Effect of hypertonic saline solution on the geometry of erythrocyte in rats subjected to hemorrhagic shock

AIM:To study the effect of hypertonic saline solution on the geometry of erythrocyte in hemorrhage-shocked rats,using micropippette aspiration technique. METHODS:Wistar rats were randomized into three groups of 0.9% NaCl(NS),7.5% NaCl(HS) and 5% NaCl-3.5% NaAc(HSA). The animals were bled to reach a mean arterial pressure of 5.3 kPa in 10 minutes and maintained in shock for 90 minutes. 4 mL/kg NS,HS or HSA was given intravenously and respectively in 5 minutes following hemorrhagic shock. The blood was collected to determine the geometry of erythrocyte at baseline,after shock and treatment. RESULTS:There were no significant difference in surface area/volume ratio and spherical index between baseline and shock.The diameter of erythrocyte were decreased obviously following hemorrhagic shock. The surface area/volume ratio was significantly lower in HS group than that in NS and HSA group after treatment. HS and HSA raised spherical index of erythrocyte remarkbly compared with NS.Otherwise,the spherical index of erythrocyte was reduced significantly in HSA group compared to HS group. Both HS and HSA decreased diameter of erythrocyte notably compared with NS. CONCLUSION:Both HS and HSA have deleterious effects on the geometry of erythrocyte in rats subjected to hemorrhagic shock.Whereas, HSA could decrease the side effect compared with HS.     

Influence of fluid resuscitation on pancreatic microthrombosis in severe acute pancreatitis

AIM: To make evaluation on fluid resuscitation with either hypertonic saline (HS) or dextran 40 (Dx) on pancreatic microthrombi and dysfunction of microcirculation in rats with severe acute pancreatitis (SAP).METHODS: SD rats were allocated into 4 groups randomly,ie.SAP group,HS group,Dx group,which respectively received normal saline (NS,4 mL/kg),HS (4 mL/kg),Dx (4 mL/kg) for 2 h by the tail intravenous injection consecutively after being made as SAP animal models,and operate sham group (OS).12 h after the operation,all animals were blooded to assess the serum amylase levels,plasma D-dimer,von Willebrand factor and GMP-140 levels.The amount of ascites was measured and the samples of the pancreas were collected for pathologic examination under light microscopy as well as transmission electron microscope.The numbers of pancreatic microthrombi were also counted with microscopy.RESULTS: (1) 12 h later when the rats were sacrificed,the survival rate in SAP group was the lowest,significantly lower than that in the 2 fluid resuscitation groups (P<0.05).However,no significant difference was found between the latter 2 groups though the survival numbers.(2) The numbers of microthrombi in the pancreas and the histological scoring of pancreatic injury were significantly fewer in the 2 fluid resuscitation groups than those in SAP group (P<0.05) though no significant difference was revealed between the 2 treated groups.The amounts of ascites,serum amylase,plasma D-dimer,von Willebrand factor and GMP-140 showed significant improvement in HS or Dx group compared to those in SAP group(P<0.05,P<0.01).However,no significant difference was found between the 2 fluid resuscitation groups.CONCLUSION: Fluid resuscitation with HS or Dx,if applied at the early stage,increases survival rate with lessened histological injury,fewer microthrombi and improved coagulation parameters including D-dimer,von Willebrand factor and GMP-140.Either hypertonic saline (HS) or dextran 40 (Dx) demonstrates the same improvement in the relevant laboratory parameters in SAP.     

Dexmedetomidine attenuates acute kidney injury induced by hemorrhagic shock/resuscitation in rats

AIM: To investigate the effects of dexmedetomidine on hemorrhagic shock/resuscitation (HS/R)-induced acute kidney injury (AKI) in rats, and to explore the possible mechanisms. METHODS: Wistar rats (n=32) were randomly divided into 4 groups (n=8):normal saline control group (NS group), dexmedetomidine group (D group), HS/R group and HS/R+D group. The animals were sacrificed at 6 h after resuscitation. The levels of serum creatinine (Cr) and blood urine nitrogen (BUN) were examined. The kidneys of all rats were removed for evaluation of histological characteristics, and the levels of malondialdehyde (MDA), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and superoxide dismutase (SOD) were measured. The expression of nuclear factor-κB (NF-κB) and hemeoxygenase-1 (HO-1) was determined by Western blot. RESULTS: Compared with NS group, the levels of Cr, BUN, MDA, TNF-α and IL-1β were obviously increased in HS/R group, which were obviously decreased in HS/R+D group (P<0.05). Compared with NS group, the SOD activity was obviously decreased in HS/R group, which was obviously increased in HS/R+D group (P<0.05). Compared with NS group, the protein expression of NF-κB was obviously increased in HS/R group, which was obviously decreased in HS/R+D group (P<0.05). Compared with NS group, the protein expression of HO-1 was increased in HS/R group. Compared with HS/R group, the protein expression of HO-1 was obviously increased in HS/R+D group. Compared with NS group, HS/R induced marked kidney histological injury, which was less pronounced in HS/R+D group.CONCLUSION: Dexmedetomidine effectively protects rats against AKI caused by HS/R, and its mechanism may be associated with the increase in HO-1 expression and the inhibition of NF-κB expression.     

Potassium antagonizes damage of coronary artery induced by high salt intake

AIM：To investigate the effect of potassium treatment on coronary arterial impairment induced by high salt intake. METHODS：Sprague-Dawley rats (4-week-old, n=10 in each group) received distilled water (NS), water containing 1.5% NaCl (HS), or 1.5% NaCl and 0.5% KCl (HS+HP) for 16 weeks. Systolic blood pressure (SBP) was determined by tail plethysmography every 2 weeks. After 16 weeks of treatment, vascular remodeling, superoxide production, malondialdehyde (MDA) content, and endothelial nitric oxide synthase (eNOS) and gp91 expression in the coronary arteries were detected. RESULTS：After 16 weeks of salt loading, the rats in HS group was divided into salt sensitive subgroup and salt resistance subgroup according to the tail-cuff blood pressure. In this experiment, the salt-sensitive rats were selected as HS group. In HS group, salt loading significantly increased SBP, serum MDA and gp91 expression, decreased serum NO and eNOS expression in the coronary arteries, and induced the coronary artery remodeling compared with NS group. In salt-loaded SD rats, 16-week potassium treatment abrogated the effects induced by salt loading. CONCLUSION：High salt may affect structural and functional changes in coronary arteries by activating oxidative stress. Potassium treatment antagonizes the effect of high salt intake.     

Experimental study on pathogenesis of salt-induced hypertension

AIM:To reveal the pathogenesis of salt-induced hypertension.METHODS:Forty male SD rats were divided into five groups, which received on same chow but different drink. Control(NC)group: deionized water; High salt(HS)group: 1.5% NaCl solution; L-arginine(HS+Arg)group: L-arginine(4 g·kg^-1·d^-1)in 1.5% NaCl solution; Enalapril (HS+En) group: enalapril (30 mg·kg^-1·d^-1) in 1.5% NaCl solution; Terazozin(HS+Ter)group: terazozin(4 mg·kg^-1·d^-1)in 1.5% NaCl solution. At the end of 8 weeks, rats were anesthetized with pentobarbital sodium. Blood pressure(BP)were recorded and blood were drawn from inferior vena cava and kidneys, adrenals were removed. NO(x),ET and AngII, Na^＋-K^＋-ATPase and proscillaridin-like compound(PLC)were assayed.RESULTS:BP, PLC and ET in plasma and AngII in adrenal were increased, NO(x)and AngII in plasma and kidney were decreased in HS group compared with NC group.CONCLUSION:High salt intake may induce hypertension in SD rats. In addition to the Na^＋-K^＋-ATPase activity was inhibited by increased sodium-pump inhibitors, NO release decrease may also play an important role in the pathogenesis of hypertension.     

Role of Cdk5-CRMP pathway in sevoflurane-induced dendritic developmental disorder of neurons in prefrontal cortex of neonatal rats

AIM: To investigate the effect of sevoflurane (Sevo) on the dendritic development in prefrontal cortex (PFC) of neonatal rats and the role of cyclin dependent kinase 5 (Cdk5)- collapsin response mediator protein (CRMP) pathway in it. METHODS: Eighty-eight postnatal day 7 Sprague Dawley (SD) rats were randomly divided into 4 groups (n=22): Air+NS group, Air+roscovitine (Ros) group, Sevo+NS group and Sevo+Ros group. The rats in Air+NS group and Air+Ros group were exposed to the air for 4 h, while the rats in the other 2 groups were exposed to 2.8% sevoflurane for 4 h. The rats received intraperitoneal injection of 150 μL normal saline 15 min before exposure in the Air+NS group and Sevo+NS group, while the rats in the Air+Ros group and Sevo+Ros group received intraperitoneal injection of 150 μL roscovitine (in DMSO solution, 10 mg/kg) 15 min before exposure. At the end of exposure, the cortices of the rat brain were collected and the protein levels of P35, P25, Cdk5, CRMP1, CRMP2, CRMP4 and p-CRMP2 Ser522 in PFC were detected by Western blot. On the postnatal day 30, the rat brains were sectioned for Golgi-Cox staining and morphological analysis of dendrites in the PFC neurons. Open-field test and contextual fear conditioning test were performed on postnatal days 25~27 and 31~32, respectively. RESULTS: Compared with Air+NS group, the expression of P35 in the Sevo+NS group was significantly decreased, and the expression of P25 was dramatically increased (P <0.05), whereas roscovitine partly reversed the changes above induced by sevoflurane (P <0.05). The expression of Cdk5 was not significantly different among all groups. Compared with the Air+NS group, the expression of CRMP1, 2, and 4 in the Sevo+NS group were decreased, and the protein level of p-CRMP2 Ser522/CRMP2 was increased (P <0.05), whereas roscovitine partly reversed the changes above induced by sevoflurane (P <0.05), except for the expression of CRMP2. Compared with Air+NS group, the total dendrite length, secondary dendritic length and interactions on 60 and 80 μm shells in the Sevo+NS group were decreased (P <0.01), whereas roscovitine partly reversed the changes above induced by sevoflurane (P <0.05). Compared with Air+NS group, the percentage of freezing time in the Sevo+NS group was decreased (P <0.01), whereas roscovitine partly reversed the changes induced by sevoflurane (P <0.05). No significant difference among groups in the open-field test was observed. CONCLUSION: Sevoflurane exposure disturbed dendritic development of neurons in PFC, learning and memory ability of neonatal rats, which may be mediated by Cdk5-CRMP pathway.     

Effects of pyrrolidine dithiocaramate and Vit C on ICAM-1 and SOD gene expression in rats with acute pancreatitis

AIM:To investigate the effects of pyrrolidine dithiocaramate (PDTC) and vitamin C (Vit C) on the intercellular adhesion molecule 1 (ICAM-1) gene expression and the superoxide dismutase (Mn-SOD, Cu, Zn-SOD) gene expression in a rat model of acute pancreatitis (AP). METHODS:Rat AP model was induced by retrograde injection of 3% sodium taurocholate in the bilio-pancreatic duct. The AP rats were divided into three groups: the normal saline (NS) group, the PDTC group and the Vit C group. The three groups were separately administered NS, PDTC and Vit C at 10 min and 5 h after operation. Rat sham operation (SO) group was subjected to laparotomy only with no further treatment. The levels of ICAM expression, Mn-SOD and Cu, Zn-SOD expression in pancreas and liver were determined by RT-PCR at 1, 5 and 10 h after the induction of AP. RESULTS:Comparing with SO group, the NS group showed a higher expression of ICAM-1 and a lower expression of Mn-SOD both in pancreas and in liver, and a lower expression of Cu, Zn-SOD in pancreas. PDTC treatment reduced the ICAM-1 expression and increased the Mn-SOD and Cu, Zn-SOD expression to a certain degree comparing with NS group, especially at 5 h. Vit C treatment also increased the Mn-SOD and Cu, Zn-SOD expression. But no obvious suppressive effect on ICAM-1 expression was observed in Vit C group.CONCLUSIONS:PDTC and Vit C have the effects of enhancing Mn-SOD and Cu, Zn-SOD expression.PDT C has an effect of inhibiting ICAM-1 expression in the pancreas and liver of rats with AP, whereas Vit C dose not.The results suggest that one of the mechanisms of PDTC and Vit C treat ing AP not only anti-oxidate directly by themselves, but also enhance Mn-SOD and Cu, Zn-SOD expression, and that the anti-inflammatory ef ect of PDTC is involved in the suppression of ICAM-1 expression.     

Effect of caveolin-1 expression on high-salt diet-induced endothelial dysfunction in type 1 diabetic rats

AIM: To investigate the underlying mechanisms responsible for endothelial dysfunction of type 1 diabetes mellitus (DM) rats fed with high-salt diet. METHODS: Type 1 DM was induced by intraperitoneal injection of streptozotocin (70 mg/kg). Normal and diabetic rats were fed high-salt food (HS, 8% NaCl) and standard food for 6 weeks, respectively. Isometric tension of the mesenteric arteries were measured. The expression of Akt, endothelial nitric oxide synthase (eNOS) and caveolin-1 (Cav-1) was examined by Western blot. RESULTS: The rats in DM+HS group exhibited more pronounced impairment of vasorelaxation to acetylcholine and insulin compared with either DM group or HS group (P<0.01). Akt and eNOS phosphorylation levels, and nitric oxide (NO) concentration in DM+HS group were significantly lower than those in DM group (P<0.01). The level of Cav-1 in DM+HS group was significantly higher than that in DM group and HS group. CONCLUSION: Impaired endothelial Akt activation, increased Cav-1 expression and resultant decreased eNOS activation contribute to aggravate high-salt diet-induced endothelial dysfunction and hypertension in DM rats.     

Change of cPKC-α level in the ascitic white blood cells of AP rats and effects of tetrandrine and aspirin

AIM: To investigate the alteration of classical protein kinase C-α(cPKC-α)in the ascitic white blood cells of rats with acute pancreatitis (AP), and explore the effects of aspirin (ASP) or tetrandrine (Tet) on them. METHODS: The total of 56 health SD rats were divided into four groups, AP+ASP group, AP+Tet group, AP+normal saline (NS) group, and sham operation control (SO) group. AP model was induced by a retrograde injection of 3% sodium deoxycholate into the pancreatic duct. The AP+Tet group received a intraperitoneal injection of Tet (80 mg/kg), respectively. The AP+ASP group received an infusion of ASP (12.5 mg/100 g) by use of a nose-gastric catheter. At 1 h, 5 h after the treatment, cPKC-α in the ascitic white blood cells of AP rats was measured by Western blot and chemiluminescence, and the semi-quantitative values was obtained by Gel-pro analyzer. RESULTS: The values of cPKC-α decreased in AP+NS group, but increased significantly in the groups treated with ASP and Tet (P＜0.05, or P＜0.01). The levels of AMY increased in AP+NS group and decreased in the treated groups significantly (P＜0.05, or P＜0.01). The morphological lesions in the pancreas were attenuated by these medicines. CONCLUSIONS: 1. ASP and Tet increase cPKC-α levels in the ascitic white blood cells of rats with acute pancreatitis. 2. ASP and Tet have protective effects on the morphology and function of the pancreas in the course of AP.     

Effects of human urotension Ⅱ on in vivo mesenteric microcirculation in rats

AIM: To investigate the effects of human urotensin Ⅱ (hUII) on in vivo mesenteric microcirculation in rats. METHODS: For recording of microcirculation images in the mesentery, the intestinal loop was mounted on the stage of an intravital microscope equipped with a TV camera. Video images of microcirculation were stored by a video cassette recorder. Temporal changes in internal diameter and microcirculatory velocity of microvesseles were measured by computer using the ImagePro software. The blood flow in intestinal wall was measured with PIMII laser Doppler perfusion Imager (Lisca Sweden). RESULTS: The internal diameters of arterioles and venules in control group were (21.4±2.3) μm and (38.1±3.6) μm,respectively. In UII group, the arterioles and venules contracted immediately after treated with UII and up to the peak at 1 min . Both microcirculatory velocity of arterioles and venules showed no significant changes in UII group (compared with control, P＞0.05). The blood flow in intestinal wall increased 1 min after treated with UII and up to high peak at 5 min(6.4±1.1 perfusion unit vs control 4.2±0.9,P＜0.05). CONCLUSION: hUII contracted mesenteric microvesseles in rats and increased microcirculatory blood perfusion in intestinal wall.     

Protective effects and mechanism of heat shock response on cardiovascular system in rats after heat exposure

AIM:To study protective effects and mechanism of heat shock response (HSR) on cardiovascular system in rats after heat exposure.METHODS:The study was divided into 2 experiments:①Protective effects of HSR on cardiovascular system in rats after heat exposure.SD rats randomly allocated into 2 groups:heat shock group (HS group), sham control group(SC group).HS group were treated with heat shock, but SC group weren't.After re-covering for 20 h at room temperature, two groups exposed to death in thermal environment, and blood pressure and elec-trocardiogram were measured continuously.Through Chart software mean arterial pressure(MAP), existent time etc were acquired.②SD male rats randomly allocated into 3 groups:HS group, SC group and normal temperature control group(NC group).NC group weren't treated.The treatment in HS and SC group was identical with in the first experiment, but it would be terminated at 73 min after heat exposure, meanwhile content of MDA of myocardium were measured.RESULTS:① Existent time in HS group was longer than that in SC group and shock arrived later; ② During earlier period after heat exposure MAP had no significant changes between HS and SC group, but after 60 mins MAP in HS group were higher than that in SC group; ③ Compared with NC group, content of MDA in myocardium in SC group was higher significantly at 73 min after heat exposure. Howerer, content of MDA in HS group was lower than in SC group, and had no significant changes with NC group.CONCLUSION:Through decreasing production of MDA in myocardium, HSR has a protective effect on cardiovascular system in rats after heat exposure.     

Protective effect of dexmedetomidine-ulinastatin combination on lipopolysaccharide-induced acute lung injury in rats

AIM：To investigate the effects of dexmedetomidine-ulinastatin combination on acute lung injury induced by lipopolysaccharide (LPS) in rats. METHODS：Male Wistar rats were randomly divided into 5 groups: saline control group (NS group) was given saline (5 mL/kg, iv) alone; LPS group (L group) was given LPS (10 mg/kg, over 10 min); dexmedetomidine+LPS group (L+D group) was treated with the additional administration of dexmedetomidine (1 μg·kg -1·h -1) immediately after LPS injection; ulinastatin+LPS group (L+U group) was treated with the addi-tional administration of ulinastatin (50 000 U/kg, ip) immediately after LPS injection; dexmedetomidine+ulinastatin+LPS group (L+D+U group) received dexmedetomidine (1 μg·kg -1·h -1) and ulinastatin (50 000 U/kg) immediately after LPS injection. The animals were sacrificed at 6 h after LPS or NS administration. Partial pressure of arterial oxygen (PaO 2), pH and base excess (BE) were measured, and the lungs were removed for evaluation of histological characteristics and determining the concentrations of TNF-α, IL-1β, macrophage inflammatory protein 2 (MIP-2), malondialdehyde (MDA), nitric oxide (NO), prostaglandin E 2 (PGE 2) and myeloperoxidase (MPO) in lung tissues, lung wet/dry weight ratio (W/D), and albumin in brochoalveolar lavage fluid (BLAF). The pulmonary expression of nuclear factor kappa B (NF-κB) p65 was evaluated by Western blotting. RESULTS：Compared with NS group, PaO 2, pH and BE was lower in L group, which was increased by treatment with dexmedetomidine-ulinastatin combination but not by dexmedetomidine or ulinastatin alone. Compared with NS group, LPS induced marked lung histological injury, which was less pronounced in the animals treated with dexmedetomidine-ulinastatin combination but not dexmedetomidine or ulinastatin alone. The levels of IL-1β, IL-6, MIP-2, MDA, NO and PGE 2 in the lung tissues increased in L group compared with NS group, which were reduced by dexmedetomidine-ulinastatin combination but not by dexmedetomidine or ulinastatin alone. The MPO activity, MDA level and W/D increased in the lung tissues in L group compared with NS group, which was reduced by dexmedetomidine-ulinastatin combination but not by dexmedetomidine or ulinastatin alone. Compared with NS group, the albumin concentration in the BLAF increased, which was reduced by dexmedetomidine-ulinastatin combination but not by dexmedetomidine or ulinastatin alone. Compared with NS group, the expression of NF-κB p65 increased in L group, which was reduced by dexmedetomidine-ulinastatin combination but not by dexmedetomidine or ulinastatin alone.CONCLUSION：Dexmedetomidine-ulinastatin combination has a protective effect on LPS-induced acute lung injury in the rats.     

Effect of glycine on lipopolysaccharide and hypoxia-induced necrotizing enterocolitis in rats

AIM:To investigate the effect of glycine on endotoxin and hypoxia-induced necrotizing exterocolitos (NEC) in rats. METHODS:In glycine+NEC group, twenty anesthetized and artificially ventilated rats received 1g/kg glycine (20%, iv). Five minutes later, the rats were treated with 2 mg/kg lipopolysaccharide (LPS). In control group (NS+NEC), twenty rats were treated with normal saline as a substitute for glycine. In all animals, FiO₂ was reduced after 90 min from 21% to 5% and ventilation continued until 180 min or death. At the end of the experiment, the samples of blood and intestine were obtained immediately. Serum TNFα was measured with ELISA, serum NO was determined by nitrate reductase. The histopathology of the necrotic lesions were categoried: grade Ⅰ, focal mild injury confined to villous tips; grade Ⅱ, partial loss of villi; grade Ⅲ, necrosis extending to submucosa; grade Ⅳ, transmural necrosis. RESULTS:The survival time was shorter in the NS+NEC group (P＜0.01). The intestinal injury of the rats in glycine+NEC group was markedly alleviated (P＜0.01). The levels of TNF-α and NO₂^-/NO₃^- in serum decreased significantly in animals treated with glycine (P＜0.01, P＜0.05). CONCLUSION:Glycine alleviated LPS-induced NEC by inhibiting excessive production of TNFα and nitric oxide.     

Hyperhomocysteinemia induces endothelial dysfunction and aggravates microcirculation dysfunction and microthrombosis

AIM: To establish a microthrombus model by carrageenan (Ca)/ lipopolysaccharides (LPS) intraperitoneal injection in rats with hyperhomocysteinemia (HHcy) and endothelial dysfunction induced by L-methionine intake.METHODS: ① Male Sprague Dawley rats were randomly divided into 2 groups: control and endothelial dysfunction (HHcy) groups. L-methionine was administered by gavage in HHcy group for total 4 weeks.Purified water was administered by gavage in control rats.Plasma Hcy,NO and vWF were examined and the thoracic aorta were excised after 4 weeks of L-methionine treatment to evaluate endothelial function.② Male Sprague Dawley rats were randomly divided into 3 groups to establish a microthrombus formation model with Ca/ LPS: control,microthrombus formation (Ca/LPS) and endothelial dysfunction plus mitoarothrombus formation (HHcy＋Ca/LPS) groups.Control rats were injected with normal saline (NS).Ca/LPS rats were intraperitoneally injected with carrageenan (Ca) and followed by lipopolysaccharides (LPS) 16 h later.HHcy＋Ca/LPS rats were intragastric gavaged by L-methionine for total 4 weeks,and then were injected with Ca/LPS in the same way as Ca/LPS group.Cruor parameters and platelet count were detected at 20 h after LPS or NS injection and the mesentery microcirculation was monitored.Plasma NO and vWF were also detected at 24 h after LPS or NS injection.RESULTS: ① Plasma Hcy concentrations and vWF level were significantly increased in HHcy group,while plasma NO content was significantly decreased compared with that in control group.Endothelial dependent relaxation (EDR) of aortic rings was significantly decreased in HHcy group,suggesting endothelial damage/dysfunction was induced by HHcy.② Mesentery capillary was obviously blocked by microthrombus in Ca/LPS rats and was blocked more seriously in HHcy＋Ca/LPS rats.Cruor parameter results suggested that Ca/LPS rats were in hypercoagulable phase and HHcy＋Ca/LPS rats were in hypocoagulable phase at 20 h after LPS injection.Platelet count and plasma NO content in HHcy＋Ca/LPS group were significantly decreased,while plasma vWF level was significantly increased compared with Ca/LPS group.CONCLUSION: L-methionine intake induces severe HHcy and causes endothelial dysfunction in rats.Microcirculation dysfunction and microthrombosis can be caused by Ca/LPS intraperitoneal injection and may be aggravated by endothelial dysfunction.     

Effect of granulocyte-colony stimulating factor on the reendothelialization and intima hyperplasia of ballon-injured rat carotid artery

AIM:To study the effect of mobilization of stem cells by exogenous recombinant human granulocyte-colony stimulating factor (rhG-CSF) on the repairing process of reendothelialization and neointima hyperplasy on ballon injured rat carotid arteries.METHODS:Male Wistar rats were randomly divided into rhG-CSF group and NS+injury group.The animals were injected daily with 30 μg/kg rhG-CSF or 0.9% NaCl for 7 days,then underwent balloon angioplasty of the common carotid arteries which were harvested and processed for scanning electron microscopy (SEM),Evans blue staining,morphometric analysis of endothelialization and neointimal formation at 1 h,3 d,5 d,7 d,14 d after injury.Immunohistochemistry for proliferation cell nuclear antigen (PCNA) and RT-PCR for eNOS mRNA were also conducted for evaluating the proliferation of cells of the vessel wall and the possible mechanism of the repairing.RESULTS:SEM and Evan’s blue staining showed increased reendothelialization of the denuded vessels in rhG-CSF-treated animals compared with that NS+injury animals ［(60.6±7.3)% vs (41.6±3.3)%，P<0.01］.Neointima thickness was reduced by 37.3% in rhG-CSF group compared with NS+injury group 2 weeks after injury.Immunohistochemical staining for PCNA positive cells was less in rhG-CSF group compared with that in NS+injury group (42.6% vs 72.8%,P<0.01).CONCLUSION:rhG-CSF has beneficial effects on the reendothelialization and neointima thickness of the ballon-injured arteries.Mobilization of EPCs by exogenous granulocyte colony stimulating factor may be a potential therapeutic strategy for prevention of restenosis after percutenous coronary artery intervention.     

Changes of lymphatic vessel response to norepinephrine in hemorrhagic shock rats

AIM: To observe the changes of lymphatic vessel response to norepinephrine (NE) in hemorrhagic shock (HS) rats, and to explore the role of lymphatic reactivity in the pathogenesis of shock. METHODS: The lymphatic vessel pressure was observed through intubating into abdomen thoracic duct in 8 rats in sham group and HS group (which was bled from femoral artery until the mean arterial pressure to 40 mmHg). The changes of lymphatic vessel pressure response to NE at different time points were observed by injection of NE (5 μg/kg) through femoral vein. The spontaneous contraction frequency (F), maximal contraction diameter (a), maximal diastolic diameter (b) and static diameter (c) of mesenteric lymphatic (ML) living samples in 8 rats of each group were recorded through microcirculation video systems continuously. The changes of lymphatic fractional contraction index (index I), total contractile activity index (index II) and lymphatic dynamic index (LD-index) (to show the value using △F, △index I, △indexⅡ, △LD-index) were calculated after injection of NE at different time points. RESULTS: The changes of lymphatic boosting pressure response to NE in HS group was started to diminish 30 min after shock, and showed a progressive decreasing trend which significantly reduced than that in sham group at all time points of shock 1 h-3 h. In HS group, the △F, △indexⅡ, △LD-index at shock 1 h, the △F, △index I, △indexⅡ, △LD-index at shock 1.5 h and 2 h were significantly lower than those in sham group, and the △F, △index I, △indexⅡ, △LD-index at all time points were significantly decreased as compared to the values of pre-shock. CONCLUSION: Lymphatic vessel reactivity in shock rats is progressive declined in the process of hemorrhagic shock. The lymphatic vessel hypo-reactivity might play an important role in the pathogenesis of shock.     

Effect of hydroxylamine on pulmonary arterial hypertension induced by chronic hypoxic hypercapnia in rats

AIM: To explore the effects of hydroxylamine on the pulmonary arterial pressure in chronic hypoxic hypercapnic rats. METHODS: Twenty-four male Sprague-Dawley rats were randomly divided into 3 groups (8 rats in each group): the normal control group (NC), hypoxic hypercapnia+normal saline group (NS), hypoxic hypercapnia+hydroxylamine group (HA). The animals in NS and HA groups were kept in the O₂ (9%-11%) and CO₂ (5%-6%) cabin, 8 h a day and 6 days a week for 4 weeks. Before entering the cabin, the rats in HA group were administered with 1 mL hydroxylamine (12.5 mg/kg) by intraperitoneal injection, while the rats in NS group were given intraperitoneal injection of 1 mL saline solution. The mean pulmonary arterial pressure (mPAP) was measured by external jugular vein cannulation. The heart was removed, and the right ventricle (RV) and the left ventricle plus the septum (LV+S) were dissected. The ratio of the wet weight of the RV to that of the LV+S was calculated. The changes of the pulmonary vascular construction were observed under optical microscope. The concentration of H₂S in the plasma was measured with a spectrometer. The expression of cystathionine-γ-lyase (CSE) in the pulmonary arterioles and bronchi was measured by immunohistochemistry and RT-PCR. RESULTS: The values of mPAP, RV/(LV+S),vessel wall area/total area (WA/TA) and media thickness of pulmonary arterioles (PAMT) in NS group and HA group were significantly higher than those in NC group (P<0.05). The level of H₂S in the plasma, the content of CSE protein and the expression of CSE mRNA in NC group were significantly lower than those in NS group (P<0.05). The values of mPAP, RV/(LV+S), WA/TA and PAMT in HA group were significantly lower than those in NS group (P<0.05). The level of H₂S in the plasma, the content of CSE protein and the expression of CSE mRNA in HA group were significantly higher than those in NS group (P<0.05). CONCLUSION: Hydroxylamine may decrease the pulmonary arterial hypertension induced by chronic hypoxic hypercapnia in rats by increasing the level of H₂S in the plasma, the content of CSE protein and the mRNA expression of CSE, thus improving the pulmonary vascular structural remodeling.     

Effects of baicalin on glial fibrillary acidic protein and nuclear factor-κB expression in juvenile rat hippocampus after status convulsion

AIM: To study the effects of baicalin (BC) on glial fibrillary acidic protein (GFAP) and nuclear factor-κB (NF-κB) expression and neuronal apoptosis in juvenile rat hippocampus after status convulsion (SC). METHODS: One hundred and ninety five juvenile male Sprague-Dawley rats were randomly divided into 3 groups: normal saline pretreatment group (NS group), SC group and SC with BC pretreatment group (BC group). Each of these 3 groups would be subdivided into 5 subgroups sacrificed at 4 h, 12 h, 24 h, 48 h and 72 h after SC. The rat SC model was prepared by lithium-pilocarpine chemical method. The protein expression of GFAP and NF-κB was detected by the method of immunohistochemistry. The mRNA expression of GFAP was detected by RT-PCR. The neuronal apoptosis was observed by TdT-mediated dUTP nick end labeling (TUNEL). RESULTS: Compared with NS group, the GFAP positive cells was increased in SC group (P<0.05). Compared with SC group, the expression of GFAP was significantly reduced in BC group (P<0.05). Compared with NS group, the NF-κB positive cells was increased in SC group (P<0.05). Compared with SC group, the expression of NF-κB was significantly reduced in BC group. RT-PCR showed that the expression trend of GFAP mRNA was similar to that of the protein. Compared with NS group, the TUNEL positive cells in the hippocampus CA1 area in SC group increased significantly 12 h after SC (P<0.01), and reached a peak at 48 h. After the intervention with BC, the TUNEL positive cells decreased significantly between 12~48 h after SC (P<0.05 or P<0.01), but the number of TUNEL positive cells remained significantly greater than that in NS group (P<0.05). CONCLUSION: The expression of GFAP and NF-κB in the hippocampus increased after SC in rats. Baicalin decreases the expression of GFAP and NF-κB in hippocampus of rats with pilocarpine-induced seizures, and reduces the number of neuronal apoptosis, suggesting that baicalin may protect against the brain damage caused by status convulsion.     

Evaluation of the inflammatory response in two-hit acute lung injury model of rats using ［8F］ FDG microPET

AIM: To investigate whether two-hit acute lung injury (ALI) model is better than one-hit, and to evaluate the inflammatory response in the lungs during these models by using ［8F］FDG microPET. METHODS: Thirty three, adult, male Sprague-Dawley rats weighing 180-210 g were used and divided into 4 groups. Rats in LPS group (n=10) and LPS-HCl group (n=10) were challenged with intraperitoneal administration of LPS at the dose of 5 mg/kg, while rats in NS group (n=3) and HCl group (n=10) received normal saline solution intraperitoneally at the dose of 1 mL/kg, after 16 h, all animals were anesthetized with an intraperitoneal injection of sodium pentobarbital (40 mg/kg) and placed in a 60°inclined position, the femoral artery was cannulated and connected to a pressure transducer to record the arterial pressure on a polygraph recorder, the trachea was surgically exposed. Rats in HCl group and LPS-HCl group received direct intratracheal injection of HCl (pH=1.2) at the dose of 0.5 mL/kg while rats in NS group and LPS group received the same volume of normal saline solution. Blood gas samples (each 0.3 mL) were obtained at 30, 90 and 240 min after the instillation and replaced by the same volume of saline solution, the samples were analyzed using a blood gas analyzer. 240 min after HCl or NS administration, the rats underwent a microPET scanning, then, all the rats were sacrificed and the lungs were obtained for histological analysis. RESULTS: Blood gas analysis showed that rats in LPS-HCl group had higher PaO2 and lower PaCO2 than the other groups. MAP decreased markedly in LPS-HCl group, while MAP in other groups remained stable. The results of microPET showed that the ratio of ROI between the right lung and the muscle tissue of the right arm in LPS-HCl group was 9.00±1.41, and was significantly higher than that in LPS group (4.01±0.60) and HCl group (3.33±0.55). Histological examination showed that the mean lung injury score in LPS-HCl group was 12.70±0.95, while that was 8.40±1.26 in HCl group and 7.00±0.82 in LPS group, and there were significant differences (both P<0.01). CONCLUSION: LPS pretreatment significantly magnifies and prolongs the inflammatory response to the subsequent acid instillation in both lungs. When compared with “one-hit”, “two-hit” is easier to induce the ALI, and ［8F］FDG microPET is a useful tool to evaluate the inflammatory reaction during ALI.     

Role of myosin-light-chain kinase in biphasic contractile activity of lymphatics isolated from hemorrhagic shock rats

AIM: To elucidate the mechanism by which myosin-light-chain kinase (MLCK) modulates the biphasic contractile activity of lymphatics isolated from the rats subject to hemorrhagic shock (HS). METHODS: Male Wistar rats were randomiz to control group and HS group. In HS group, the rats were subject to HS and then further divided into HS 0 h, 0.5 h, 1 h, 2 h and 3 h subgroups. Thoracic ducts of control and shock rats were isolated and used to determine the protein levels of phosphorylated MLCK (p-MLCK). In addition, thoracic ducts obtained from control, 0.5 h- and 2 h-shocked rats were used to observe the contractile properties of lymphatics by a pressure myograph in vitro . Lymphatic rings were prepared and incubated with ML-7 (a specific inhibitor of MLCK) or substance P (SP, an agonist of MLCK). During the experiment, the contractile frequency (CF), end-diastolic diameter, end-systolic diameter and passive diameter in Ca²⁺-free PSS buffer were measured and used to calculate the lymphatic tonic index (TI), contractile amplitude (CA) and fractional pump flow (FPF) as the indexes of lymphatic contraction activity. RESULTS: The levels of p-MLCK in lymphatics in 0 h- and 0.5 h-shocked rats were significantly increased compared with the control rats, and it was gradually decreased with the development of shock. The values of CF, TI and FPF in 0.5 h-shocked lymphatics were significantly increased at transmural pressure of 1, 3 and 5 cmH₂O compared with those in control group, and significantly blunted by ML-7. SP obviously increased the suppressive effects induced by ML-7 and restored the values of CF, TI and FPF to the levels of HS 0.5 h group. CF, TI and FPF in 2 h-shocked lymphatics significantly declined under different transmural pressure as compared with those in control group, and significantly elevated by SP. Similarly, ML-7 depressed the effects of SP. No significant difference was found in CA between 0.5 h- and 2 h-shocked lymphatics. SP decreased the CA of lymphatics obtained from 2 h-shocked rats and this effect was suppressed by ML-7. However, both agents had no effects on CA of 0.5 h-shocked lymphatics. CONCLUSION: MLCK, as an essential enzyme that influences the contraction of lymphatic smooth muscle cells, involves in the modulation of biphasic changes of lymphatic contractile activity during the process of HS.