation role in the process of cardiac hypertrophy, and the role of PTEN may be closely related with renin-angiotensin system.

[Effect of captopril on expression of PTEN in aorta of aortic-induced hypertensive rats].

This study inquired about the role of tumor suppressor PTEN in the arterial remodeling of Ang II induced hypertension. The expression of PTEN of aorta was examined in the aortic-constricted hypertensive rats (hypertension group), in the aortic-constricted hypertensive rats treated with captopril(hypertension and captopril group), and in the rats having undergone sham operation (control group). At day 28 after surgery, the aortas were collected from the groups. The expression of PTEN mRNA was detected by RT-PCR. The expression and location of PTEN protein were determined by immunohistochemistry. The results showed that the expression of PTEN in aorta of the hypertension group was significantly lower than that of the hypertension and captopril group, and similarly lower than that of the control group. The intensity of PTEN-positive immunohistochemical production in aorta of the hypertension group was weaker than that of the hypertension and captopril group, and likewise, it was weaker than the control. PTEN-positive immunohistochemical production was located in VSMC of aorta. The findings indicated that the expression of PTEN is reduced in hypertensive aorta, that the reduced PTEN experession can be reversed by captopril treatment, that AngII and the increased mechanical strain may participate in regulating expression of PTEN, and that PTEN may play a role in the arterial remodeling induced by hypertension.

[The role of TLR4-mediated MyD88-dependent pathway in neuroinflammation in hippocampal neurons of rats].

OBJECTIVE: To investigate weather there is a toll-like receptor 4 (TLR4)-mediated myeloid differentiation factor 88 (MyD88)-dependent pathway in hippocampal neurons of rats and the probable role of the pathway in neuroinflammation. METHODS: To establish the proper model, primarily cultured hippocampal neurons were treated with lipopolysaccharides (LPS), or pretreated with TLR4 antibody then co-treated with LPS. The expression of mRNA of MyD88 and TNF-alpha receptor associated factor 6 (TRAF6) were tested by RT-qPCR. The content of MyD88 and TRAF6 were tested by Western blot. The nuclear translocation of nuclear factor-kappaB/P65 (NF-kappaB/p65) was tested by immunofluorescence. The content of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta) and nitric oxide (NO) were tested by ELISA. RESULTS: LPS could increase MyD88 and TRAF6 mRNA, upregulate protein level of MyD88 and TRAF6 and increase the level of TNF-alpha, IL-1beta and NO in cell culture supernatant. LPS also could promote NF-kappa B/p65 translation to the nucleus. The pretreatment with TLR4 antibody reduced the translocation to nucleus for NF-kappaB/P65 and the contents of TNF-alpha, IL-1beta and NO in the culture supernatant. CONCLUSION: There is a TLR4-mediated MyD88-dependent pathway in hippocampal neurons. The activation of this pathway can increase the level of TNF-alpha, IL-1beta and NO in cell culture supernatant. TLR4-mediated MyD88-dependent pathway in hippocampal neurons participate in neuroinflammation, that means neurons are not passive in inflammation.

[The effect of meloxicam on the inflammatory reaction induced by beta amyloid protein in Alzheimer's disease rats].

OBJECTIVE: To investigate the effect and mechanism of meloxicam on the inflammatory reaction induced by beta amyloid protein (AB) in Alzheimer's disease (AD) rats. METHODS: The rat model was established by microinjection of Abeta(1-40) into hippocampus. The expression of NF-kappaB p65 and glial fibrillary acidic protein (GFAP) in hippocampus were detected by immunohistochemistry. The content of GFAP in cortex was tested by Western-blot. The content of TNF-alpha in cortex was tested by ELISA. The expression of IL-1beta mRNA was tested by RT-PCR. RESULTS: The expression of NF-kappaB p65, GFAP and TNF-alpha as well as IL-1beta mRNA were decreased by meloxicam. CONCLUSION: Meloxicam can reduce the proliferation of astrocyte by decreasing the expression of GFAP in AD model rat's hippocampus and cortex. And the depression of NF-kappaB p65 may significantly decrease the expression of TNF-alpha1 and IL-1beta to lessen the inflammatory reaction in cerebral tissue.

[Protective effect of Naoyikang on the Alzheimer's disease model mice induced by D-galactose and NaNO2].

AIM: To investigate the mechanisms of Naoyikang (Traditional Chinese Medicine) on the Alzheimer's Disease (AD) model mice induced by D-galactose (D-gal) and NaNO2. METHODS: The mouse model was established by intraperitoneal injection of D-gal and NaNO2. The capacity of learning and memory was tested on mice with electrical maze; the content of nitric oxide (NO) and the activity of monoamine oxidase-B (MAO-B), glutathione peroxidase (GSH-PX), Na(+) -K(+) -ATP enzyme and Ca(2+) -ATP enzyme in cerebral cortex and hippocampus were assayed by biochemical methods; expression of Bax and Bcl-2 mRNA was detested by RT-PCR. RESULTS: Naoyikang could ameliorate the capacity of learning and memory of AD model mice and reduce MAO-B activity in the brain tissue and activate the activity of Na(+) -K(+) -ATP enzyme and Ca(2+) -ATP enzyme in the brain tissue and decrease the expression of Bax mRNA, but increase the expression of Bcl-2 mRNA in the model brain tissue. CONCLUSION: Naoyikang could protect AD model mice induced by D-gal and NaNO2. It could modify the metabolism of monoamine neurotransmitter in brain through reducing MAO-B activity and protect neurons by activating the activity of Na(+) -K(+) -ATP enzyme and Ca(2+) -ATP enzyme and decrease Bax expression and increase Bcl-2 expression in the model brain tissue.

[Effect of acupoint-injection of oxymatrine on experimental hepatic carcinoma and study on the mechanism].

OBJECTIVE: To investigate the effect of acupoint injection of oxymatrine (OM) on experimental hepatocellular carcinoma and the mechanism. METHODS: The rats of hepatocellular carcinoma induced by 2-acetoaminoflurence (2-AAF) were randomly divided into a normal control group (group N), a model group (group M), a control group of oxymatrine intraperitoneal injection (OM ip group) and a treatment group of small dose oxymatrine injection into Zusanli (OM ZSL group). At the end of 12h week, the serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and gamma-glutamyl transferase (gamma-GT) were determined. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the expressions of cyclin D1 and cyclin-dependent kinase 4 (CDK4) mRNA in hepatocellular carcinoma tissues. RESULTS: The number of cancer nodes on the surface of liver in th Om ip group and the Om ZSL group was lower than in the group M, with the serum ALT, AST, and gamma-GT levels significantly decreased (P<0. 01), and significantly inhibited expressions of cyclin D1, CDK4 mRNA (P<0. 01). CONCLUSION: OM ip and small dose oxymatrine injection into ZSL can treat or delay hepatocarcinogenisis of hepatocellular carcinoma induced by 2-AAF. Partial mechanism of this anti-carcinoma is protecting hepatocytes possibly through improving hepatic functions, and inhibiting excessive proliferation of liver cancer cells via inhibiting the expressions of cyclin Dl, CDK4 mRNA.

[Protective effect of Naoyikang on the damage induced by glutamate in hippocampal neuron].

AIM: To investigate the effect of Naoyikang serum on the damage induced by glutamate in hippocampal neuron. METHODS: Morphological observation, MTT assay and nuclear DNA-associated fluorescence with DAPI dye were applied to evaluate the viability of hippocampal neuron, immunocytochemistry and RT-PCR were used to determine the expression of PTEN. RESULTS: A decreased viability and increased expression of PTEN were shown in hippocampal neuron in response to the treatment with glutamate. It was shown that the percentage of cell death and the expression of PTEN were reduced by the treatment with Naoyikang serum. CONCLUSION: These results suggest that Naoyikang may prevent the toxicity of glutamate by suppressing the expression of PTEN.

[Effect of NOS and PTEN on cardiomyocyte hypertrophy induced by angiotensin II].

AIM: To investigate the effect of NOS and PTEN on the hypertrophic response induced by angiotensin II in the primary culture of neonatal rat cardiomyocytes. METHODS: Total protein content of cardiomyocytes was used as the index of cardiac myocyte hypertrophy. eNOS mRNA, iNOS mRNA and PTEN mRNA expression were assessed using RT-PCR normalized with GAPDH. PTEN protein was determined by Western blot and immunohistochemistry method. RESULTS: (1) On day 1 after Ang II treatment, the expression of eNOS mRNA was significantly decreased whereas iNOS mRNA expression was significantly increased. The effect of Ang II on NOS expression was inhibited by L-arginine. (2) Total protein content of cardiomyocytes increased significantly on day 5 after Ang II treatment, and PTEN protein expression was significantly decreased. The increased protein content and the decreased expression of PTEN protein were inhibited by L-Arg. The L-arginine effect was blocked by L-NAME(NOS inhibitor). (3) The positive immunocytochemical product of PTEN was mainly located in the nucleus of myocardiocyte. CONCLUSION: These results indicate that NOS and PTEN may take part in the process of cardiac myocyte hypertrophy induced by Ang II. The effect of L-arginine on cardiomyocytes may be mediated by NOS/NO and PTEN.

[A study of the effect of PTEN on cardiac hypertrophy].

AIM: To investigate the role of tumor suppressor PTEN in cardiac hypertrophy, the expression of PTEN mRNA in left ventricle of abdominal aorta constricted-induced cardiac hypertrophic rats which treated with and without captopril was analyzed. METHODS: SD rats were divided into control group, hypertrophy group and captopril group. The expression of PTEN mRNA in left ventricle was detected by RT-PCR in different groups in 4 weeks after operation. RESULTS: (1) Compared with control group, the expression of PTEN mRNA in left ventricle of hypertrophy group was reduced. (2) Compared with hypertrophy group, the expression of PTEN mRNA in left ventricle of captopril group was upregulated, which were similar to that of control group. CONCLUSION: PTEN maybe plays a negative regulation role in the process of cardiac hypertrophy, and the role of PTEN is closely relative with renin-angiotensin system.