Huizhen Yu

Resveratrol Protects against TNF-α-Induced Injury in Human Umbilical Endothelial Cells through Promoting Sirtuin-1-Induced Repression of NF-KB and p38 MAPK.

Inflammation and reactive oxygen species (ROS) play important roles in the pathogenesis of atherosclerosis. Resveratrol has been shown to possess anti-inflammatory and antioxidative stress activities, but the underlying mechanisms are not fully understood. In the present study, we investigated the molecular basis associated with the protective effects of resveratrol on tumor necrosis factor-alpha (TNF-α)-induced injury in human umbilical endothelial cells (HUVECs) using a variety of approaches including a cell viability assay, reverse transcription and quantitative polymerase chain reaction, western blot, and immunofluorescence staining. We showed that TNF-α induced CD40 expression and ROS production in cultured HUVECs, which were attenuated by resveratrol treatment. Also, resveratrol increased the expression of sirtuin 1 (SIRT1); and repression of SIRT1 by small-interfering RNA (siRNA) and the SIRT1 inhibitor Ex527 reduced the inhibitory effects of resveratrol on CD40 expression and ROS generation. In addition, resveratrol downregulated the levels of p65 and phospho-p38 MAPK, but this inhibitory effect was attenuated by the suppression of SIRT1 activity. Moreover, the p38 MAPK inhibitor SD203580 and the nuclear factor (NF)-κB inhibitor pyrrolidine dithiocarbamate (PDTC) achieved similar repressive effects as resveratrol on TNF-α-induced ROS generation and CD40 expression. Thus, our study provides a mechanistic link between resveratrol and the activation of SIRT1, the latter of which is involved in resveratrol-mediated repression of the p38 MAPK/NF-κB pathway and ROS production in TNF-α-treated HUVECs.

Synergistic effects of telmisartan and pyridoxamine on early renal damage in spontaneously hypertensive rats.

The aim of this study was to investigate the protective effects of telmisartan and/or pyridoxamine on spontaneously hypertensive rats (SHRs). Rats were treated with telmisartan (T group) or pyridoxamine (P group), or telmisartan and pyridoxamine (TP group). The serum levels of advanced glycation end products (AGEs), superoxide dismutase (SOD), malonaldehyde and the level of 24-h urinary albumin were measured. Morphological changes in renal tissues were observed under light (H&E or Massons trichrome) and transmission electron microscopy. Expression of NF-κBp65 and p-ERK1/2 in renal tissue was detected by immunohistochemistry. Expression of receptors for advanced glycation end products (RAGE) and TGF-β in the renal cortex was investigated by western blotting. We found that early renal structural and functional damage was alleviated in the three intervention groups. SOD activity was significantly elevated in the P and TP groups (P<0.05) compared to that in the T group. Of note, both the positive expression of NF-κBp65 (P<0.01 vs. the T and P groups) and p-ERK1/2 (P<0.05 vs. the P group) was lowest in the TP group. Our results suggest that the combined use of telmisartan and pyridoxamine is superior to the single use of either drug on renoprotection, which may result from the alleviation of oxidative stress and the reduction of NF-κBp65 and p-ERK1/2 activation.

GW27-e1069 Synergistic effect of a tissue kallikrein 1 and tissue inhibitor of matrix metalloproteinase 1 co expression vector on the proliferation of rat vascular smooth muscle cells

Tissue kallikrein 1 (TK1) and tissue inhibitor of matrix metalloproteinase 1 (TIMP1) are important in inhibiting vascular smooth muscle cell (VSMC) proliferation and improving vascular remodeling, respectively. It was hypothesized that a combination of TK1 and TIMP1 genes, mediated by an adenovirus

[PS 01-23] SYNERGISTIC EFFECT OF A TISSUE KALLIKREIN 1 AND TISSUE INHIBITOR OF MATRIX METALLOPROTEINASE 1 CO—EXPRESSION VECTOR ON THE PROLIFERATION OF RAT VASCULAR SMOOTH MUSCLE CELLS

Objective: Tissue kallikrein 1 (TK1) and tissue inhibitor of matrix metalloproteinase 1 (TIMP1) are important in inhibiting vascular smooth muscle cell (VSMC) proliferation and improving vascular remodeling, respectively. It was hypothesized that a combination of TK1 and TIMP1 genes, mediated by an adenovirus vector could augment or act in synergy to enhance the inhibitory effects. Design and Method: The promoter, mCMV carrying hTIMP1 cDNA was subcloned into pDC316—hTK1 to construct a recombinant plasmid carrying hTK1 and hTIMP1 genes. Subsequently, the double gene plasmid and adenovirus backbone plasmid were packaged into HEK293A cells. Gene transcription and protein expression were examined, respectively using reverse transcription—quantitative polymerase chain reaction (PCR) and western blotting assays. VSMC proliferation was assessed using cell counting and methyl—thiazolyl—tetrazoliuin methods. Results: The constructed plasmid containing hTK1 and hTIMP1 genes was correctly identified by means of PCR, double digestion and sequencing analysis. The co—expression vector, Ad—hTK1—hTIMP1 was successfully constructed and packaged into HEK293A cells. When VSMCs were transfected with the co—expression vector, the mRNA transcription and protein expression of hTK1 and hTIMP1 exhibited abundant expression in a concentration—dependent and time—dependent manner, independently. Conclusions: The co—expression vector synergistically inhibited the cell growth and proliferation induced by platelet—derived growth factor—BB compared with the single gene vector.

PS 07-06 RESVERATROL PROTECTS AGAINST TNF-α-INDUCED INJURY IN HUMAN UMBILICAL ENDOTHELIAL CELLS THROUGH PROMOTING SIRTUIN-1-INDUCED REPRESSION OF NF-KB AND P38 MAPK

Objective: Inflammation and reactive oxygen species (ROS) play important roles in the pathogenesis of atherosclerosis. Resveratrol has been shown to possess anti-inflammatory and antioxidative stress activities, but the underlying mechanisms are not fully understood. Design and Method: In the present study, we investigated the molecular basis associated with the protective effects of resveratrol on tumor necrosis factor-alpha (TNF-&agr;)-induced injury in human umbilical endothelial cells (HUVECs) using a variety of approaches including a cell viability assay, reverse transcription and quantitative polymerase chain reaction, western blot, and immunofluorescence staining. Results: We showed that TNF-&agr; induced CD40 expression and ROS production in cultured HUVECs, which were attenuated by resveratrol treatment. Also, resveratrol increased the expression of sirtuin 1 (SIRT1); and repression of SIRT1 by small-interfering RNA (siRNA) and the SIRT1 inhibitor Ex527 reduced the inhibitory effects of resveratrol on CD40 expression and ROS generation. In addition, resveratrol downregulated the levels of p65 and phospho-p38 MAPK, but this inhibitory effect was attenuated by the suppression of SIRT1 activity. Moreover, the p38 MAPK inhibitor SD203580 and the nuclear factor (NF)-&kgr;B inhibitor pyrrolidine dithiocarbamate (PDTC) achieved similar repressive effects as resveratrol on TNF-&agr;-induced ROS generation and CD40 expression. Conclusions: Our study provides a mechanistic link between resveratrol and the activation of SIRT1, the latter of which is involved in resveratrol-mediated repression of the p38 MAPK/NF-&kgr;B pathway and ROS production in TNF-&agr;-treated HUVECs.

Synergistic effect of a tissue kallikrein 1 and tissue inhibitor of matrix metalloproteinase 1 co‑expression vector on the proliferation of rat vascular smooth muscle cells

Tissue kallikrein 1 (TK1) and tissue inhibitor of matrix metalloproteinase 1 (TIMP1) are important in inhibiting vascular smooth muscle cell (VSMC) proliferation and improving vascular remodeling, respectively. It was hypothesized that a combination of TK1 and TIMP1 genes, mediated by an adenovirus vector could augment or act in synergy to enhance the inhibitory effects. The promoter, mCMV carrying hTIMP1 cDNA was subcloned into pDC316-hTK1 to construct a recombinant plasmid carrying hTK1 and hTIMP1 genes. Subsequently, the double gene plasmid and adenovirus backbone plasmid were packaged into HEK293A cells. Gene transcription and protein expression were examined, respectively using reverse transcription-quantitative polymerase chain reaction (PCR) and western blotting assays. VSMC proliferation was assessed using cell counting and methyl-thiazolyl-tetrazoliuin methods. The constructed plasmid containing hTK1 and hTIMP1 genes was correctly identified by means of PCR, double digestion and sequencing analysis. The co-expression vector, Ad-hTK1-hTIMP1 was successfully constructed and packaged into HEK293A cells. When VSMCs were transfected with the co-expression vector, the mRNA transcription and protein expression of hTK1 and hTIMP1 exhibited abundant expression in a concentration-dependent and time-dependent manner, independently. In conclusion, the co-expression vector synergistically inhibited the cell growth and proliferation induced by platelet-derived growth factor-BB compared with the single gene vector.

Effects of pyridoxamine on preliferation of rat vascular smooth muscle cells

Objective To investigate the influence of pyridoxamine on the pre- iferation of rat vascular smooth muscle cells induced by Angiotensin (A)in vitro and its mechanism. Methods Primary VSMCs were cultivated from the thoracic aortas of spontaneously hypertensive rats, and which from the 3th to 5th generation at exponential phase of growth were selected for the experiments and induced by A (10−7 mol/l). The proliferation of VSMCs after intervention with pyridoxamine (0.1, 1 and 10 mmol/l) was determined by MTT. The levels of advanced glycation end-products in cellular supernatant was determined by enzyme-linked immunosorbent assay (ELISA) and the intracellular level of reactive oxygen species (ROS) was detected by f1ow cytometry. The mRNA expressions of RAGE, NF-κB-P65, NADPH oxidaseP47phox were detected by Real-time PCR. Results The proliferation of VSMCs induced by A was inhibited by pyridoxamine in dependent-concentration manner (0.1, 1 and 10 mmol/l). The cellular supernatant AGEs level in pyridoxamine (1 mmol/l, P1) and pyridoxamine (10 mmol/l, P10) group was significantly lower (p<0.001) as compared with A group. It was decreased more in P10 group than that in P1 group (p<0.001); the intracellular ROS level significantly was decreased in P1and P10 group as compared with that in A group (p<0.001), and less in P10 group than that in P1 group (p<0.001); Compared with A group, The mRNA expressions of advanced glycation end-products receptor (RAGE), NF-κB-P65, NADPH oxidaseP47phox and MMP-9 mRNA in P1 and P10 group all were significantly lower (p<0.01), and they were much decreased in P10 group than that in P1 group (p<0.01). Conclusions The pyridoxamine inhibits the proliferation of VSMCs induced by A in dependent-concentration manner, and the potential mechanism of which pyridoxamine may reduce the expressions of RAGE by NF-κB pathway signals and inhibiting AGEs expressing and oxidative stress.

Effects of Telmisartan and Pyridoxamine on vascular smooth muscle cells from rat abdominal aorta vascular

Objective To investigate the effects of telmisartan and pyridoxamine on vascular smooth muscle cells (VSMCs) proliferation and apoptosis with abdominal aorta vascular remodelling in spontaneously hypertensive rat (SHR). Methods SHR randomly received hypertensive, telmisartan (6 mg/kg/d), pyridoxamine (200 mg/kg/d) or combination therapy (telmisartan 6 mg/kg/d, pyridoxamine 200 mg/kg/d). Drug treatment lasted for 16 weeks. Wistar-Kyoto (WKY) as control. The systolic blood pressure (SBP) of rat was measured before and after experimentation every weeks. The serum advanced glycation end-products (AGEs) were detected by competitive ELISA. The serum super oxide dismutase (SOD), nitric oxide (NO) were measured chemical method. The abdominal aorta were assessed by image analysis in HE stained sections. The VSMCs apoptosis and proliferation in abdominal aorta were detected with in situ end labelling technique and proliferating cell nuclear antigen (PCNA) immunohistochemistry staining, respectively. Results The levels of SBP were significantly lower in telmisartan and combination therapy treated SHR than that in hypertensive rats. There was no significant difference between pyridoxamine and WKY (p>0.05). The activity of SOD and NO were significantly higher and AGEs significantly lower in telmisartan, pyridoxamine and combination therapy treated SHR than that in hypertensive rats (p<0.01). The telmisartan, pyridoxamine and combination therapy can significantly inhibit the PCNA expression and significantly enhanced the apoptosis value in abdominal aorta (p<0.01). The effect of combined treatment have been showed significant difference from telmisartan and pyridoxamine (p<0.05). Conclusion Telmisartan and pyridoxamine could partial synergistically improved the blood remodelling associated with attenuation in oxidative stress status and improve the VSMCs unbalance relationship of proliferation and apoptosis in SHR abdominal aorta. Telmisartan was also associated with significant reducing blood pressure.

Effect of serum uric acid level on blood pressure response to antihypertensive drug in male hypertensives

Objective The authors explore the relationship between the serum uric acid levels and blood pressure (BP) response to antihypertensive drugs treatment in male hypertensives by a hospitalisation-based retrospective analysis. Methods According to serum uric acid (SUA) levels, 804 male inpatients with diagnosis of hypertension were divided into two groups, the hyperuricaemia patients (SUA>420 μmol/l, n=305) and patients with normal SUA levels (SUA<420 μmol/l, n=499). Multiple regression analysis models was used to examine the effect of serum uric acid levels on BP response to antihypertensive drugs during hospitalisation, after adjustment for age, diabetes mellitus, chronic kidney disease (CKD) and dyslipidaemia. Result The hypertensive patients with hyperuricaemia have higher body mass index (BMI), dyslipidaemia ratio, systolic BP (diastolic BP), serum creatinine, antihypertensive therapeutic intensity score (p<0.05) and lower estimated glomerular filtration rate (eGFR), BP compliance rate (p<0.05) than the normal group. Serum uric acid levels was positively correlated with BMI, total cholesterol, triglyceride, serum creatinine, systolic BP, diastolic BP (p<0.05), and was negatively correlated with high-density lipoprotein cholesterol and eGFR (p<0.001). In multivariable analyses adjusting for age, the mean difference from systolic BP (diastolic BP) goal, BMI, diabetes, smoking, family history, lipid metabolic disorders, hospitalisation time, antihypertension therapeutic intensity score and CKD, the male patients with hyperuricaemia have less decrease in systolic BP of 5.20 mm Hg (95% CI: 3.11–7.30 mm Hg), and have less decrease in diastolic BP of 1.55 mm Hg (95% CI 0.22 to 2.88 mm Hg). Conclusions The results suggest that serum uric acid levels could affect blood pressure response to the antihypertensive therapy in male hypertensives.