Cheng-Ming Tsao

Perivascular Adipose Tissue Inhibits Endothelial Function of Rat Aortas via Caveolin-1

Perivascular adipose tissue (PVAT)-derived factors have been proposed to play an important role in the pathogenesis of atherosclerosis. Caveolin-1 (Cav-1), occupying the calcium/calmodulin binding site of endothelial NO synthase (eNOS) and then inhibiting nitric oxide (NO) production, is also involved in the development of atherosclerosis. Thus, we investigated whether PVAT regulated vascular tone via Cav-1 and/or endothelial NO pathways. Isometric tension studies were carried out in isolated thoracic aortas from Wistar rats in the presence and absence of PVAT. Concentration-response curves of phenylephrine, acetylcholine, and sodium nitroprusside were illustrated to examine the vascular reactivity and endothelial function. The protein expressions of eNOS and Cav-1 were also examined in aortic homogenates. Our results demonstrated that PVAT significantly enhanced vasoconstriction and inhibited vasodilatation via endothelium-dependent mechanism. The aortic NO production was diminished after PVAT treatment, whereas protein expression and activity of eNOS were not significantly affected. In addition, Cav-1 protein expression was significantly increased in aortas with PVAT transfer. Furthermore, a caveolae depleter methyl-β-cyclodextrin abolished the effect of PVAT on the enhancement of vasoconstriction, and reversed the impairment of aortic NO production. In conclusion, unknown factor(s) released from PVAT may inhibit endothelial NO production and induce vasocontraction via an increase of Cav-1 protein expression.

RhoA/Rho-Kinase and Nitric Oxide in Vascular Reactivity in Rats with Endotoxaemia

RhoA/Rho-kinase (RhoA/ROK) pathway promotes vasoconstriction by calcium sensitivity mechanism. LPS causes nitric oxide (NO) overproduction to induce vascular hyporeactivity. Thus, we tried to examine the role of RhoA/ROK and NO in the regulation of vascular reactivity in different time-point of endotoxaemia. Male Wistar rats were intravenously infused for 10 min with saline or E. coli endotoxin (lipopolysaccharide, LPS, 10 mg/kg) and divided to five groups (n = 8 in each group): (i) Control, sacrificed at 6 h after saline infusion; (ii) LPS1h, sacrificed at 1 h after LPS infusion; (iii) LPS2h, sacrificed at 2 h after LPS infusion; (iv) LPS4h, sacrificed at 4 h after LPS infusion; and (v) LPS6h, sacrificed at 6 h after LPS infusion. LPS1h and LPS2h were regarded as early endotoxaemia, whereas LPS4h and LPS6h were regarded as late endotoxaemia. Indeed, our results showed that LPS reproduced a biphasic hypotension and sustained vascular hyporeactivity to noradrenaline (NA) in vivo. Interestingly, this hyporeactivity did not occur in ex vivo during early endotoxaemia. This could be due to increases of aortic RhoA activity (n = 5, P<0.05) and myosin phosphatase targeting subunit 1 phosphorylation (n = 3, P<0.05). In addition, pressor response to NA and vascular reactivity in early endotoxaemia were inhibited by ROK inhibitor, Y27632. Furthermore, plasma bradykinin was increased at 10 min (24.6±13.7 ng/mL, n = 5, P<0.05) and aortic endothelial NO synthase expression was increased at 1 h (+200%. n = 3, P<0.05) after LPS. In late endotoxaemia, the vascular hyporeactivity was associated with aortic inducible NO synthase expression (n = 3, P<0.05) and an increased serum NO level (n = 8, P<0.05). Thus, an increased RhoA activity could compensate vascular hyporeactivity in early endotoxaemia, and the large NO production inhibiting RhoA activity would lead to vascular hyporeactivity eventually.

Application of thrombelastography in liver injury induced by endotoxin in rat.

Liver injury developing in patients with sepsis may lead to an increased risk of mortality. Thrombelastography (TEG) is generally applied to evaluate hemostatic disturbance in patients undergoing liver transplantation or cardiopulmonary bypass. The aim of this study was to investigate the development of liver injury and coagulopathy in a lipopolysaccharide (LPS)-induced animal model and to assess the relationship between TEG variables and liver injury. Male Wistar rats received LPS (30 mg/kg over a 4-h intravenous infusion) to induce experimental liver injury or isotonic saline as a control. Variables of hemodynamics and liver biochemistry were measured during the subsequent 6 h after the start of infusion. TEG variables (R-time, K-time, &agr;-angle and maximal amplitude), thrombin–antithrombin complex and plasminogen activator inhibitor-1 were also measured. After LPS infusion, liver injury [examined by biochemical variables (e.g. alanine aminotransferase, ALT) and histological studies] was developed and inflammatory cytokines (tumor necrosis factor-&agr; and interleukin-6) were raised. At the initial period of LPS infusion, R-time was shortened and &agr;-angle was increased. Thereafter, &agr;-angle and maximal amplitude were decreased progressively, demonstrating that endotoxin induced coagulation disturbances. Furthermore, there were strong positive correlation between K-time and natural log (Ln)(ALT) (r = 0.823, P = 0.001); also, there were strong negative correlations between &agr;-angle and Ln(ALT) (r = −0.762, P = 0.002) as well as maximal amplitude and Ln(ALT) (r = −0.732, P = 0.004) at 6 h after LPS infusion. These results demonstrated that TEG could be a potential tool to evaluate the development of liver injury in endotoxemia.

Carbon dioxide embolism diagnosed by transesophageal echocardiography during endoscopic vein harvesting for coronary artery bypass grafting.

IMPLICATIONS We describe a case of massive carbon dioxide embolism with an abrupt decrease in arterial blood pressure and continuous mixed venous oxygen saturation during endoscopic vein harvesting that was immediately diagnosed by intraoperative transesophageal echocardiography.

Beneficial effects of hyperoncotic albumin on liver injury and survival in peritonitis-induced sepsis rats.

Liver injury/dysfunction developing in patients with sepsis may lead to an increased risk of death. Small-volume resuscitation with hyperoncotic albumin (HA) has been proposed to restore physiologic hemodynamics in hemorrhagic and septic shock. We evaluated whether HA resuscitation could alleviate the development of liver injury/dysfunction in rats with polymicrobial sepsis induced by cecal ligation and puncture (CLP). The male Wistar rats received 0.9% saline or HA (25%, 3 mL/kg intravenously) at 3 h after CLP or sham operation. All hemodynamic and biochemical variables were measured during the 18-h observation. After 18 h of CLP, the septic rats developed circulatory failure (i.e., hypotension, tachycardia, and poor tissue perfusion), liver injury (examined by biochemical variables and histologic studies), and a higher mortality. Hyperoncotic albumin not only ameliorated the deterioration of hemodynamic changes but also attenuated neutrophil infiltration and cell death in the liver of septic animals. The septic rats treated with HA had a higher survival when compared with those with 0.9% saline treatment. Moreover, the increased plasma IL-1&bgr;, plasma IL-6, plasma nitrite/nitrate concentrations, liver iNOS expression, and liver superoxide levels in CLP rats were attenuated after administration of HA. Thus, HA may be regarded as a potential therapeutic agent in the early treatment of septic shock to prevent or reduce subsequent liver failure.

Effects of small-volume hypertonic saline on acid-base and electrolytes balance in rats with peritonitis-induced sepsis.

ABSTRACT Our previous study has demonstrated that hypertonic saline (HS) given at 3 h after cecal ligation and puncture (CLP) surgery alleviates circulatory failure, multiple organ dysfunction syndrome, and mortality rate in rats. However, only few data exist on the application of HS in acid-base and electrolyte imbalance of sepsis. In addition, early one-dose HS administration seems to have only modest improvement on mortality rate. Thus, we evaluated the effects of HS on acid-base equilibrium and electrolyte balance in CLP-induced sepsis model and further compared with the effects of two- and one-dose HS administration. Male Wistar rats received CLP or sham operation followed by the administration of saline or HS (7.5% NaCl, 4 mL/kg, intravenously at 3 and 9 h after laparotomy or CLP). The changes in hemodynamics, biochemical variables, blood gas, electrolytes, organ histology, and plasma levels of nitric oxide (NO) and interleukin 1&bgr; (IL-1&bgr;) were examined during the 18-h observation. Hypertonic saline given either at 3 h (one-dose administration) or at 3 and 9 h (two-dose administration) after CLP attenuated circulatory failure, multiple organ dysfunction syndrome, metabolic acidosis, hyperkalemia, neutrophil infiltration, and 18-h mortality. Moreover, both one- and two-dose HS administrations significantly diminished plasma NO and IL-1&bgr; levels in CLP rats. However, only the two-dose HS administration significantly improved hyponatremia and hypocalcemia in septic rats. Beneficial effects of HS in septic rats may be attributed to not only reducing plasma levels of NO and IL-1&bgr;, but also improving metabolic acidosis and electrolyte imbalance. In addition, two-dose HS administration could reverse electrolyte imbalance caused by CLP.

Electro-acupuncture at 'Neiguan' (PC6) attenuates liver injury in endotoxaemic rats.

Background Intravenous injection of lipopolysaccharide (LPS) stimulates macrophages to release proinflammatory cytokines and nitric oxide (NO). This results in hypotension, vascular hyporeactivity and multiple organ failure (eg, liver injury) in rats. In rats with endotoxin shock, electro-acupuncture (EA) of ‘Neiguan’ (PC6) retrieved blood pressure and reduced plasma concentrations of NO. The authors evaluated whether EA at PC6 could alleviate the development of liver injury and dysfunction in endotoxic rats. Methods A total of 28 male adult Wistar rats were included in this study. Rats received intravenous LPS (10 mg/kg for 4 h) or saline for 4 h followed by EA at PC6 acupuncture point. Results Elevated biochemical parameters of liver injury and marked infiltration of neutrophils into liver tissues caused by LPS were significantly attenuated by EA. However, hypotension, tachycardia and raised production of plasma NO were not suppressed by EA at PC6. Conclusions These results indicate that EA at PC6 should be further investigated as a possible adjuvant therapy for endotoxin-induced liver dysfunction. Its mechanism of action needs further investigation.

Combined effects of propofol and dexamethasone on rats with endotoxemia.

Objective:To clarify the effect of combined treatment with propofol and dexamethasone on hemodynamics, organ injury, and survival rate in rats with endotoxemia. Design:Randomized, prospective animal experiment. Setting:Academic research laboratory. Subjects:Male Wistar rats. Interventions:Rats were divided into five groups: a control group, a group of conscious rats infused with Escherichia coli lipopolysaccharide, two groups of lipopolysaccharide rats treated with either propofol or dexamethasone, and a group of lipopolysaccharide rats with combined treatment of propofol and dexamethasone. Measurements and Main Results:All hemodynamic and biochemical variables were measured during the 6-hr observation. Propofol plus dexamethasone attenuated hypotension and delayed hypoglycemia and metabolic acidosis caused by coadministration of E. coli lipopolysaccharide. In addition, propofol plus dexamethasone attenuated the lipopolysaccharide-induced multiple organ dysfunctions, such as lung, liver, and kidney. The increases in serum tumor necrosis factor-&agr;, tissue nitric oxide, and superoxide anion levels were attenuated by propofol plus dexamethasone in lipopolysaccharide rats. Microscopic findings confirmed that propofol plus dexamethasone attenuated the substantial swelling and cell infiltration in lung and kidney caused by endotoxin. The 22-hr survival rate after endotoxin injection was markedly increased in lipopolysaccharide rats with combined treatment compared with the lipopolysaccharide rats (80% vs. 0%). Conclusions:The combined treatment with propofol plus dexamethasone reduced mortality rate and attenuated organ injury in conscious rats treated with lipopolysaccharide. These protective effects may be associated with their anti-inflammatory capacity and antioxidant activity.

Possible biomarkers of early mortality in peritonitis-induced sepsis rats

BACKGROUND Sepsis induced by cecal ligation and puncture (CLP) is accompanied by circulatory failure, multiple organ dysfunction syndrome, metabolic acidosis, and electrolyte imbalance in rats. However, it remains uncertain which parameters can be used to predict the mortality of septic rats. Thus, the aim of this study was to examine which possible biomarkers were associated with mortality in the CLP-induced sepsis model. MATERIALS AND METHODS After the carotid artery and vein were cannulated, rats were subsequently subjected to CLP or sham operation. The changes of hemodynamics, biochemical variables, blood gas, and electrolytes were monitored during the 18-h observation. RESULTS The CLP surgery caused circulatory failure, multiple organ dysfunction syndrome, metabolic acidosis, electrolyte imbalance, and death. Compared with survivors, nonsurvivors showed significant difference in (1) blood glucose; (2) lactate dehydrogenase, alanine aminotransferase, aspartate aminotransferase, creatinine, and blood urea nitrogen in serum; and (3) base excess, HCO3(-), PaCO2, potassium, and calcium in whole blood at 9 h after CLP. No significant difference in blood pressure, heart rate, pressor response to noradrenaline, rectal temperature, total protein, albumin, PaO2, and sodium was observed between nonsurvivors and survivors. However, after multifactor dimensionality reduction analysis, the union of HCO3(-) and blood glucose had the biggest testing balanced accuracy. CONCLUSIONS These results indicate that HCO3(-) plus blood glucose serves as the best biomarker of early death in rats with CLP-induced sepsis. Thus, these parameters could guide experimental procedures for making the right interventions when utilizing CLP as a sepsis model in rats.

Effect of propofol on vascular reactivity in thoracic aortas from rats with endotoxemia

Background: This study examined the effect of propofol on thoracic aortas isolated from endotoxic rats to assess endothelium‐dependent and ‐independent relaxant responses. Methods: Adult male Wistar rats were assigned randomly to one of two groups, a saline control group or an experimental group treated with lipopolysaccharide (LPS, 10 mg/kg intravenously). At 6 hours after saline or LPS infusion, the thoracic aorta was excised and cut into 3‐mm rings. Aortic rings with or without endothelium were suspended in organ baths for isometric tension recording. Results: Both norepinephrine (NE)‐induced vascular contraction and acetylcholine‐induced vasodilation were attenuated in aortasfrom LPS‐treated rats. Furthermore, preincubation with propofol caused a rightward shift in the NE concentration–response curve for aortasfrom LPS‐treated rats compared to sham controls. The slow and sustained, but not the initial fast, contractile response to NE was significantly suppressed by propofol in LPS‐treated aortas. In addition, vascular relaxation induced by propofol in LPS‐treated aortas was partially suppressed by inhibitors of either nitric oxide (NO) synthase or soluble guanylate cyclase, but not bypotassium channel inhibitors. Conclusion: These data suggest that propofol reduces the sensitivity to NE in aortic rings from endotoxic rats. This appears to be caused by (i) blockade of the extracellular calcium influx rather than a reduction in intracellular calcium release and (ii) an increased response to, at least in part, NO–cGMP in rat aortas.