Effect of elevated intra-abdominal pressure and hyperoxia on portal vein blood flow, hepatocyte proliferation and apoptosis in a rat model.

BACKGROUND/PURPOSE: Indications for a laparoscopic approach for the management of biliary atresia in children are not clearly defined. We have recently shown that persistent intra-abdominal pressure (IAP) significantly decreased portal vein (PV) flow. Ventilation with a high concentration of oxygen after abdomen deflation raises concerns of increased oxidative stress but has also been shown to exert beneficial effects on splanchnic ischemia/reperfusion. The purpose of the present study was to evaluate the effects of IAP and hyperoxia on liver histology, hepatocyte proliferation and apoptosis in a rat model of abdominal compartment syndrome (ACS). METHODS: Male Sprague-Dawley rats were anesthetized with intraperitoneal ketamine and xylasine. After a midline laparotomy, the PV was isolated. Ultrasonic blood flow probes were placed on the vessel for continuous measurement of regional blood flow. Mean arterial blood pressure (MABP) was continuously measured. Two large-caliber percutaneous peripheral intravenous catheters were introduced into the peritoneal cavity for inflation of air and measurement of IAP. Rats were divided into three experimental groups: 1) Sham rats were subjected to IAP of 0 mmHg; 2) ACS rats were subjected to IAP of 6 mmHg for 2 hours and were ventilated with air; and 3) ACS-O (2) rats were subjected to IAP of 6 mmHg for 2 hours and were ventilated with 100 % O (2) during the operation and ventilation was continued for 6 hours after operation. Liver structural changes, hepatocyte proliferation (using BrdU assay) and apoptosis (using Tunel assay) were determined 24 hours following operation. RESULTS: IAP at 6 mmHg caused a twofold decrease in PV flow compared to sham animals. Hyperoxia resulted in a less significant decrease in PV flow compared to air-ventilated animals. Despite a significant decrease in PV blood flow, 24 hours after abdominal deflation only a few animals demonstrated histological signs of liver damage. The small histological changes were accompanied by increased hepatocyte apoptosis and enhanced hepatocyte proliferation in 25 % of animals, suggesting a liver repair response. CONCLUSIONS: Despite a significant decrease in PV blood flow, persistent IAP for 2 hours results in few changes in liver histology, and stimulates hepatocyte proliferation and apoptosis in only a few animals, supporting the presence of a recovering mechanism. Treatment with hyperoxia did not significantly change hepatocyte proliferation and apoptosis.

Divergent effects of ischemia/reperfusion and nitric oxide donor on TNFalpha mRNA accumulation in rat organs.

We previously showed that serum TNFalpha bioactivity in rats is proportional to the extent of graded tissue injury caused by laparotomy, intestinal ischemia, and reperfusion and that the spleen is an important source of TNFalpha secretion in this condition. TNFalpha production varies, depending on the type and duration of tissue injury. It is also affected by other mediators, such as nitric oxide (NO). TNFalpha is known to increase NO production, but the effect of NO on the production of TNFalpha has not yet been fully elucidated. In this study we determined the levels of TNFalpha mRNA in rat organs after graded injury caused by anesthesia, laparotomy, intestinal ischemia, and reperfusion and evaluated the effects of the NO donor S-nitroso-N-acetylpenicillamine (SNAP) on it. Samples from different organs were removed, and TNFalpha gene expression was evaluated by semiquantitative RT-PCR. TNFalpha mRNA was not detected in the intestine (the ischemic organ) and in the kidney, brain, heart, or liver after all 4 experimental protocols. In the mesenteric lymph node (draining the ischemic organ) a basal level of expression of TNFalpha mRNA was detected in the control (anesthesia alone) group, which was increased significantly after ischemia. In the spleen (a remote immune organ not directly involved in the ischemia), a significant gradual increase in TNFalpha mRNA, which correlated to the severity of the experimental protocol, was observed. In the lung (a central participant in post-injury multiple organ failure), all interventions increased TNFalpha mRNA. Infusion of SNAP exerted a differential effect on TNFalpha mRNA: diminished its accumulation in the lymph node, enhanced it in the lung, and had no effect in the spleen. The divergent organ pattern of TNFalpha transcription emphasizes the importance of its localized expression, which is critical to the understanding of its autocrine and paracrine actions in ischemia and reperfusion.

Abdominal surgery reduces the ability of rat spleen cells to synthesize and secrete active tumour necrosis factor-alpha (TNF-alpha) by a multilevel regulation.

We have previously shown that abdominal surgery (explorative laparotomy) reduces the ability of lipopolysaccharide (LPS)-triggered spleen macrophages to secrete TNF-alpha. In this study we characterize possible mechanisms which could be responsible for the reduction in splenic production of TNF-alpha. Post-operative and control (unoperated) rat splenocytes or enriched splenic macrophages were cultured with LPS. Steady-state levels of TNF-alpha mRNA were determined by Northern and slot blot analyses, and validated by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). The amount of TNF-alpha protein was measured by Western blot analysis, and its biological activity was determined by the fibroblast L-929 cytotoxicity assay. Surgery induced a 12-fold inhibition in TNF-alpha activity (P < 0.02), caused up to two-fold reduction in the accumulation of TNF-alpha mRNA (P < 0.01), and suppressed TNF-alpha protein maturation into its 17-kD form in cellular extracts. Post-surgical spleen supernatants revealed mainly a band of a lower molecular weight (14 kD). Our data suggest a multilevel regulation of post-operative inhibition of TNF-alpha response to LPS, at the accumulation of mRNA, translational and secretory levels. We also suggest that the reduced bioactivity could be partially caused by a proteolytic cleavage of TNF-alpha. Since TNF-alpha is an important participant in immune responses, its reduced production and activity may be a central mechanism of post-operative immunosuppression.

[The form of the circulatory tree and the organ specificity of the blood supply]. / Forma krovenosnogo dereva i organospetsifichnost' krovosnabzheniia.

An original method of quantitative investigation of vascular bed is based on a vascular bed representation as a system of successively connected bifurcations. Parameters of arterial and venous bifurcations were measured in brain hemispheres and femoral biceps of Wistar rats, relationships between these values and stem diameter found, arterial-venous route numerically reconstructed, and its hemodynamic parameters (linear bloodflow velocity, parietal shift tension, hydraulic pressure, and vascular resistance) estimated. Structural functional differences in the vascular bed of the brain and muscle were detected and modeled.

Effects of severe hemorrhage on plasma ANP and glomerular ANP receptors.

Atrial natriuretic peptide (ANP) plays an important role in blood volume and electrolyte homeostasis in normovolemia and in hypervolemic states. The currently available information on the effects of hypovolemia on plasma ANP is contradictory. Moreover, possible regulation of ANP receptors during severe hemorrhagic hypovolemia has not been investigated. This study evaluated the effects of severe hemorrhage on plasma ANP and on the regulation of glomerular ANP receptor subtypes in anesthetized rats. Constant rate bleeding of 50% of total blood volume within 2 h induced a reproducible shock state characterized by marked decreases in blood pressure, heart rate, and hematocrit and an increase in plasma renin activity and aldosterone. Hemorrhaged rats exhibited a gradual significant increase in plasma ANP from 39.3 +/- 2.9 to 114.7 +/- 20.0 pmol/l 1 h after the bleeding (P < 0.001 from the initial value and P < 0.02 from the final value of sham-shock rats). Hemorrhage induced a significant decrease in total glomerular ANP binding sites (172 +/- 25 vs. 363 +/- 39 fmol/mg protein in hemorrhaged and sham-shock rats, respectively, P < 0.05). This decrease was mainly due to a significant decrease in ANPC receptors (132 +/- 22 vs. 312 +/- 40 fmol/mg protein in hemorrhaged and sham-shock rats, respectively, P < 0.05). Hemorrhage did not change glomerular ANPA receptor density. No significant differences in the affinity of the glomerular receptor subtypes for ANP were detected. Our data indicate that plasma ANP increases after prolonged severe hemorrhage. It is suggested that downregulation of renal ANPC receptors leads to reduced clearance of ANP and contributes to elevation of its plasma level after severe hemorrhage.

Effects of oxygen on regional hemodynamics in hemorrhagic shock.

This study investigated mechanisms of the hemodynamic effects of oxygen in hemorrhagic shock induced by bleeding 30% of the total blood volume in anesthetized rats. An ultrasonic flowmeter was used to monitor regional blood flow. Changes in tissue perfusion were assessed by the laser-Doppler technique. The inhalation of 100% oxygen induced a significant increase in mean arterial blood pressure (MABP) and vascular resistance in the hindquarters, with a concomitant decrease in blood flow in the distal aorta and biceps femoris muscle. In contrast, oxygen did not change vascular resistance in the superior mesenteric artery (SMA) and renal beds and induced a significant increase in blood flow to the renal artery, SMA, and small bowel in hemorrhaged rats. L-Arginine (100 mg/kg iv) but not D-arginine or the vehicle (0.9% NaCl) completely abolished the effects of oxygen on blood pressure and reversed its effects on blood flow and resistance in the hindquarters and biceps femoris muscle. Administration of the nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine methyl ester (50 mg/kg iv) significantly increased MABP and the resistance in the three vascular beds. Pretreatment of hemorrhaged rats with a superoxide dismutase mimic, the NO-stable radical 2,2,6,6-tetramethylpiperidine-N-oxyl (5 mg/kg iv), resulted in significantly diminished effects of oxygen on hindquarter hemodynamics. These results demonstrate a differential effect of oxygen, which increases vascular resistance in the hindquarters without a significant effect in the splanchnic and renal beds, thus favoring an increase in splanchnic and renal perfusion. It is suggested that inactivation of NO by reactive oxygen species may underlie the effects of oxygen on hindquarter vascular tone during shock.

[Age-related changes in the parameters of the capillary blood flow in rat skeletal muscle: allometric relations and the hypothesis of capillary functional length]. / Vozrastnye izmeneniia parametrov kapilliarnogo krovotoka v skeletnoi myshtse krysy: allometricheskie zavisimosti i gipoteza funktsional'noi dliny kapilliara.

The effect of growth on the capillary i.d. and density, the RBC velocity and tube hematocrit in the rat m. cremaster was studied in vivo. All these parameters decreased with growing as described by allometric equations. The mean capillary diameter and the hematocrit did not change significantly under the same conditions. A hypothetic explanation of the discrepancy between the bulk flow and capillary flow is suggested.

[The reactions of the capillary bed of the skeletal muscles to normobaric hyperoxia and a perfluoro-organic blood substitute]. / Reaktsii kapilliarnogo rusla skeletnoi myshtsy na normobaricheskuiu giperoksiiu i perftororganicheskii krovezamenitel'.

The capillary density of the m. cremaster decreased while the RBC velocity and tube hematocrit remained constant during oxygen respiration in rats. The blood substitute induced opposite changes. The data suggest that the RBC velocity decrease depended on the blood rheological properties due to addition of submicron particles of the blood substitute and not to its oxygen capacity.

[Structural and functional organization of the microcirculatory bed of the frog and rat mesentery]. / Strukturno-funktsional'naia organizatsiia mikrotsirkuliatornogo rusla bryzheiki liagushki i krysy.

The arteriolar and venular network and the blood flow distribution were studied in mesenteric microvascular beds of frogs and rats. Some principles of spatial organization were established: an increase of branching coefficient, a decrease of branching asymmetry with the internal diameter of parent vessel progressively diminishing, the more frequent branching of venous tree including the fragment of a relatively greater volume. The values of geometrical and hemodynamical parameters were different in frogs and rats. Experimental dependence between pressure gradients and vessel radii was found to be exponential, thus being in agreement with prediction of a mathematical model based on physiologically significant criteria.

[Cardiac minute volume and its pattern in Rana ridibunda frogs]. / Minutnyi ob"em serdtsa i ego raspredelenie u liagushek Rana ridibunda.

In frogs with an average body mass 56 g, the minute volume of the heart is equal to 4.5 ml/min X 100 g, which is approximately an order lower than in mammals with the same body mass. Pulmonary fraction constitutes 52% of the minute volume of the heart. The main bulk of systemic fraction of the minute volume of the heart (78%) passes to locomotor system and skin, whereas 19% of this volume are adressed to vegetative organs. This pattern of distribution significantly differs from that in mammals with a similar body mass, in which the vegetative and locomotor fractions are approximately equal. Differentiation in muscular blood supply was noted--there is a threefold difference in the volume of blood flow between gastrocnemius and submandibular muscles.

[Architectonics of the arterial bed of the cerebral hemispheres in the normal rat and following a stay at an "altitude" of 5600 m]. / Arkhitektonika arterial'nogo rusla polusharii golovnogo mozga u krys v norme i posle prebyvaniia na "vysote" 5600 m.

Post mortem morphometric study of the arterial bed of the brain hemispheres in rats after 90-day stay at "the altitude" 5600 m revealed a noticeably larger diameter of arteries (D), a greater coefficient of their branching, lesser distances between branches (L), larger angles of branch deviation and a longer capillary bed. Calculations based on these data show that amount of arterial vessels and capillaries must increase in hypoxia, their length and linear blood flow velocity (V) must diminish, and the value of V/D2 and epsilon (LV/D2) characterizing the gradient of impetuous pressure and total losses of the impetuosity in the vascular bed must decrease.