Effects of metformin on nuclear factor-κB expression and serum high sensitivity C-reactive protein level in atherosclerosis model of rabbit

AIM: To investigate the effects of metformin on nuclear factor-κB (NF-κB),its inhibitor IκB,and the level of serum high sensitivity C-reactive protein (hs-CRP) in rabbits.METHODS: 24 New Zealand male rabbits were randomly divided into control group,atherosclerosis (AS) group and metformin (Met) group.AS group and Met group were made as models by cholesterolenriched diets feeding and vascular intimal immunologic injury.The AS model was confirmed by high frequency ultrasound.Met group were given metformin 150 mg·kg-1·d-1 for 8 weeks.At the end of experiment,serum hs-CRP and serum lipids in all three groups were detected.Immunohistochemistry and Western blotting technique were applied to detect the expression of nucleus NF-κB p65 and cytoplasma IκBα in aorta in all three groups.RESULTS: Compared to normal control group,the level of serum hs-CRP was elevated (1.27±0.43 vs 3.96±0.63,P<0.01),the expression of nucleus NF-κB p65 increased significantly (P<0.01) while the expression of IκBα reduced significantly (P<0.01).Compared to AS group,metformin significantly reduced the level of serum hs-CRP (2.79±0.40 vs 3.96±0.63,P<0.05) and the expression of nucleus NF-κB p65 (P<0.01),and increased the expression of IκBα (P<0.05).CONCLUSION: Metformin inhibits the activation of NF-κB p65 and the degradation of IκBα,and decreases the levels of serum hs-CRP in AS rabbits.These results suggest that metformin exerts direct vascular anti-inflammatory effects.It may be one important mechanism of metformins antiatherogenic properties.     

Effects of CARHSP1 gene expression on viability,apoptosis and immune factors of vascular endothe-lial cells induced by hypoxia in atherosclerosis

AIM: To investigate the effect of calcium-regulated heat stable protein 1 (CARHSP1) gene expression on the viability, apoptosis and expression of interleukin-6 (IL-6) and C-reactive protein (CRP) in vascular endothe-lial cells induced by hypoxia.METHODS: The protein expression of CARHSP1 was detected by Western blot in atherosclerotic plaques. Human umbilical vein endothelial cells (HUVECs) were treated with hypoxia, and the cells were divided into normal culture group, hypoxia group, hypoxia+CARHSP1-siRNA group and hypoxia+pcDNA3.1-CARHSP1 group. The viability and apoptotic rate of the HUVECs were measured by CCK-8 assay and flow cytometry, respectively. The mRNA expression of IL-6 and CRP was detected by RT-PCR. The protein levels of caspase-3, cleaved caspase-3, Bcl-2 and Bax were determined by Western blot.RESULTS: The protein expression of CARHSP1 in atherosclerotic plaques was significantly higher than that in control group (P<0.05). Hypoxia significantly increased the expression of CARHSP1. The cell viability and the protein expression of Bcl-2 were significantly lower in hypoxia group than those in normal culture group (P<0.05). The apoptotic rate and the protein levels of IL-6, CRP, cleaved caspase-3 and Bax were significantly higher than those in normal culture group (P<0.05). Compared with hypoxia group, the cell viability and protein expression of Bcl-2 were significantly increased in hypoxia+CARHSP1-siRNA group, while the apoptotic rate and the protein levels of IL-6, CRP, cleaved caspase-3 and Bax were decreased significantly (P<0.05). The cell viability and protein expression of Bcl-2 were decreased significantly in hypoxia+pcDNA3.1-CARHSP1 group, while the apoptotic rate and the protein le-vels of IL-6, CRP, cleaved caspase-3 and Bax were increased significantly (P<0.05).CONCLUSION: The expression of CARHSP1 is increased in atherosclerotic plaques, and inhibition of CARHSP1 expression improves the viability, reduces the apoptosis, and down-regulates the expression of IL-6 and CRP in the HUVECs. Over-expression of CARHSP1 exerts the opposite effect.     

Inhibitory effect of high-dose Xuezhikang on inflammatory response induced by percutaneous coronary intervention in patients with unstable angina

AIM: To study the inhibitory effect of high-dose Xuezhikang,administered before percutaneous coronary intervention (PCI) on inflammatory response induced by PCI in patients with unstable angina (UA).METHODS: All patients with UA in class Ⅲ and ⅡB according to Braunwald classification were considered for inclusion in the present study.Finally,196 patients received Xuezhikang treatment 72 h before coronary angiography and successfully performed PCI with elevated C-reactive protein (CRP) level (>3 mg/L) were randomised to 2 groups: 1.2 g/d of Xuezhikang as group A,or 2.4 g/d of Xuezhikang as group B.The levels of CRP were measured at baseline,after 3 days of therapy (before procedure) and 48 hours after PCI.The patients were followed-up for 6 months for major adverse coronary events and left ventricular ejection fraction.RESULTS: There was no significant difference in the mean CRP level among the two randomized groups (P>0.05),however,after three days of pharmacological treatment,there was significantly reduced CRP content in group A ［(5.44±1.57） mg/L vs （4.04±1.54） mg/L,P<0.05］ and in group B ［(5.42±1.36） mg/L vs （3.60±1.14） mg/L,P<0.05］ compared with admission.Measurements performed 48 hours after the procedure revealed a marked CRP level increase in group A (up to 9.22 mg/L±5.03 mg/L) and an obvious increase in groups B (up to 4.97 mg/L±1.75 mg/L,P<0.05) compared with pre-procedure.The serum level of CRP in B group was distinctly lower than that in A group before (P<0.05) and after the procedure (P<0.05),respectively.Major adverse coronary events during the 6-month clinical follow-up occurred less in group A than that in group B ［21/104 (20.2%) vs 9/92 (9.8%); patients,P<0.05］.Follow-up echocardiography revealed lower left ventricular ejection fraction in group A than that in group B (55.41%±10.93% vs 59.30%±9.99%,P<0.05).CONCLUSION: High-dose Xuezhikang therapy,administered before PCI,has better inhibition effect than low-dose on inflammatory response induced by PCI in patients with UA.Attenuation of inflammatory response may be crucial for the reduction of coronary events following invasive coronary interventions.     

Effects of tirofiban on no-reflow and activation of NF-κB after myocardial ischemia/reperfusion in rats

AIM: To investigate the effects of platelet glycoprotein Ⅱb/Ⅲa receptor inhibitor tirofiban on myocardial no-reflow and activation of NF-κB after acute ischemia/reperfusion in rats. METHODS: Male Wistar rats were randomized into sham operation group, control group and tirofiban treatment group. Control group and tirofiban group were subjected ischemia for 90 min by ligation of coronary artery after thoracotomy and subsequently reperfusion for 120 min to establish acute myocardial ischemia/reperfusion no-reflow models. Thioflavine S, Evans blue and triphenyltetra zolium chloride (TTC) staining were performed to evaluate the area of no-reflow (ANR), infracted area (IA) and risk area (RA) of the heart. Immunohistochemistry was used for semi-quantitative analysis of the expression of nuclear factor-κB p65 (NF-κB p65) protein in myocytes and arteriole. Activity of myeloperoxidase (MPO) and content of malondialdehyde (MDA) in risk area of the heart were detected by ultraviolet spectrophotometer. RESULTS: After 120 min for reperfusion, compared to sham group, the statistical differences of higher positive expression of NF-κB p65 in myocytes and arteriole, activity of MPO and content of MDA both in control and tirofiban group were observed. Compared to control group, lower positive expression of NF-κB p65 in myocyte and arteriole, activity of MPO and content of MDA in tirofiban group were found (P<0.05, P<0.01). A markedly reduced ANR and IA were observed in tirofiban group than those in control group (34.36%±6.04% vs 52.09%±6.89%, P<0.01; 80.41%±8.48% vs 90.13%±5.72%, P<0.05). CONCLUSION: After myocardial ischemia/reperfusion for 120 min, no-reflow phenomenon can be observed in rats. Tirofiban reduces the areas of anatomic no-reflow and infarction, inhibits the activation of NF-κB in myocyte and arteriole, and decreases the infiltration of neutrophils and release of oxygen free radicals.     

Sodium valproate induces cycle arrest,apoptosis and elevates p21WAF1 mRNA expression in chronic myeloid leukemia cell line K562

AIM:To investigate the effects of sodium valproate (VPA) on the cell cycle and apoptosis of chronic myeloid leukemia cell line K562, and to explore the possible mechanisms. METHODS:K562 cells were treated with VPA. Cell cycle and apoptosis were analyzed by flow cytometry. The expression of p21WAF1 mRNA was detected by RT-PCR. RESULTS:After treatment with VPA, cell cycle was arrested obviously at G0/G1 phase ［(82.30±9.41)% vs (40.13±2.12)%, P<0.05］. The apoptotic rate was significantly higher in the cells treated with VPA than that in untreated cells ［(11.47%±0.25%) vs (4.77%±0.40%), P<0.05］. The level of p21WAF1 mRNA was increased ［(1.65±0.91) vs (0.25±0.04), P<0.05］. CONCLUSION:VPA induces elevated expression of p21WAF1 mRNA in K562 cells, resulting in G0/G1 phase arrest and apoptosis in vitro.     

Ischemic preconditioning relieves the ischemia/reperfusion injury of neurons in hippocampus by inhibiting the expression of p53

AIM: To examine whether ischemic preconditioning (IPC) can protect against apoptosis in CA1 subfield of hippocampus following reperfusion of a lethal ischemia in rats and explore the role of IPC by inhibiting the expression of p53 in this process. METHODS: Wistar rats were used in the experiment. A global ischemia/reperfusion model was induced by 4-vessel occlusion. The rats were divided into the following three groups randomly: (1) ischemic preconditioning group (IPC group); (2) ischemia/reperfusion group (IR group); (3) control group. The histopathological changes, the percentage of apoptosis and the expression of p53 gene in CA1 region of rat hippocampus were examined by HE staining, FCM, RT-PCR and immunohistochemistry techniques. RESULTS: The neuronal density of CA1 region in IPC group ［(217±9)/0.72 mm²］ was significantly higher than that in IR group ［(29±5)/0.72 mm², P<0.01］. The percentage of apoptotic neurons in IPC group (2.07%±0.21%) was lower than that in IR group (4.26%±0.08%), P<0.01. Compared with IR group, the expression of p53 gene in IPC group was significantly weakened. CONCLUSION: Ischemic preconditioning protects the ischemic neurons in CA1 region of rat hippocampus by inhibiting the expression of p53 gene.     

Physiological and pathological effects of body position changes on experimental acute lung injury in rabbits

AIM: To study the effects of body position on the physiological and pathological changes in an oleic acid-induced acute lung injury (ALI) model. METHODS: One control group and three experimental groups were set up in which the rabbits were put on supine, prone and rotation, respectively. The changes of blood pressure, heart rate and partial pressure of arterial oxygen, lung mechanics, cytokines in the course of experiment and pathologic changes were observed. RESULTS: At the end of experiment, heart rate in rotated rabbits was lower than that in prone rabbits ［(176.13±26.55) beats/min vs (217.75±14.44) beats/min, P<0.05］. Partial pressure of arterial oxygen and lung compliance in prone and rotated rabbits were higher than that in supine rabbits ［(157.75±51.19) mmHg and (166.08±37.07) mmHg vs (86.59±23.82) mmHg, (2.75±0.89) mL/cmH2O and (2.63±0.74) mL/cmH2O vs (1.63±0.52) mL/cmH2O, respectively, P<0.05］, but intrapulmonary shunt was just up side down. Airway peak inspiration pressure in rotated rabbits was higher than that in prone group rabbits, ［(19.63±2.45) cmH2O vs (16.00±2.27) cmH2O, P<0.05］. TNF-α in supine and rotated rabbits was higher than that in control rabbits, ［(3.12±0.83 and 2.59±0.79) μg/L vs (1.36±0.34) μg/L］, but it was not the case in prone rabbits. Partial pressure of arterial oxygen was positive correlation to lung compliance and negative correlation to intrapulmonary shunt. Dorsal lung edema was more severe than ventral in group 2, ventral lung edema was more severe than dorsal in group 3 and lung edema was diffused in group 4. CONCLUSIONS: Prone and rotation positions increased lung compliance and decreased intrapulmonary shunt accompanied with oxygenation improvement in ALI model, but prone was safer than rotation position, the former can inhibit the secretion of TNF-α. The distribution of lung edema changed with body position changes.     

Influences of wild-type p53 gene overexpression on the differentiation,apoptosis and expression of scavenger receptor CD36 in U937 cells*

AIM: To study the effect of wild-type p53 gene on the differentiation, apoptosis and expression of scavenger receptor CD36 in U937 cells. METHODS： Recombinant adenovirus vector with wild-type p53 gene was constructed and used to transfect U937 cells. With the expression of wild-type p53 gene following adenoviral infection, transfected U937 cells were largely promoted to differentiate into macrophages. RESUITS： Trypanblue-staining test demonstrated that the percentage of positive cells increased from (14.2±5.5)% to (64.6±9.2)% and nitroblue tetrazolium (NBT) reduction test reached similar results (6.3±1.8)% vs (49.7±12.6)%. Furthermore, CD36 mRNA was up-regulated as confirmed by RT-PCR. The increased expression level of CD 36 was also detected by flow cytometry analysis. CONCLUSION： These results suggest that wild-type p53 gene can affect U937 cells differentiation and apoptosis, up-regulate expression of scavenger receptor CD36. It may have a potential significance on atherogenesis.     

Expression of galectin-3 in human coronary artery and its correlation with plaque structural stability

AIM To investigate the relationship between the expression level of galectin-3 and the stability of plaque structure in human atherosclerotic plaques. METHODS The coronary specimens from autopsy cases （n=84） were collected. Among them, 22 cases had coronary atherosclerotic lesions without sudden death of coronary heart disease （A1 group）, 20 cases were sudden death of coronary heart disease without secondary lesions （A2 group）, 24 cases were sudden death of coronary heart disease with secondary lesions （A3 group）, and 18 cases without heart disease were used as normal control group （control group）. The intimal thickness, necrotic lesion thickness, fibrous cap thickness and the degree of lumen stenosis were measured by routine HE staining in all coronary arteries. The foam cells in the lesion were marked by CD68 and counted. The expression of galectin-3, CD68 and matrix metalloproteinase-2 （MMP-2） in coronary artery intima was detected by immunohistochemical staining, Western blot and RT-qPCR. The correlation between above factors and the structural stability of atherosclerotic plaques was also analyzed. RESULTS Compared with control group, the intima and necrotic lesions were thickened, the fiber cap was thinned, and the degree of lumen stenosis were increased in A1~3 groups （P<0.05）. The number of foam cells in the atherosclerotic focus was increased （P<0.05）. The protein and mRNA levels of galectin-3, CD68 and MMP-2 in the lesions showed an increasing trend from normal group to A1~3 groups （P<0.05）. The expression of galectin-3, CD68 and MMP-2 in atherosclerotic lesions was positively correlated with intimal thickness and necrotic lesion thickness, and negatively correlated with fibrous cap thickness. CONCLUSION The expression of galectin-3 in human coronary atherosclerotic lesions is increased, which is related to the stability of atherosclerotic plaques.     

Correlation between intracellular magnesium and expression of beta 2-adrenergic receptor mRNA in the lung of C57BL/6 asthmatic mice

AIM: To explore the effect of intracellular magnesium on expression of beta 2-adrenergic receptor mRNA in the lung of C57BL/6 asthmatic mice. METHODS: Ninety-six healthy, 4-6 weeks old and female C57BL/6 mice, weighting (12±2) g, were randomly divided into the following A, B, C, D groups with 24 mice in each group. The mice were sensitized and challenged by ovalbumin to establish the asthmatic model. A and B groups were fed with magnesium deficient diet. C and D groups were fed with normal magnesium level diet. B and D groups were injected intraperitoneally with 0.2 mL sulphate salbutamol solution after OVA provocation. A and C groups were injected intraperitoneally with 0.2 mL saline as control. Eight mice in each group were randomly taken out at 1 d, 21 d, 34 d to detect plasma Mg2+, intracellular Mg2+, the beta 2-AR mRNA and protein in lung tissue. RESULTS: No significant difference in plasma Mg2+, intracellular Mg2+, the beta 2-AR mRNA and protein in lung tissue among all groups at 1st d was observed (P>0.05, respectively). Plasma Mg2+, intracellular Mg2+, the beta 2-AR mRNA and protein in lung tissue in group C at 21st d and 34th d were significantly higher than those in group A at 21st d and 34th d ［21st d:(0.84±0.09)mmol/L vs 0.57±0.10)mmol/L, (2.39±0.14)mmol/L vs (2.11±0.08) mmol/L,(0.75±0.09)pmol/g vs (0.59±0.06)pmol/g, (88.50±8.50)pmol/g vs (60.10±7.70)pmol/g, P<0.05, respectively; 34th d:(0.67±0.10)mmol/L vs (0.51±0.09)mmol/L, (2.17±0.08)mmol/L vs (2.05±0.09)mmol/L,(0.61±0.05)pmol/g vs (0.53±0.06)pmol/g, (76.60±7.10)pmol/g vs (58.00±7.60)pmol/g, P<0.05, respectively］. Also, plasma Mg2+, intracellular Mg2+, the beta 2-AR, mRNA and protein of lung tissue in group D at 21st d and 34th d were significantly higher than those in group B at 21st d and 34th d ［21st d:(0.95±0.33)mmol/L vs (0.46±0.09)mmol/L,(2.32±0.18)mmol/L vs (1.87±0.14)mmol/L,(0.73±0.10)pmol/g vs (0.43±0.07)pmol/g, (96.90±8.00)pmol/g vs (47.90±4.90)pmol/g, P<0.05, respectively; 34th d:(0.71±0.10)mmol/L vs (0.31±0.08)mmol/L, (1.66±0.13)mmol/L vs (1.45±0.16)mmol/L,(0.40±0.07)pmol/g vs (0.33±0.05)pmol/g, (61.50±3.20)pmol/g vs (35.30±7.10)pmol/g, P<0.05, respectively］.CONCLUSION: The expression of β2-adrenergic receptor mRNA in the lung of C57BL/6 asthmatic mice with deficient intracellular magnesium is suppressed and C57BL/6 asthmatic mice with deficient intracellular magnesium are even easier to induce downregulation of β2-adrenergic receptor when β2-AR agonist is administered.     

Triptolide induces apoptosis of rat synoviocytes in collagen-induced arthritis

AIM: To determine whether triptolide induce apoptosis of synovial cells in collagen-induced arthritis (CIA) in rats. METHODS: The male Wistar rats were used to make CIA models by immunized with Bovine collagen Ⅱ (BCⅡ) in Freund's complete adjuvant (FCA). A total of 20 CIA rats were randomly divided into 2 groups, triptolide group (10 rats) and CIA control group (10 rats). Triptolide group were administered with triptolide at 40 μg/kg body weight intramuscularly every three days. CIA control group and another 10 age-matched normal rats were given normal saline instead. The rats were sacrificed on the 31st day after the triptolide administration. The pieces of synovium of the rat knee joints were harvested. The synovium was examined by HE staining and electron microscope. The apoptosis was tested by TUNEL and flow cytometer. RESULTS: The earlier phase of apoptotic synoviocytes were observed under the electron microscope. The flow cytometry showed that the percentage of the apoptotic cells was (3.98±1.16)% in the triptolide group, (1.83±0.82)% in the CIA control group, and (0.87±0.24)% in the normal group (P<0.01: triptolide vs control group). While the percentage of the cells in DNA synthesis phase was (3.3±1.2)% in the triptolide group, (8.0±1.4)% in the CIA control group, and (3.4±0.7)% in the normal group. There is significantly different in the apoptosis changes between the triptolide group and the CIA control group (P<0.01: triptolide vs CIA control group). The TUNEL labeling demonstrated that the percentage of the apoptotic cells was (4.5±1.0)% in the triptolide group, (2.2±1.0)% in the CIA control group, and (1.0±0.4)% in the normal group. The difference of apoptotic rate between the triptolide group and the CIA control group is significant (P<0.01). CONCLUSION: This study demonstrates that triptolide can induce apoptosis in CIA rats, which may be one of the mechanisms that triptolide treats the rheumatoid arthritis.     

Cell transplantation combined with transmuscle laser revascularization augments neovascularization in rat ischemia hindlimb

AIM: To determine the combined effect of transmuscle laser revascularization (TMR) and endothelial progenitor cells(EPCs) treatment on ischemic hindlimb of nude rats.METHODS: Mononuclear cells (MNCs) isolated from human umbilical cord-blood (HUCB) by density gradient centrifugation were expanded in vitro. Immunocytochemistry and flow cytometry studies were performed. EPCs were labeled with 1, 1- dioctadecyl-1 to 3, 3, 3, 3- tetramethyl-indocarbocyanine perchlorate (DiI) before injected into the laser induced channels or ischemic region. Acute ischemic limb was created in 4 groups of nude rats by ligating right external iliac artery. All animals were divided randomly into the following four groups: TMR+EPCs group: local transplantation of EPCs into laser channels; TMR group: transmuscular channels were created without EPCs; EPCs group: EPCs were injected into ischemic hindlimb; control group: ischemic model without TMR or EPCs. All rats underwent femoral artery ultrasonic blood flow measurements of the ischemic and nonischemic limbs to obtained a flow ratio ［femoral artery flow index (FAFI): right femoral artery flow /left femoral artery flow］ at baseline (after ligating artery immediately) and 28 days postoperation, and then the samples of ischemic limb muscle underwent histochemical and immunohistologic analysis. RESULTS: The attached cells expressed endothelial cell (ECs) markers (KDR, CD34, CD31, AC133 and von Willebrand factor) and exhibited function similar to that of ECs judged by Ac-LDL incorporation. Flow cytometric analysis disclosed that AT cells were positive for CD34 (62%±7%) and AC133 (57.2%±9.8%) at day 7 of culture. 28 days after therapy, FAFI was significantly higher in the TMR +EPCs (0.66±0.09, P<0.01) and EPCs group (0.59±0.09, P<0.05) compared to control group (0.47±0.05). It was significantly higher in TMR +EPCs-，EPCs- and TMR group compared to baseline (TMR+EPCs group: 0.66±0.09 vs 0.39±0.07, P<0.01; TMR group: 0.54±0.12 vs 0.40±0.09, P<0.05; EPCs group: 0.59±0.09 vs 0.38±0.08, P<0.01; control group: 0.47 ±0.05 vs 0.39±0.08, P>0.05). FAFI in the control group was unchanged and no difference was found between TMR group and control group. TMR+EPCs (5.66±0.77), TMR (4.96±0.31) as well as EPCs (4.68±0.44) treatment resulted in an increased number of capillaries in the treated regional area compared with control group (2.60±0.31, P<0.01).CONCLUSION: Nd: YAG-laser revascularization combined with the application of EPCs transplantation significantly ameliorates perfusion and augments neovascularization in this ischemic hindlimb model.     

Correlation between CD4+CD28-T cells and lymphocytic apoptosis in rheumatoid arthritis patients

AIM: To investigate the fraction of CD4⁺CD28^-T cells and its correlation with lymphocytic apoptosis in peripheral blood of rheumatoid arthritis (RA) patients.METHODS: The RA patients and age-matched health controls were selected in the study. The lymphocytes were isolated from peripheral blood. CD4⁺ T cells without CD28 expression (CD4⁺ CD28^-) were analyzed by flow cytometry. The incidence of apoptosis in the cells cultured with or without PHA for 24 h was determined by flow cytometry with Annexin V-FITC/PI di-staining. The correlation between the fraction of CD4⁺CD28^-T cells and lymphocytic apoptosis was also observed.RESULTS: The fraction of CD4⁺CD28^-T cells was significantly higher in RA group than that in healthy control group (7.79%±3.52% vs 1.89%±1.78%, P<0.05). The apoptotic level in PHA cultured lymphocytes was significantly lower in RA group than that in healthy controls (11.38%±5.73% vs 19.46%±6.32％, P<0.05). Spearman analysis showed that there was a negative correlation between the fraction of CD4⁺CD28^-T cells and apoptotic level of activated lymphocytes (r=-0.433,P<0.01).CONCLUSION: The increased CD4⁺CD28^-T cells contribute to prolong the lifespan of activated lymphocytes in peripheral blood of RA patients, and the persistence of activated lymphocytes may contribute to the pathogenesis of RA.     

Effects of pentoxifylline on ventricular remodeling and cardiac function of dilated cardiomyopathy rats

AIM: To explore the effects of pentoxifylline (PTX) on ventricular remodeling and cardiac function in dilated cardiomyopathy (DCM) rats.METHODS: Lewis rats were randomly allocated to a myocin-induced dilated cardiomyopathy (DCM) group receiving saline (n=10), a DCM group receiving PTX (PTX group; 25 mg·kg^-1·d^-1, ip, for 30 days, n=10) or healthy control group (n=10). The levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-10 in the blood plasma were analyzed by ELISA. The extent of fibrosis was estimated using Massons staining and immunohistochemistry analyses. Cardiac structure and function were measured by echocardiography.RESULTS: PTX decreased plasma levels of TNF-α and IL-6, and increased IL-10 level in DCM animals compared with DCM group ［TNF-α: (7.21±0.24) μg/L vs (19.30±1.31) μg/L, P<0.01; IL-6: (119.60±36.58) ng/L vs (189.50±13.25) ng/L, P<0.05; IL-10: (41.26±3.27) μg/L vs (32.45±4.32) μg/L, P<0.05］. Collagen volume fraction (CVF), perivascular collagen area (PVCA) and collagen Ⅰ/Ⅲ ratio were lower in PTX group than those in DCM group ［CVF: (16.45±3.01)% vs (23.33±4.43)%, P<0.05; PVCA: 4.58±2.10 vs 13.74±4.29, P<0.05; Ⅰ/Ⅲ ratio: 2.84±0.67 vs 4.22±0.54, P<0.01］. Left ventricular end-diastolic dimension reduced ［(6.11±0.51) mm vs (6.46±0.28) mm, P<0.05］ and left ventricular ejection fraction elevated ［(77.29±5.20)% vs (62.73±10.11)%, P<0.01］ by PTX compared with DCM.CONCLUSION: PTX modulates plasma levels of inflammatory cytokines, delays the ventricle remodeling and improves the heart function in DCM rats.     

Role of Kv1.3 channel of CD4+T lymphocytes in the formation of atherosclerosis in rat spleen

AIM: To investigate the function of voltage-gated potassium channel Kv1.3 and its possible role in CD4⁺ T lymphocytes in the formation of atherosclerosis (AS) in rat spleen. METHODS: The rat atherosclerosis model was established by feeding high-fat diet. The proportion of lymphocytes was determined by flow cytometry. The CD4⁺ T lymphocytes were separated using immunomagnetic bead. The mRNA expression of Kv1.3 in CD4⁺ T lymphocytes was detected. The concentrations of intracellular calcium and cytokines were also measured. RESULTS: (1) The proportion of CD4⁺ T lymphocytes in AS group was significantly higher than that in control group (74.93%±2.15% vs 67.80%±2.54%, P<0.05). (2) After stimulated with concanavalin A (ConA), the proliferation of CD4⁺ T lymphocytes in AS group was significantly higher than that in control group (1.1321±0.1750 vs 0.7971±0.0955, P<0.05). (3) After stimulated with ConA, the concentration of intracellular calcium in AS group was higher than that in control group. (4) In AS group, the releases of cytokines of IL-2 and TNF-α in AS group were significantly higher when stimulated with ConA for 48 h than that for 24 h. (5) The mRNA expression of Kv1.3 in CD4⁺ T lymphocytes was greatly higher in AS group than that in control group (3.670±1.579 vs 1). CONCLUSION: In AS rats, the increase in CD4⁺ T lymphocytes as well as the augmentation of Kv1.3 mRNA expression in the cells suggest that up-regulation of Kv1.3 mRNA expression in CD4⁺ T lymphocytes may be involved in the mechanism of atherosclerotic formation in rat spleen.     

Effects of pituitary adenylate cyclase-activating polypeptide on cardiac remodeling in coronary atherosclerotic rabbits

AIM:To study the intervention effects of pituitary adenylate cyclase-activating polypeptide (PACAP) on cardiac remodeling during the development of rabbit coronary atherosclerosis. METHODS:60 male New Zealand rabbits were equally divided into 3 groups randomly: control group (C group), atherosclerosis model group (A group) and PACAP intervention group (P group). At the 4th, 8th and 12th week, 5-6 cases of rabbits in each group were sacrificed, cardiac tissue with coronary arteries were harvested to make paraffin sections. The sections were stained with hematoxylin-eosin and van Gieson separately. The qualitative observation and/or quantitative analysis were made by light microscope. RESULTS:（1）There was no lesion in C group. For A group and P group, there were plaques in large epicardial coronary arteries and small coronary arteries; an impressive accumulation of collagen was also observed in myocardium. In P group, the lesions of small coronary arteries were less serious, and the degrees of perivascular and myocardial fibrosis also appeared to be less.（2）For A group, the wall-to-lumen ratios in small coronary arteries were significantly greater at the 12th week (2.58±1.54) than C group (1.34±0.58) and P group (1.39±0.48) （P<0.05）； and the width of cardiomyocyte (13.85 μm±2.27 μm) was already remarkably narrower than C group (14.68 μm±2.40 μm) at the 8th week (P<0.05) and narrower significantly than C group and P group at the 12th week. （3）There were not difference significantly between the above-related parameters of P group and C group (P>0.05). CONCLUSION:Structure changes exist in coronary arteries and myocardium during the development of rabbit coronary atherosclerosis, PACAP can inhibit the cardiac remodeling.     

Effect of atorvastatin on blood pressure in spontaneously hypertensive rats

AIM: To explore the mechanisms underlying the effect of atorvastatin on blood pressure in spontaneously hypertensive rats (SHR). METHODS: The effects of atorvastatin on plasma endothelin-1, aortic nitric oxide synthase, aortic smooth muscle cell (ASMC) apoptosis and p27 expression in SHR were evaluated. 12 eight-week-old SHR were randomized into atorvastatin group (ATV, n=6) and SHR group (n=6). 6 age-matched normotensive Wistar-Kyoto rats (WKY) were served as controls. 50 mg·kg-1·d-1 of atorvastatin was administered to ATV by gavage for 10 weeks. Serum cholesterol and triglycerides were measured, and systolic blood pressure of caudal artery was examined. Plasma endothelin-1 and nitric oxide synthase activity of aortic tissue were measured. ASMC apoptosis rate was detected by TUNEL technique, and positive expression rate of P27 in ASMC was analyzed. RESULTS: After 10 weeks, systolic blood pressure in ATV was significantly lower than that in SHR ［(134.17±3.60）mmHg vs （173.33±3.78）mmHg, P<0.01］. Compared with SHR, serum cholesterol and triglycerides were significantly lower (P<0.01, P<0.01) in ATV. Additionally，atorvastatin significantly decreased plasma endothelin-1 ［(130.04±40.07）ng/L vs （196.74±59.69）ng/L, P<0.05］ and increased nitric oxide synthase activity in aortic tissue ［(0.189±0.040）kU/g protein vs (0.124±0.057)kU/g protein, P<0.01］, compared with SHR. ASMC apoptosis rate was higher in ATV than that in SHR (16.94%±3.08% vs 9.01%±2.36%, P<0.01). Compared with WKY, positive expression rate of p27 in ASMC from ATV was higher (33.02%±5.01% vs 24.25%±4.41%, P<0.05), whereas that was lower in SHR (16.08%±7.09% vs 24.25%±4.41%, P<0.01). CONCLUSION: Atorvastatin may reduce the plasma endothelin-1， up-regulate nitric oxide synthase activity and ASMC P27 expression and facilitate ASMC apoptosis，which may effectively reduce blood pressure in SHR.