Renal expression of ACE/AngⅡ in ACE2 knockout mice after tourniquet shock and its relationship with renal injury

AIM: To investigate the effect of angiotensin-converting enzyme 2 (ACE2) on tourniquet shock (TS)-induced acute renal injury by observing the renal expression of angiotensin-converting enzyme/angiotensin Ⅱ (ACE/AngⅡ) and injury severity in ACE2 knockout (KO) mice. METHODS: The TS animal model was produced by bilateral tourniquet ligation in the inguinal region on both hind legs for 2 h to induce ischemia, and reperfusion was initiated by cutting latex rings for 4 h. Six-month-old male wild-type (WT) and ACE2 KO C57BL/6 mice were selected, and divided into 4 groups (6 mice in each group) including WT group, WT+TS group, KO group and KO+TS group. The expression of ACE and AngⅡ in the renal tissues was determined by Western blotting and ELISA, respectively. The renal content of malondialdehyde (MDA) and activity of superoxide dismutase (SOD), blood urea nitrogen (BUN) and serum creatinine (Cr) were measured by chemical colorimetry. RESULTS: Compared with WT group, the expression of ACE/AngⅡ was obviously increased in WT+TS group, and significant renal oxidative stress injury was also observed. The expression of ACE/AngⅡ was elevated in KO mice, but no significant renal oxidative stress injury was found. Compared with WT+TS group, more highly increased expression of ACE/AngⅡ and more aggravated renal injury were exhibited in KO+TS group. CONCLUSION: Deletion of ACE2 gene exacerbates TS-induced renal oxidative stress injury by increasing local ACE/AngⅡ expression. The agonist targeting to ACE2 may be helpful for prevention and treatment of TS-induced acute renal injury.     

Changes of lung AT1R and Mas receptor protein expression in lung tissue of mice with lung injury after limb ischemia-reperfusion

AIM:To explore the role of imbalance of local renin-angiotensin system (RAS) in lung injury by observing the changes of angiotensin Ⅱ type 1 receptor (AT1R) and Mas receptor protein expression in the lung and the degree of lung injury subject to limb ischemia-reperfusion (LIR) in the mice.METHODS:Male ICR mice (n=42,8 weeks old) were randomly assigned into 7 groups (6 in each group),including control group and 6 model groups with LIR of 0.5 h,1 h,2 h,4 h,6 h and 12 h reperfusion.Tourniquets were used to block the blood flow of the hind limbs of the ICR mice and were released after 2 h ischemia to initiate reperfusion.The mice were sacrificed by eyeball blood withdrawal at different time points after reperfusion.The organ coefficient and wet/dry weight ratio (W/D) of the lung tissue were calculated.Bronchoalveolar lavage fluid (BALF) was taken for cell counting and protein concentration measurement.The histopathological changes of the lung tissues was observed,and the pathological score was calculated.The protein expression of AT1R and Mas receptor in the lung tissues was determined by Western blot.RESULTS:The organ coefficient,W/D of lung tissue,and cell number and protein concentration in BALF of model groups were significantly higher than those in control group after LIR.The pathological changes were found in the lung tissue of model mice,including alveolar capillary dilation and congestion,edema,inflammatory cell infiltration in peripheral vascular,alveolar and bronchial walls,alveolar septal thickening and inflammatory cell infiltration.The lung injury score was elevated gradually along with the extension of reperfusion time.The protein expression of AT1R began to increase at reperfusion time points of 0.5 h and 1 h.With the extension of reperfusion time,the protein expression of AT1R decreased gradually.Conversely,the protein expression of Mas receptor increased gradually with prolonged reperfusion.CONCLUSION:LIR induces acute lung injury gradually.The imbalance of AT1R and Mas receptor expression may be involved in the damage process.     

Change of ACE/ACE2 expression in mouse kidney after tourniquet shock

AIM: To investigate the change of angiotensin-converting enzyme/angiotensin-converting enzyme 2(ACE/ACE2) expression in mouse kidney after tourniquet shock (TS) and to study the effects of ACE/ACE2 imbalance on the kidney of TS mice. METHODS: The male ICR mice were used in the study. The model of TS was made using bilateral tourniquets placed high in the inguinal region on both hind legs for 2 h to induce ischemia, and reperfusion was initiated by cutting latex rings. The expression of ACE/ACE2 in the kidney at different time points was determined by Western blotting and immunohistochemistry. The content of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) in the kidney were determined by chemical colorimetric method. The morphological changes of the kidney were observed under microscope with HE staining. RESULTS: Western blotting showed that ACE was increased and ACE2 was decreased in the kidney after TS. MDA was increased and SOD was decreased in TS groups as compared with control group. The kidney of TS groups displayed distinct injuries including inflammatory cell infiltration and degeneration of tubule epithelial cells. The results of immunohistochemistry showed that ACE expressed in endochylema of the renal tubular epithelial cells was increased and ACE2 expressed in epithelium of proximal tubules was decreased in the TS mice. CONCLUSION: Both ACE and ACE2 are expressed in the kidney. The imbalance of ACE/ACE2 expression in the kidney after TS may be related to the kidney injury.     

Effects of dexamethasone on acute lung injury by inhibiting renin-angiotensin system in rats

AIM:To explore the relationship between renin-angiotensin system (RAS) and acute lung injury (ALI) in rats, and the effect of dexamethasone (DEX) on RAS was observed.METHODS: The rat model of ALI was induced by hemorrhagic shock and LPS administered intraperitoneally. The changes of MAP and the effects of DEX on MAP were observed. The mRNA expressions of ACE, AGT, AT1 and AT2 in lung tissue were assayed by RT-PCR. The changes of Ang I and Ang II in the serum and the effects of DEX on them were observed.RESULTS: The increasing of MAP was statistically obvious. MAP in hemorrhagic shock+LPS (HL) group recovered more slowly than that in HL+DEX(HLD) group. ACE, AGT, AT1 and AT2 mRNA expressions in HL group were increased, and higher than those in HLD group. The change of Ang II in serum in HL group was obviously higher than that in HLD group, while that of Ang I was not obvious.CONCLUSION:ALI activates RAS in rat lung, and promotes the production of Ang II then aggravates the injury of lung through increasing the expression of ACE and AGT. DEX decreases expression of Ang II.     

Role of post-hemorrhagic shock mesenteric lymph drainage in restoring balance of ACE/ACE2 in kidney of mice

AIM: To study the role of post-hemorrhagic shock mesenteric lymph (PHSML) drainage on the balance of angiotensin-converting enzyme (ACE) and ACE2 in the kidney. METHODS: A hemorrhagic shock model was established and then fluid resuscitation was performed to the animals in shock and shock+drainage groups, and the PHMSL was drained in shock+drainage group after fluid resuscitation. After 6 h of resuscitation, the mRNA expression of ACE, ACE2, angiotensin Ⅱ (Ang Ⅱ) type 1 receptor (AT1R) and Mas-related G-protein-coupled receptor (MasR), and the levels of Ang Ⅱ and Ang (1-7) in the renal tissues were observed. RESULTS: Hemorrhagic shock increased the levels of ACE mRNA, AT1R mRNA and Ang Ⅱ, and decreased the levels of ACE2 mRNA, MasR mRNA and Ang(1-7) in the kidney. PHSML drainage abolished the effect of hemorrhagic shock on ACE2 and AT1R mRNA expression. Meanwhile, PHSML drainage reduced the hemorrhagic shock-induced increases in the ratios of ACE/ACE2, Ang Ⅱ/Ang(1-7) and AT1R/MasR. CONCLUSION: The PHSML drainage restores the balance of ACE/ACE2, which is beneficial to alleviate acute kidney injury following hemorrhagic shock in the mice.     

Mechanism of calcium sensitivity in lymphatic hypo-reactivity in hemorrhagic shock rats

AIM: To observe the changes of lymphatic reactivity to norepinephrine (NE) and calcium sensitivity in vitro in hemorrhagic shock (HS) rats. METHODS: Male Wistar rats were randomly divided into sham group (with only operation), HS group (duplicating HS model, and divided into shock 1 h and shock 2 h subgroups). The thoracic duct rings (n=48 in each group) were prepared for assaying the lymphatic reactivity to NE and calcium sensitivity by lymphatic tension measurement technique in vitro with isolated perfusion system. Meanwhile, the effects of angiotensin Ⅱ (Ang Ⅱ) and insulin (Ins) on lymphatic reactivity were also observed. RESULTS: Compared with sham group, the NE concentration-response curves in HS 1 h and HS 2 h groups, and calcium concentration-response curves in HS 2 h group were obviously shifted to right. The lymphatic reactivity to NE, contraction to calcium, maximum effect(E_max)and avidity index (pD₂) were markedly reduced. In HS group, after incubating with calcium sensitizer Ang Ⅱ, the lymphatic reactivity to NE and calcium sensitivity were significantly increased but reduced in sham group. However, calcium sensitivity inhibitor Ins decreased the lymphatic contractile response to NE and Ca²⁺. CONCLUSION: The lymphatic hypo-reactivity in hemorrhagic shock rats is related to calcium desensitization, indicating a mechanism of lymphatic hypo-contraction.     

Angiotensin 1-7 attenuates angiotensin Ⅱ-induced human glomerular endothelial cell injury via Mas receptor

AIM:To investigate the effect of angiotensin 1-7 (Ang1-7) on the human glomerular endothelial cells (HGECs) injury induced by angiotensin Ⅱ (Ang Ⅱ) and its possible mechanism. METHODS:Cultured HGECs were divided into 6 groups randomly:control group, Ang Ⅱ group, Ang1-7 group, Ang Ⅱ +Ang1-7 group, Ang Ⅱ +Ang1-7+A779 (an inhibitor of Mas receptor) group and A779 group. The apoptotic rate and reactive oxygen species (ROS) of HGECs were analyzed by flow cytometry and photographed by fluorescence microscopy. The levels of lactate dehydrogenase (LDH), nitric oxide (NO), endothelin-1 (ET-1), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), transforming growth factor-β (TGF-β), monocyte chemoattractant protein-1 (MCP-1) and intercellular adhesion molecule-1 (ICAM-1) in the supernatant of cell cultures were measured. RESULTS:Compared with the control group, the apoptotic rate and the average fluorescence intensity of ROS were increased in the Ang Ⅱ group, IL-6, TNF-α, TGF-β, ICAM-1 and MCP-1 in cell supernatants were also increased in the Ang Ⅱ group (P<0.05). Compared with the Ang Ⅱ group, the apoptotic rate, ROS level, and the above inflammatory factors were decreased in Ang Ⅱ +Ang1-7 group (P<0.05). Compared with the Ang Ⅱ +Ang1-7 group, adding A779 increased the cell apoptotic rate, ROS production and the releases of the above inflammatory factors in cell supernatants (P<0.05). Compared with the Ang Ⅱ group, adding Ang1-7 inhibited the LDH leakage, ET-1 secretion and promoted the release of NO in a dose-dependent manner (P<0.05). CONCLUSION:Ang1-7 attenuates the HGECs injury induced by Ang Ⅱ by inhibiting the Mas receptor.     

Blockage of mesenteric lymph return promotes the vascular reactivity and calcium sensitivity in hemorrhagic shock rats

AIM: To observe the effects of mesenteric lymph duct ligation and mesenteric lymph drainage on the vascular reactivity and calcium sensitivity in hemorrhagic shock (HS) rats, and to investigate the role of mesenteric lymph on the vascular hyporeactivity during shock. METHODS: Seventy-two male Wistar rats were randomly divided into sham group (only operation), shock (duplicating HS model) group, shock+ligation group (duplicating HS model and mesenteric lymph duct ligation) and shock+drainage group (duplicating HS model and mesenteric lymph drainage). The changes of mean artery pressure (MAP) after injection of norepinephrine (NE, 3 μg/kg) at different time points were recorded. After hypotension (40 mmHg) for 3 h, the vascular ring of superior mesenteric artery (SMA) was made for determining the vascular reactivity and sensitivity to calcium by observing the contraction initiated by NE and Ca²⁺ under depolarizing conditions (120 mmol/L K⁺) in the isolated organ perfusion system. Meanwhile, the effects of angiotensin Ⅱ (AngⅡ) and insulin (Ins) on the vascular reactivity were also observed. RESULTS: Compared to sham group, the △MAP in shock group was increased significantly at 0 h and 0.5 h after shock, and that was decreased markedly at 1.5 h, 2 h, 2.5 h and 3 h after shock, respectively, and that in shock+ligation group and shock+drainage group was increased at 0 h, 0.5 h and 1 h after shock, decreased at 2.5 h and 3 h after shock, respectively. The △MAP in shock+ligation group and shock+drainage group was higher than that in shock group at 0.5 h after shock and all the time points followed. The SMA reactivity to NE and sensibility to Ca²⁺ in shock group, shock+ligation group and shock+drainage group were lower markedly than those in sham group. The vascular reactivity and calcium sensitivity in shock+ligation and shock+drainage groups were higher than those in shock group. The vascular reactivity and calcium sensitivity in shock group, shock+ligation group and shock+drainage group were lower than those in sham group, and those in shock+ligation and shock+drainage groups were increased as compared to shock group, respectively. CONCLUSION: Blockage of mesenteric lymphatic return with the methods of mesenteric lymph duct ligation and mesenteric lymph drainage promotes the vascular reactivity of HS rats. The mechanism may be related to improving the calcium sensitivity in the vasculature.     

Over-expression of ACE2 gene alleviates Ang Ⅱ-induced oxidative stress in neural cells

AIM:To investigate the effect of over-expression of angiotensin-converting enzyme 2 (ACE2) gene on angiotensin Ⅱ (Ang Ⅱ)-induced oxidative stress and NADPH oxidase (NOX) expression in mouse neuroblastoma Neuro-2A cells. METHODS:The recombinant lentivirus encoding ACE2 gene was constructed and transfected into the Neuro-2A cells at a multiplicity of infection (MOI) of 10 for 72 h. The transfection efficiency of ACE2 gene and protein expression of ACE2 were detected, and the Neuro-2A cells were identified by detection of a neural cell marker. The Neuro-2A cells were divided into 7 groups:control group, eGFP group, ACE2-eGFP group, Ang Ⅱ treatment group, Ang Ⅱ-eGFP group, Ang Ⅱ-ACE2-eGFP group and Ang Ⅱ-ACE2-eGFP-A779 group. The Ang(1-7) level was determined by ELISA. The level of reactive oxygen species (ROS) in the cells was measured with a method of DHE staining. The protein expression of MAS receptor and NOX subunits (NOX2, NOX4, p47^phox and p67^phox) was detected by Western blot. RESULTS:Ang Ⅱ signi-ficantly increased ROS levels (P<0.01) and up-regulated the protein expression of NOX2, NOX4, p47^phox and p67^phox (P<0.01), but down-regulated MAS protein expression (P<0.01). Over-expression of ACE2 inhibited Ang Ⅱ-induced increase in ROS, down-regulated the protein expression of NOX2, NOX4, p47^phox and p67^phox,and still increased the Ang(1-7) level (P<0.01) and MAS receptor expression (P<0.01). An antagonist of the MAS receptor, A779, blocked the down-regulating effect of ACE2 on NOX expression (P<0.05). CONCLUSION:ACE2 over-expression antagonizes Ang Ⅱ-induced oxidative stress via MAS receptor in the neural cells.     

Effects of Radix Angelicae Sinensis on changes of IL-6, TNF-α and bFGF in renal ischemia/reperfusion injury in rabbits

AIM: To determine the effect of Radix Angelicae Sinensis(RAS) on renal ischemia/reperfusion injury in rabbits and to explore its mechanism. METHODS: Twenty-five rabbits were divided randomly into the sham operated group(Control group), renal ischemia/reperfusion injury group(IR group) and RAS+IR group. At the time point of reperfusion 48 h after renal ischemia 1 h, the renal tissue were observed by electron-microscope and the contents of creatinine(Cr) in serum, tumor necrosis factor-α(TNF-α), interleukin-6(IL-6)and basic fibroblast growth factor(bFGF) in the renal tissue were measured. RESULTS: A remarkably degenerative changes in renal tissue were showed under electronmicroscope in IR group, but the changes in RAS+IR group were slight. The contents of Cr, TNF-α and IL-6 in IR group were higher than those in Control group, these parameters in RAS+IR group were lower than those in IR group, the difference between these groups was significant(P< 0.05 or P< 0.01). At the same time, the content of bFGF in IR group was lower than that in Control group(P< 0.01), while the content of bFGF in RAS+IR group was higher than that in IR group(P< 0.01) and Control group(P< 0.05). CONCLUSION: RAS has an effect of alleviating the renal ischemia/reperfusion injury by modulating the production or release of TNF-α, IL-6 and bFGF.     

Role of angiotensin Ⅱ in the regulation of platelet-derived growth factor receptor β subunit of vascular smooth muscle

AIM:To investigate the crosstalk between angiotensin Ⅱ (AngⅡ)-mediated and platelet-derived growth factor (PDGF)-mediated signal transduction in vascular smooth muscle proliferation.METHODS:A model of renal hypertension was made by two kidney/one-clip operation. Level of PDGF receptor β subunit of aorta was measured by Western Blot analysis. The effect of Ang Ⅱ on PDGF receptor β subunit expression was investigated in culture rat aortic vascular smooth muscle cells (VSMC).RESULTS:Systolic blood pressure obviously increased at 8th week after operation, whereas the level of PDGF receptor β subunit of aorta significantly increased by 126.6% (P＜0.05) in 2K1C rats compared with control group. The expression of PDGF receptor β subunit in cultured VSMC stimulated by AngⅡ was higher than that of control by 192.74%(P＜0.01). The effect of AngⅡ was inhibited remarkably by pretreated with losartan, a kind of specific AngⅡ receptor 1 (AT₁) subtype antagonist and U73122, a kind of phospholipase C inhibitor. The effect was partly blocked by PD98059, which inhibit the activity of mitogen-activated, ERK-activating kinase (MEK).CONCLUSION:AngⅡ-induced PDGF receptor β subunit expression is regulated by the AT₁ and its downstream signal molecule-PLC and ERK, might participate in the intracellular signal transduction pathway.     

Role of endogenous angiotensin II in the pathogenesis of aortic calcification in rats

AIM: To explore the effects of angiotensin II on aortic calcification in the rat. METHODS: Arterial calcification of Sprague-Dawley rats was induced by vitamin D₃ plus nicotine. Calcification was confirmed by Von Kossa staining, measurement of calcium content, [⁴⁵Ca²⁺]accumulation and alkaline phosphatase (ALP) activity of vascular tissue. RESULTS: The results showed that calcium content, [⁴⁵Ca²⁺]accumulation and ALP activity in calcified arteries increased significantly compared with those of control. Ang Ⅱ levels in plasma and aortic tissues and the amount of angiotensinogen mRNA in calcified aorta were also increased as compared with control. Captopril (inhibitor of ACE) and losartan (Ang Ⅱ receptor inhibitor) decreased significantly the content of calcium, [⁴⁵Ca²⁺] uptake and ALP activity in calcified aorta. Ang Ⅱ levels in plasma and aortic tissues and the amount of angiotensinogen mRNA in aortic tissue were down-regulated by captopril. The amount of angiotensinogen mRNA and the content of Ang Ⅱ in the calcified aorta were also decreased by losartan. CONCLUSION: The captopril and losartan significantly alleviate the vascular calcification.     

Effect of ischemic preconditioning on vascular reactivity and calcium sensitivity in hemorrhagic shock rats

AIM: To investigate the effect of ischemic preconditioning (IPC) on vascular reactivity and calcium sensitivity during hemorrhagic shock. METHODS: Appropriate method of IPC was selected by observing the effect of different strategies of IPC on the survival time and the survival rate in hemorrhagic shock rats. The effect of IPC on the pressor effect of norepinephrine (NE, 3 μg/kg) and the contractile response of superior mesenteric artery (SMA) to NE and calcium in vivo and in vitro were observed. RESULTS: Among 3 strategies of IPC, 3 cycles of abdominal aorta occlusion for 1 min and loosing for 5 min increased the survival time and 24 h survival rate significantly, which was superior to the other two IPC methods. In vivo, IPC significantly increased the pressor response to NE and the contractile response of SMA to NE (P<0.01). In vitro, IPC significantly improved the reactivity of SMA to NE and Ca²⁺. The E_max values of SMA to NE and Ca²⁺ in IPC group were significantly higher than that in shock control group (P<0.01). CONCLUSION: Ischemic preconditioning reverses Shock-induced vascular hyporeactivity via improving calcium sensitivity of the vasculatures.     

Effect of angiotensin Ⅱ on the remodeling of aorta in spontaneously hypertensive rats

AIM: To investigate the effects of a 10-weeks treatment with angiotensin Ⅱ (Ang Ⅱ) subtype I receptor antagonist losartan on vascular remodeling of thoracic aorta in male spontaneously hypertensive rats (SHR). METHODS: SHR were treated from 16 to 26 weeks of age with losartan at 15 mg/kg·d^-1 or 0.75 mg/kg·d^-1. RESULTS: Losartan (15 mg/kg·d^-1) treatment significantly decreased systolic blood pressure compared with the control group, while losartan (0.75 mg/kg·d^-1) had no the effect, losartan(15 mg) prevents the development of aortic hypertrophy by preventing hypertrophy of vascular smooth muscle cells (VSMC). In the losartan 0.75 group, these parameters were not changed. But in the losartan 15 and losartan 0.75 groups, the collagen content of the aortic media decreased significantly. CONCLUSION: It is inferred that the effect of Ang Ⅱ on stimulating VSMC growth of the aorta in SHR is dependent on arterial pressure, while the effect on collagen fibers is through pressure independent mechanism.     

Effect of non-invasive ischemic preconditioning on nitric oxide/endothelin-1 imbalance and gas exchange impairment following limb ischemia reperfusion: a clinical study

AIM: To investigate the effects of non-invasive ischemic preconditioning on nitric oxide (NO)/endothelin-1 (ET-1) imbalance and gas exchange impairment following limb ischemia reperfusion in patients undergoing unilateral lower extremity surgery with tourniquet. METHODS: Twenty-seven patients aged 25-65 years, whose tourniquets duration varied from 1 h to 1.5 h and matched American Society of Anesthesiologists Physical Status Ⅰ-Ⅱ, were randomized into two groups: a control group (n=14) and a ischemic preconditioning group (IPC group, n=13) in which patients received three cycles of 5 min of ischemia/5 min of reperfusion before tourniquet inflation. Radial arterial blood gas, plasma malondialdehyde (MDA) and NO, serum ET-1 and interleukin-6 (IL-6) were measured just before tourniquet inflation(T0), 1 h after inflation(T1), and 0.5 h(T2), 2 h(T3), 6 h(T4), 24 h(T5) after tourniquet deflation. Meanwhile NO/ET-1 ratio, alveolar-arterial oxygen gradient (PA-aDO2) and intrapulmonary shunt (Qs/Qt) were calculated. RESULTS: In control group, arterial partial pressure of oxygen (PaO2) were decreased, while PA-aDO2 and Qs/Qt were increased significantly at T4 compared to the baselines at T0 (P<0.01). Plasma NO levels and NO/ET-1 ratios decreased gradually after tourniquets deflation and statistical significances were observed at T3 (P<0.01) with a valley at T4 (P<0.01) and recovered to baselines at T5. Serum ET-1, IL-6 and plasma MDA began to increase remarkably after T3 (P<0.05 or P<0.01), peaked at T4 and dropped slightly at T5. The changes above-mentioned could be well attenuated by the application of IPC (P<0.05 or P<0.01) except PaO2 (P>0.05). CONCLUSION: Clinical application of unilateral tourniquet within safe time limit (1.5 h) may lead to limb ischemia reperfusion and further pulmonary gas exchange impairment, which could be partially attenuated by the application of IPC via alleviating NO/ET-1 imbalance.     

Effect of high-fructose diet on adipose tissue renin-angiotensin system in rats

AIM: To observe the effects of high-fructose diet on adipose tissue inflammation and renin-angiotensin system (RAS), and to reveal the role of Toll-like receptor 2 (TLR2) in this process.METHODS: Male SD rats (n=16) were randomly divided into control group, high fructose group, high fructose+siRNA negative control group, and high fructose+TLR2-siRNA group. The rats in control group were fed with a standard chow diet. The rats in high fructose group were fed with a diet with 60% fructose, and the rats in high fructose+TLR2-siRNA group and high fructose+siRNA negative control group were transfected with TLR2 siRNA and scrambled siRNA, respectively. Serum uric acid was measured and visceral adipose tissue was weighed at the 14th week. The levels of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), angiotensinogen (AGT), and angiotensin Ⅱ (Ang Ⅱ) were measured by ELISA. Infiltrating macrophages in the adipose tissues were measured with immunohistochemistry. The mRNA expression of IL-6, TNF-α, monocyte chemoattractant protein-1 (MCP-1), AGT, angiotensin-converting enzyme 1 (ACE1), angiotensin Ⅱ type 1 receptor (AT1R), and angiotensin Ⅱ type 2 receptor (AT2R) was detected by RT-qPCR. The protein level of TLR2 was determined by Western blot.RESULTS: High fructose-fed rats showed elevated serum uric acid, raising fat content, higher serum concentrations of IL-6, TNF-α, AGT and AngⅡ, and more infiltrating macrophages in the adipose tissues (P<0.05). Moreover, the mRNA levels of IL-6, TNF-α,MCP-1, AGT, ACE1, AT1R and AT2R in the adipose tissues were increased (P<0.05). When high fructose-fed rats were transfected with TLR2-siRNA, the dramatic decreases in TLR2 protein level and number of infiltrating macrophages in the adipose tissues were found. Both in serum and adipose tissues, the mRNA levels of inflammatory cytokines and RAS components were all significantly decreased (P<0.05).CONCLUSION: High-fructose diet up-regulates RAS in adipose tissues via activation of TLR2 inflammation signaling pathway.     

Expression of calcium sensing receptor during ischemia/reperfusion myocardial damage and relationship between CaSR and injury of myocardium

AIM: To investigate the expression of calcium sensing receptor(CaSR) during myocardial injury induced by ischemia/reperfusion and disclose the relationship between CaSR and myocardial ischemia/reperfusion. METHODS: The experimental model was established by the 30 min ligating and 1 h, 2 h, 4 h, 6 h reperfusing the left descending coronary artery (LAD) in rats. Wistar rats were randomly divided into 5 groups: sham group, ischemia/reperfusion 1 h, 2 h, 4 h, 6 h groups (I/R 1 h, 2 h, 4 h, 6 h group). CaSR mRNA expression was detected by RT-PCR. Left ventricular function was recorded. The levels of plasma lactate dehydrogenase (LDH), alondialdehyde (MDA) and superoxide dismutase (SOD) were measured. The change of ultrastructure in the ischemia/reperfusion myocardium of rats was observed by electron microscopy. RESULTS: LVSP,±dp/dtmax and SOD activity decreased gradually with the reperfusion time prolonged. LDH and MDA peaked at 2 h. The ultramicro-structural injury at the 1 h and 2 h was more serious than that at 4 h and 6 h. The expression of CaSR increased significantly after reperfusion of 1 h and 2 h, and decreased after 4 h and 6 h. CONCLUSION: The increased expression of CaSR mRNA and serious injure of myocardium were observed. CaSR may be associated with the myocardial ischemia/reperfusion injury.     

Effects of external counterpulsation on renin-angiotensin system and hemodynamics in dogs with myocardial ischemia

AIM:To examine the effect of external counterpulsation(ECP) on renin-angiotensin system(RAS) and hemodynamics in dogs with myocardial ischemia and their relationship. METHODS:Acute myocardial ischemia was induced by occluding the left anterior descending of the dogs. The local renin activity, angiotensin Ⅱ(AngⅡ) level, angiotension-converting enzyme(ACE) activity were tested by biochemical methods and hemodynamics was recorded by a 8-channel physiological recorder.RESULTS:Ischemia could actiivate renin,ACE and AngⅡ in cardiovasculature and ECP reduced them except for renin in ischemic myocardium. Ischemia also could activate RAS in lung and kidney, which play an important role on circulating RAS, and ECP reduced them. Furthermore, ECP could improve hemodynamics and there existed a close relationship between local AngⅡlevel and hemodynamics. CONCLUSION:ECP can reduce local RAS and improve hemodynamics in dogs with myocardial ischemia, which might be one of mechanisms underlying the protective effect of ECP on ischemic myocardium.     

Role of A3 adenosine receptor on vascular reactivity of superior mesenteric artery in hemorrhagic shock in rat

AIM: To explore whether A3 adenosine receptor plays a role in the modulation of vascular reactivity after hemorrhagic shock in rat, and to find out the prospective drug target to restore the decreased vascular reactivity following hemorrhagic shock. METHODS: The hemorrhagic shock (40 mmHg) model was established in rat, and the reactivity of superior mesenteric artery (SMA) to norepinephrine (NE) was observed. A3AR expression at protein level and mRNA level were measured by Western blotting and RT-PCR respectively. RESULTS: The vascular reactivity of SMA to NE after hemorrhagic shock (40 mmHg) was decreased significantly in a biphasic response manner. The expression of A3AR mRNA in SMA after hemorrhagic shock decreased without significant difference. The expression of A3AR protein has a slight increase without statistical difference after 30 min of hemorrhagic shock and then has a significant decrease (especially at 2 h and 4 h after hemorrhagic shock). The usage of IB-MECA, a selective A3AR agonist, significantly increased the responsiveness of SMA to NE in hemorrhagic shock in rat. MRS1523, the selective A3AR antagonist, significantly abolished the restoration of the vascular reactivity to NE by IB-MECA in hemorrhagic shock in rat. CONCLUSION: A3AR plays a role in the modulation of vascular responsiveness to NE in hemorrhagic shock in rat, and the selective agonist of A3AR could restore the reactivity of SMA to NE in hemorrhagic shock in rat.     

Effects of imidapril on blood gas,oxidative stress and inflammatory factors in rats with acute lung injury induced by ammonium chloride

AIM: In this study, the rat lung injury model was induced by ammonium chloride for studying the effect of imidapril on blood gas, serum TNF-α, IL-6 and MDA concentrations, and AngⅡ and CD54 protein expression in rat lung tissue. METHODS: Male rats were randomly divided into 3 groups: control group, lung injury model group and drug group. The rats in control group were given saline (2 mL/kg), while the rats in lung injury model group were given 6% ammonium chloride (2 mL/kg). In drug group, imidapril (3 mg·kg^-1·d^-1) was given to the rats once daily for 1 week by intragastric gavage after given 6% ammonium chloride. On the 7th day, the rats were anesthetized with 2% so-dium pentobarbital. Abdominal aorta blood, venous blood and lung tissue were collected. The blood gas indexes and serum TNF-α, IL-6 and MDA concentrations were determined. The lung tissues were fixed and sliced, and the expression of AngⅡ and CD54 proteins was detected by immunohistochemistry. RESULTS: The PaCO₂ increased in lung injury model group compared with control group and drug group (P < 0.05).The expression of AngⅡ and CD54, and the concentrations of TNF-α, IL-6 and MDA also increased significantly (P < 0.01) in model group. Pulmonary edema, inflammation, alveolus congestion, hemorrhage and hyperplasia in model group were obvious compared with control group and drug group. CONCLUSION: Imidapril improves blood gas indexes, and reduces lipid peroxidation and inflammatory responses in the rats with lung injury induced by ammonium chloride.