Fuhong Su

Evaluation of sublingual and gut mucosal microcirculation in sepsis: A quantitative analysis

Objective:To determine the relationship between sublingual and intestinal mucosal microcirculatory perfusion. Design:Observational, experimental study. Setting:University-affiliated large animal laboratory. Subjects:Ten fasted, anesthetized, mechanically ventilated, male pigs randomized to a sham group (n = 3) or to a hyperdynamic septic shock group (n = 7) in which cholangitis was induced by direct infusion of Escherichia coli into the common bile duct. This model was developed because it is not accompanied by changes in intra-abdominal pressure. Measurements and Main Results:The sublingual and intestinal microcirculations were simultaneously assessed at 4-hr intervals for up to 12 hrs with a modified orthogonal polarization spectral device and functional microvessel density and erythrocyte velocity were measured quantitatively. In sham animals, both regions maintained a stable functional microvessel density and erythrocyte velocity throughout the study period. In contrast, in septic animals, already after 4 hrs of sepsis, functional microvessel density was markedly decreased (>50%) in the sublingual and gut regions; mean erythrocyte velocity decreased dramatically and similarly in both regions, from 1022 ± 80 to 265 ± 43 &mgr;m/sec in the sublingual region and from 1068 ± 45 to 243 ± 115 &mgr;m/sec in the gut (p < 0.001, at T12). There was a significant correlation between the sublingual and gut microcirculations in septic animals (r2 = 0.92, p < 0.0001). Conclusions:The severity and the time course of microcirculatory changes were similar in the sublingual and in the gut region in this clinically relevant model of severe sepsis. These findings support the sublingual region as an appropriate region to monitor the microcirculation in sepsis.

Cerebral microcirculation is impaired during sepsis: an experimental study

IntroductionPathophysiology of brain dysfunction due to sepsis remains poorly understood. Cerebral microcirculatory alterations may play a role; however, experimental data are scarce. This study sought to investigate whether the cerebral microcirculation is altered in a clinically relevant animal model of septic shock.MethodsFifteen anesthetized, invasively monitored, and mechanically ventilated female sheep were allocated to a sham procedure (n = 5) or sepsis (n = 10), in which peritonitis was induced by intra-abdominal injection of autologous faeces. Animals were observed until spontaneous death or for a maximum of 20 hours. In addition to global hemodynamic assessment, the microcirculation of the cerebral cortex was evaluated using Sidestream Dark-Field (SDF) videomicroscopy at baseline, 6 hours, 12 hours and at shock onset. At least five images of 20 seconds each from separate areas were recorded at each time point and stored under a random number to be analyzed, using a semi-quantitative method, by an investigator blinded to time and condition.ResultsAll septic animals developed a hyperdynamic state associated with organ dysfunction and, ultimately, septic shock. In the septic animals, there was a progressive decrease in cerebral total perfused vessel density (from 5.9 ± 0.9 at baseline to 4.8 ± 0.7 n/mm at shock onset, P = 0.009), functional capillary density (from 2.8 ± 0.4 to 2.1 ± 0.7 n/mm, P = 0.049), the proportion of small perfused vessels (from 95 ± 3 to 85 ± 8%, P = 0.02), and the total number of perfused capillaries (from 22.7 ± 2.7 to 17.5 ± 5.2 n/mm, P = 0.04). There were no significant changes in microcirculatory flow index over time. In sham animals, the cerebral microcirculation was unaltered during the study period.ConclusionsIn this model of peritonitis, the cerebral microcirculation was impaired during sepsis, with a significant reduction in perfused small vessels at the onset of septic shock. These alterations may play a role in the pathogenesis of septic encephalopathy.

Fever control in septic shock: beneficial or harmful?

The beneficial effects of interventions to control fever in sepsis are controversial. We investigated whether the use of acetaminophen and external cooling is beneficial to control fever in septic shock. We studied 24 fasted, anesthetized, invasively monitored, mechanically ventilated female sheep (27.0 ± 4.6 kg) that received 0.5 g/kg body weight of feces into the abdominal cavity to induce sepsis. Ringers lactate (RL) was titrated to maintain pulmonary artery occlusion pressure (PAOP) at baseline levels throughout the experimental period. During the 2 h after the surgical operation, animals were placed in the hypothermia group if their temperature fell below 36.0°C; the other animals were randomized to three groups: high fever (T > 39.0°C), mild fever (37.5°C < T < 38.5°C), and normothermia (36.0°C < T < 37.0°C). The administration of 25 mg/kg acetaminophen every 4 to 6 h combined with external cooling (ice pad) was used to control core temperature in these three groups. The PaO2/FiO2 ratio was higher and blood lactate concentration was lower in the high fever than in the other groups (P < 0.01 and 0.05, respectively). Survival time was longer in the high fever group (25.2 ± 3.0 h) than in the mild fever (17.7 ± 3.5 h), normothermia (16.0 ± 1.9 h), and hypothermia (18.5 ± 2.5 h) groups (P < 0.05 for all). Plasma heat shock protein (HSP) 70 levels were higher in the two fever groups than in the other groups (P < 0.05). In this clinically relevant septic shock model, the febrile response thus resulted in better respiratory function, lower blood lactate concentration, and prolonged survival time. Antipyretic interventions including acetaminophen and external cooling were associated with lower circulating HSP70 levels. These data challenge the temperature control practices often used routinely in acutely ill patients.

Rapid alterations in transferrin sialylation during sepsis

The inflammatory process is associated with alterations in iron metabolism. Transferrin, an acute-phase N-glycosylated glycoprotein, plays an important role in iron transport. Human serum transferrin contains two biantennary glycans, each consisting of 0 to 4 molecules of sialic acid (SA); its SA content is heterogeneous with high concentration of tetrasialotransferrin (4SA) and low amounts of disialo-, trisialo-, penta-, and hexasialotransferrin. The hepatic uptake of iron is greater for desialylated transferrin isoforms (disialotransferrin) than for the other forms. We hypothesized that serum levels of carbohydrate-deficient transferrin (CDT, disialotransferrin) may increase rapidly in septic patients. Blood samples were obtained from critically ill patients with (n = 15) and without (n = 14) documented sepsis and compared with healthy volunteers. The different forms of transferrin were studied by capillary zone electrophoresis; SA concentrations were measured by enzymatic colorimetric assay. There was a significant increase in the proportion of CDT in septic compared with nonseptic patients and volunteers (18.3% [1.3-30.5] vs. 0.7% [0.5-0.9]; P < 0.01 and 0.9% [0.5-1.1]; P < 0.05). Conversely, tri- and tetrasialotransferrin levels were lower in septic patients. Total and free SA concentrations were significantly higher in septic patients than in healthy volunteers. In a sheep model of septic shock secondary to peritonitis, serum free SA was already increased after 15 h. Sepsis is associated with decreased SA content on circulating transferrin and with an increase in blood free SA concentrations. In view of these rapid modifications and the long half-life of transferrin, the most likely explanation is degradation of transferrin by neuraminidase. Further studies including measurement of blood neuraminidase concentration and activity are needed to understand the process and exact role of SA decrease in septic patients.

Fluid resuscitation in severe sepsis and septic shock: albumin, hydroxyethyl starch, gelatin or ringer's lactate-does it really make a difference?

The aim of this study was to investigate whether the type of i.v. fluid administered has an impact on outcome in an animal model of septic shock. The study included 28 anesthetized, invasively monitored, mechanically ventilated female sheep (29.5 ± 4.0 kg), which received 0.5 g/kg body weight of feces into the abdominal cavity to induce peritonitis. During the surgical operation and 4 h after feces spillage, only Ringers lactate (RL) was administered in all animals. Thereafter, animals were randomized to receive continuous infusions of RL (n = 7) alone or combined with either 20% albumin (n = 7, volume ratio to RL 1:10) or 6% hydroxyethyl starch (HES) (n = 7, volume ratio to RL 1:1), or gelatin alone (n= 7, no volume limitation). Fluid resuscitation was titrated to maintain pulmonary artery occlusion pressure at baseline levels throughout the experiment. No antibiotics or vasoactive drugs were administered, and animals were monitored until their spontaneous death. Hemodynamic variables were better with HES and albumin than with the other fluids, as reflected by higher stroke volume, cardiac index, and oxygen delivery (all P < 0.05). Hydroxyethyl-starch-treated animals also had lower arterial lactate concentrations (P < 0.01). However, times to develop hypotension and oliguria were similar in all groups. Blood interleukin (IL) 6 concentrations were significantly increased in all groups. The mean survival time was similar in all groups. In this clinically relevant model of prolonged septic shock, albumin and HES solution resulted in higher cardiac output, oxygen delivery, and lower blood lactate levels than gelatin and RL; however, the choice of i.v. fluid did not affect outcome.ABBREVIATIONS-RL-Ringers lactate; HES-Hydroxyethyl starch; PAOP-Pulmonary arterial occlusion pressure; IL-Interleukin; FiO2-Inspired oxygen fraction; PaO2-Arterial partial pressure of oxygen; PaCO2-Arterial partial pressure of carbon dioxide; MAP-Mean arterial pressure; CI-Cardiac index; DO2-Oxygen delivery; COP-Colloid osmotic pressure; ELISA-Enzyme-linked immunosorbent assay; PBS-Phosphate-buffered saline; HRP-Horseradish peroxidase; SD-Standard deviation; SE-Standard error; ANOVA-Analysis of variance

Sepsis is associated with altered cerebral microcirculation and tissue hypoxia in experimental peritonitis.

Objective:Alterations in cerebral microvascular blood flow may develop during sepsis, but the consequences of these abnormalities on tissue oxygenation and metabolism are not well defined. We studied the evolution of microvascular blood flow, brain oxygen tension (PbO2), and metabolism in a clinically relevant animal model of septic shock. Design:Prospective randomized animal study. Setting:University hospital research laboratory. Subjects:Fifteen invasively monitored and mechanically ventilated female sheep. Interventions:The sheep were randomized to fecal peritonitis (n = 10) or a sham procedure (n = 5), and craniectomies were performed to enable evaluation of cerebral microvascular blood flow, PbO2, and metabolism. The microvascular network of the left frontal cortex was evaluated (at baseline, 6, 12, and 18 hr) using sidestream dark-field videomicroscopy. Using an off-line semiquantitative method, functional capillary density and the proportion of small perfused vessels were calculated. PbO2 was measured hourly by a parenchymal Clark electrode, and cerebral metabolism was assessed by the lactate/pyruvate ratio using brain microdialysis; both these systems were placed in the right frontal cortex. Measurement and Main Results:In septic animals, cerebral functional capillary density (from 3.1 ± 0.5 to 1.9 ± 0.4 n/mm, p < 0.001) and proportion of small perfused vessels (from 98% ± 2% to 84% ± 7%, p = 0.004) decreased over the 18-hour study period. Concomitantly, PbO2 decreased (61 ± 5 to 41 ± 7 mm Hg, p < 0.001) and lactate/pyruvate ratio increased (23 ± 5 to 36 ± 19, p < 0.001). At 18 hours, when shock was present, animals with a mean arterial pressure less than 65 mm Hg (n = 6) had similar functional capillary density, proportion of small perfused vessels, and PbO2 values but significantly higher lactate/pyruvate ratio (46 ± 18 vs 20 ± 4, p = 0.009) compared with animals with an mean arterial pressure of 65–70 mm Hg (n = 4). Conclusions:Impaired cerebral microcirculation during sepsis is associated with progressive impairment in PbO2 and brain metabolism. Development of severe hypotension was responsible for a further increase in anaerobic metabolism. These alterations may play an important role in the pathogenesis of brain dysfunction during sepsis.

Administration of tetrahydrobiopterin improves the microcirculation and outcome in an ovine model of septic shock.

Objective: Supplementation with tetrahydrobiopterin, a nitric oxide synthase cofactor, may reduce microvascular endothelial dysfunction in severe sepsis. We studied whether tetrahydrobiopterin administration exerts beneficial effects in an ovine septic shock model. Design: Randomized animal study. Setting: University hospital animal research laboratory. Subjects: Fourteen adult female sheep. Interventions: Fecal peritonitis was induced, and the sheep were randomized to receive tetrahydrobiopterin (n = 7), given intravenously as 20mg/kg boluses at 4 and 12 hrs after sepsis induction, or placebo (n = 7). All animals were fluid resuscitated. The experiment was continued until death or for a maximum of 30 hrs. Measurements and Main Results: In addition to standard hemodynamic assessment, the sublingual microcirculation was evaluated using sidestream dark-field videomicroscopy. The first bolus of tetrahydrobiopterin blunted the increase in heart rate and cardiac index seen in the control group without affecting mean arterial pressure, and the second bolus of tetrahydrobiopterin prevented the decreases in cardiac index and mean arterial pressure. The reduction in mixed venous blood oxygen saturation and the increase in blood lactate seen in the control group were also delayed. Tetrahydrobiopterin significantly attenuated the deterioration in perfused small vessel proportion and density, microvascular flow index, and the increase in microvascular heterogeneity observed in the control group. Tetrahydrobiopterin was associated with better preserved lung compliance and PaO2/FIO2 ratio, which were associated with a lower lung wet/dry weight ratio at the end of the study. Median survival time was significantly prolonged in the tetrahydrobiopterin group (25.0 vs. 17.8 hrs, p < .01). Conclusion: In this clinically relevant model of sepsis, tetrahydrobiopterin supplementation attenuated the impairment in sublingual microvascular perfusion and permeability, which was accompanied by better preserved gas exchange, renal flow and urine output, and prolonged survival.

Beneficial effects of recombinant human activated protein C in a ewe model of septic shock.

Objective:To investigate the effects of activated protein C (APC) in a clinically relevant animal model of septic shock. Design:Prospective, randomized, controlled study. Setting:University medical center research laboratory. Subjects:Eighteen female sheep (body weight, 27–35 kg). Interventions:Animals were fasted, anesthetized, invasively monitored, and mechanically ventilated before receiving 0.5 g/kg body weight of feces intraperitoneally to induce sepsis. Fluid resuscitation with Ringer lactate was titrated to maintain pulmonary artery occlusion pressure at baseline levels. No vasoactive agents or antibiotics were used. Two hours after the induction of sepsis, animals were randomized to receive an infusion of APC (24 &mgr;g·kg−1·hr−1, n = 9) or an equivalent volume of vehicle (n = 9) throughout the experimental period. Measurements and Main Results:The APC-treated animals had significantly higher arterial pressure, urine output, Pao2/Fio2 ratios, and thoracopulmonary compliance than the control animals. They had lower pulmonary arterial pressure and arterial lactate concentrations than the control animals. Plasma colloid oncotic pressure was better maintained in the APC-treated group than in the control group (p < .05). Prothrombin time and activated partial thromboplastin time were altered less, and plasma D-dimer concentrations were significantly lower in the APC-treated group than in the control group (p < .05). The blood protein C concentration and platelet count were maintained better in the APC-treated group than in the control group (p < .05). APC administration was associated with significantly longer survival (median, 27 hrs vs. 20 hrs; p < .05). At postmortem examination, the lung wet/dry ratio was significantly lower in the APC group than in the control group (6.3 ± 0.7 vs. 7.1 ± 1.2, p < .05). Conclusions:In this clinically relevant model of septic shock due to fecal peritonitis, administration of APC had beneficial effects on hemodynamic variables, gas exchange, lactic acidosis, and coagulation abnormalities. Higher colloid oncotic pressures and lower lung wet/dry ratios at autopsy suggest preserved endothelial integrity. APC administration resulted in prolonged survival.

A selective V1A receptor agonist, selepressin, is superior to arginine vasopressin and to norepinephrine in ovine septic shock

Objective:Selective vasopressin V1A receptor agonists may have advantages over arginine vasopressin in the treatment of septic shock. We compared the effects of selepressin, a selective V1A receptor agonist, arginine vasopressin, and norepinephrine on hemodynamics, organ function, and survival in an ovine septic shock model. Design:Randomized animal study. Setting:University hospital animal research laboratory. Subjects:Forty-six adult female sheep. Interventions:Fecal peritonitis was induced in the anesthetized, mechanically ventilated, fluid-resuscitated sheep, and they were randomized in two successive phases. Three late-intervention groups (each n = 6) received IV selepressin (1 pmol/kg/min), arginine vasopressin (0.25 pmol [0.1 mU]/kg/min), or norepinephrine (3 nmol [0.5 &mgr;g]/kg/min) when mean arterial pressure remained less than 70 mm Hg despite fluid challenge; study drugs were thereafter titrated to keep mean arterial pressure at 70–80 mm Hg. Three early-intervention groups (each n = 7) received selepressin, arginine vasopressin, or norepinephrine at the same initial infusion rates as for the late intervention, but already when mean arterial pressure had decreased by 10% from baseline; doses were then titrated as for the late intervention. A control group (n = 7) received saline. All animals were observed until death or for a maximum of 30 hours. Measurements and Main Results:In addition to hemodynamic and organ function assessment, plasma interleukin-6 and nitrite/nitrate levels were measured. In the late-intervention groups, selepressin delayed the decrease in mean arterial pressure and was associated with lower lung wet/dry weight ratios than in the other two groups. In the early-intervention groups, selepressin maintained mean arterial pressure and cardiac index better than arginine vasopressin or norepinephrine, slowed the increase in blood lactate levels, and was associated with less lung edema, lower cumulative fluid balance, and lower interleukin-6 and nitrite/nitrate levels. Selepressin-treated animals survived longer than the other animals. Conclusions:In this clinically relevant model, selepressin, a selective V1A receptor agonist, was superior to arginine vasopressin and to norepinephrine in the treatment of septic shock, especially when administered early.

Cardiovascular and microvascular responses to mild hypothermia in an ovine model

AIMS Hypothermia is used for brain protection after resuscitation from cardiac arrest and other forms of brain injury, but its impact on systemic and tissue perfusion has not been well defined. The aim of this study was to evaluate the cardiovascular and microvascular responses to mild therapeutic hypothermia (MTH) in an ovine model. METHODS Seven anaesthetised, mechanically ventilated, invasively monitored sheep were cooled from a baseline temperature of 39-40°C to 34°C using cold intravenous fluids, ice packs and transnasal cooling. After 6h of MTH, sheep were progressively re-warmed to baseline temperature. Positive fluid balance was maintained during the entire study period to avoid hypovolemia. In addition to standard haemodynamic assessment, the sublingual microcirculation was evaluated using sidestream dark-field (SDF) videomicroscopy. RESULTS MTH was associated with significant decreases in cardiac index and left (LVSWI) and right (RVSWI) ventricular stroke work indexes. There was a downward shift in the relationship between LVSWI and pulmonary artery occlusion pressure during MTH, indicating myocardial depression. During MTH, mixed venous oxygen saturation increased, in association with reduced oxygen consumption, but blood lactate concentrations increased significantly. There was a significant decrease in the proportion and density of small perfused vessels. All variables returned to baseline levels during the re-warming phase. CONCLUSION In this large animal model, MTH was associated with decreased ventricular function, oxygen extraction and microvascular flow compared to normothermia. These changes were associated with increased blood lactate levels. These observations suggest that MTH may impair tissue oxygen delivery through maldistribution of capillary flow.