Effect of the angiotensin II receptor blocker valsartan on cardiac hypertrophy and myocardial histone deacetylase expression in rats with aortic constriction.

The aim of the present study was to observe the myocardial expression of members of the histone deacetylase (HDAC) family (HDAC2, HDAC5 and HDAC9) in rats with or without myocardial hypertrophy (MH) in the presence and absence of the angiotensin II receptor blocker valsartan. Adult male Wistar rats were randomly divided into three groups (n=6/group): Sham-operated control rats, treated with distilled water (1 ml/day) through gavage; rats with MH (established through aortic constriction), treated with distilled water (1 ml/day) through gavage; and MH + valsartan rats, treated with 20 mg/kg/day valsartan through gavage. Treatments commenced one day after surgery and continued for eight weeks. Body weight (BW), heart weight (HW) and plasma atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) levels were determined, and the myocardial expression of HDAC2, HDAC5 and HDAC9 was analyzed through a reverse transcription semi-quantitative polymerase chain reaction. The BWs of the rats in the three groups were similar at baseline; however, after eight weeks the BW of the rats in the MH + valsartan group was significantly reduced compared with that of the MH rats. Furthermore, the HW/BW ratio and plasma ANP and BNP levels were increased, the myocardial HDAC2 expression was significantly upregulated and the HDAC5 and HDAC9 expression was significantly downregulated in the MH rats compared with those in the control rats; however, these changes were significantly attenuated by valsartan. Modulation of myocardial HDAC5, HDAC9 and HDAC2 expression may therefore be one of the anti-hypertrophic mechanisms of valsartan in this rat MH model.

Impact of phosphomimetic and non-phosphorylatable mutations of phospholemman on L-type calcium channels gating in HEK 293T cells.

BACKGROUND: Phospholemman (PLM) is an important phosphorylation substrate for protein kinases A and C in the heart. Until now, the association between PLM phosphorylation status and L-type calcium channels (LTCCs) gating has not been fully understood. We investigated the kinetics of LTCCs in HEK 293T cells expressing phosphomimetic or nonphosphorylatable PLM mutants. METHODS: The LTCCs gating was measured in HEK 293T cells transfected with LTCC and wild-type (WT) PLM, phosphomimetic or nonphosphorylatable PLM mutants: 6263AA, 6869AA, AAAA, 6263DD, 6869DD or DDDD. RESULTS: WT PLM significantly slowed LTCCs activation and deactivation while enhanced voltage-dependent inactivation (VDI). PLM mutants 6869DD and DDDD significantly increased the peak of the currents. 6263DD accelerated channel activation, while 6263AA slowed it more than WT PLM. 6869DD significantly enhanced PLM-induced increase of VDI. AAAA slowed the channel activation more than 6263AA, and DDDD accelerated the channel VDI more than 6869DD. CONCLUSIONS: Our results demonstrate that phosphomimetic PLM could stimulate LTCCs and alter their dynamics, while PLM nonphosphorylatable mutant produced the opposite effects.

Effects of heme oxygenase-1 gene modulated mesenchymal stem cells on vasculogenesis in ischemic swine hearts.

BACKGROUND: Mesenchymal stem cells (MSCs) transplantation may partially restore heart function in the treatment of acute myocardial infarction (AMI). The aim of this study was to explore the beneficial effects of MSCs modified with heme xygenase-1 (HO-1) on post-infarct swine hearts to determine whether the induction of therapeutic angiogenesis is modified by the angiogenic cytokines released from the implanted cells. METHODS: In vitro, MSCs were divided into four groups: (1) non-transfected MSCs (MSCs group), (2) MSCs transfected with the pcDNA3.1-Lacz plasmid (Lacz-MSCs group), (3) MSCs transfected with pcDNA3.1-hHO-1 (HO-1-MSCs group), and (4) MSCs transfected with pcDNA3.1-hHO-1 and pretreatment with an HO inhibitor, tin protoporphyrin (SnPP) (HO-1-MSCs + SnPP group). Cells were cultured in an airtight incubation bottle for 24 hours, in which the oxygen concentration was maintained at < 1%, followed by 12 hours of reoxygenation. After hypoxia/reoxygen treatment, ELISA was used to measure transforming growth factor (TGF-ß) and fibroblast growth factor (FGF-2) in the supernatant. In vivo, 28 Chinese mini-pigs were randomly allocated to the following treatment groups: (1) control group (saline), (2) Lacz-MSCs group, (3) HO-1-MSCs group, and (4) HO-1-MSCs + SnPP group. About 1 × 10(7) of autologous stem cells or an identical volume of saline was injected intracoronary into porcine hearts 1 hour after MI. Magnetic resonance imaging (MRI) assay and postmortem analysis were assessed four weeks after stem cell transplantation. RESULTS: Post hypoxia/reoxygenation in vitro, TGF-ß in the supernatant was significantly increased in the HO-1-MSCs ((874.88 ± 68.23) pg/ml) compared with Lacz-MSCs ((687.81 ± 57.64) pg/ml, P < 0.001). FGF-2 was also significantly increased in the HO-1-MSCs ((1106.48 ± 107.06) pg/ml) compared with the Lacz-MSCs ((853.85 ± 74.44) pg/ml, P < 0.001). In vivo, at four weeks after transplantation, HO-1 gene transfer increased the capillary density in the peri-infarct area compared with the Lacz-MSCs group (14.24 ± 1.66/HPFs vs. 11.51 ± 1.34/HPFs, P < 0.001). Arteriolar density was also significantly higher in HO-1-MSCs group than in the Lacz-MSCs group (7.86 ± 2.00/HPFs vs. 6.45 ± 1.74/HPFs, P = 0.001). At the same time, the cardiac function was significantly improved in the HO-1-MSCs group compared with the Lacz-MSCs group ((53.17 ± 3.55)% vs. (48.82 ± 2.98)%, P < 0.05). However, all these effects were significantly abrogated by SnPP. CONCLUSION: MSCs provided a beneficial effect on cardiac function after ischemia/reperfusion by the induction of therapeutic angiogenesis, and this effect was amplified by HO-1 overexpression.

Suppression of experimental abdominal aortic aneurysm in a rat model by the phosphodiesterase 3 inhibitor cilostazol.

BACKGROUND: The present experiments sought to determine whether cilostazol, a selective inhibitor of cyclic adenosine monophosphate (cAMP) phosphodiesterase 3 (PDE3), suppressed elastase-induced abdominal aortic aneurysm (AAA) development in a rat model. METHODS: Male Sprague-Dawley rats (n = 16/each group) were randomly distributed into three groups: sham-, saline-, and cilostazol-. Rats of saline and cilostazol groups underwent intra-aortic elastase perfusion to induce AAAs, while rats of sham-group were perfused with saline. Rats of cilostazol-group received cilostazol treatment (100 mgkg(-1)d(-1)) for the entire experimental period. The areas of the lumen of the aortas at the segment with maximum diameter were measured preperfusion and on d 7, 14 after perfusion. Systolic blood pressure was measured by tail-cuff technique. Aortic tissue samples were harvested on d 14 after intra-aortic perfusion and evaluated by reverse transcription-polymerase chain reaction and Western blot for matrix metalloproteinase-2, 9 (MMP-2, 9), by immunohistochemistry for nuclear factor kappa B (NF-κB), and by Gomori aldehyde fuchsin for elastin. Activity of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and level of reactive oxygen species (ROS) in these samples were also measured. RESULTS: On d 14, rats of saline-group had significantly increased aortic sizes compared with sham-group (P < 0.01), while, cilostazol treatment significantly reduced this increase (cilostazol- versus saline-, P < 0.01) without affecting blood pressure (P > 0.05). The expression of both MMP-2 and MMP-9 and the destruction of elastic fibers in aortic tissues were significantly decreased by cilostazol treatment (P < 0.05), probably through the suppression of NF-κB activation (P < 0.01). Consistently, cilostazol significantly inhibited NADPH oxidase activity (P < 0.01), accompanied by a reduced level of ROS (P < 0.01). CONCLUSION: Cilostazol retards experimental AAAs development independently of blood pressure reduction possibly by inhibiting proteolysis, inflammation, and oxidative stress. Selective PDE3 inhibition may offer an additional method to pharmacologically inhibit AAAs.

[Effects of autologous mesenchymal stem cells transfected with heme oxygenase-1 gene transplantation on ischemic Swine hearts].

OBJECTIVE: To observe the effect of intracoronary transfer of autologous HO-1 overexpressed MSCs in porcine model of myocardial ischemia (1 h)/reperfusion. METHODS: Apoptosis was assayed and cytokine concentrations in supernatant were measured in cells exposed to hypoxia-reoxygen in vitro. In vivo, Chinese male mini-pigs were allocated to the following treatment groups: control group (saline), MSCs group (MSCs), MSCs transfected with pcDNA3.1-nHO-1 (HO-1-MSCs). 1 x 10(7) of autologous stem cells or identical volume of saline was injected intracoronary into porcine hearts 1 h after ischemia. MRI assay and postmortem analysis were assessed 3 months after stem cell transplantation. RESULTS: In vitro, cell apoptosis rate post hypoxia-reoxygen was significantly reduced in HO-1-MSCs group (30.30% +/- 7.64%) compared with that in MSCs group (56.93% +/- 4.68%, P < 0.001) and LacZ-MSCs group (55.88% +/- 4.38%, P < 0.001), VEGF was also significantly upregulated in HO-1-MSCs group [(768.44 +/- 78.38) pg/ml] compared with that in MSCs group [(555.27 +/- 67.67) pg/ml, P < 0.001] and LacZ-MSCs group [(522.97 +/- 71.45) pg/ml, P < 0.001]. In vivo, cardiac function was significantly improved in both MSCs transplantation groups compared to saline group (all P < 0.05 vs.saline) and the left ventricular ejection fraction was significantly higher in HO-1-MSCs group compared with that in MSCs group at 3 months after transplantation (53.50% +/- 2.09% vs. 49.54% +/- 2.74%, P = 0.017), capillary density in the peri-infarct area was also significantly higher in HO-1-MSC group than that in MSCs group [(14.59 +/- 2.39)/HPF vs. (11.78 +/- 2.48)/HPF, P = 0.033]. CONCLUSIONS: Efficacy of HO-1 overexpressed MSCs on improving cardiac function and promoting angiogenesis was greater than those by MSCs in this porcine ischemia/reperfusion model.