Hypothermia improves oral and gastric mucosal oxygenation during hypoxic challenges.

BACKGROUND: Therapeutic hypothermia, used primarily for protective effects after hypoxia, improves oral and gastric mucosal microvascular oxygenation (µHbO2) during additional haemorrhage. Therefore, we questioned whether hypothermia likewise improves µHbO2 during hypoxic challenges. Since both hypothermia and hypoxia reduce cardiac output (e.g. by myofilament Ca(2+) desensitization), and modulate vasomotor tone via K(+) ATP channels, we hypothesized that the Ca(2+) sensitizer levosimendan and K(+) ATP channel blocker glibenclamide would support the cardiovascular system. METHODS: The effects of mild hypothermia (34°C) on µHbO2 during hypoxia [Formula: see text] were analysed in a cross-over study on five anaesthetized dogs and compared with normothermia (37.5°C) and hypoxia. During hypothermia, but before hypoxia, glibenclamide (0.2 mg kg(-1)) or levosimendan (20 µg kg(-1)+0.25 µg kg(-1) min(-1)) was administered. Systemic haemodynamic variables, gastric and oral mucosal microvascular oxygenation (reflectance spectrophotometry), and perfusion (laser Doppler flowmetry) were recorded continuously. Data are presented as mean (sem), P<0.05. RESULTS: Hypoxia during normothermia reduced gastric µHbO2 by 27 (3)% and oral µHbO2 by 28 (3)% (absolute change). During hypothermia, this reduction was attenuated to 16 (3)% and 13 (1)% (absolute change). This effect was independent of microvascular flow that did not change during hypoxia and hypothermia. Additional administration of levosimendan during hypothermia restored reduced cardiac output but did not change flow or µHbO2 compared with hypothermia alone. Glibenclamide did not exert any additional effects during hypothermia. CONCLUSIONS: Hypothermia attenuates the decrease in µHbO2 during additional hypoxic challenges independent of systemic or regional flow changes. A reduction in cardiac output during hypothermia is prevented by Ca(2+) sensitization with levosimendan but not by K(+) ATP channel blockade with glibenclamide.

In-vitro antidiabetic activity of a Bistorta officinalis Delarbre root extract can not be confirmed in the in-vivo models hen's egg test and Drosophila melanogaster.

The potential of plant bioactives for the prevention and therapy of diabetes is increasingly being recognized. In the present study we investigated the antidiabetic properties of an aqueous Bistorta officinalis Delarbre extract (BODE) by employing both in-vitro assays and in-vivo models. Multiple targets in glucose homeostasis which are involved in the regulation of the blood glucose level were affected by BODE in-vitro. The extract exhibited inhibitory activities towards the intestinal carbohydrate-hydrolysing enzymes α-amylase and α-glucosidase with IC50 values of 81.5 µg/mL and 8.4 µg/mL, respectively. Furthermore, moderate reduction of the dipeptidyl peptidase-4 (DPP4) enzyme activity was evident when tested in the presence of 1.0 mg/mL BODE. A significant inhibition of the intestinal glucose transporter sodium-dependent glucose transporter 1 (SGLT1) in response to 1.0 mg/mL BODE was shown for Caco-2 cells mounted in Ussing chambers. High performance liquid chromatography-mass spectrometry analyses of the BODE revealed several plant bioactives including gallotannins, catechins and chlorogenic acid. Although our in-vitro data were promising, BODE-supplementation in the model organism Drosophila melanogaster lacked to confirm the antidiabetic effect of the extract in-vivo. Moreover, BODE failed to reduce blood glucose levels in chicken embryos (in-ovo). Hence, BODE is probably not a suitable candidate for developing a pharmaceutical against diabetes mellitus.

Cardiovascular stability and unchanged muscle sympathetic activity during xenon anaesthesia: role of norepinephrine uptake inhibition.

BACKGROUND: Intraoperative hypotension is associated with increased risk of perioperative complications. The N-methyl-d-aspartate (NMDA) receptor (NMDA-R) antagonist xenon (Xe) induces general anaesthesia without impairment of cardiac output and vascular resistance. Mechanisms involved in cardiovascular stability have not been identified. METHODS: Muscle sympathetic activity (MSA) (microneurography), sympathetic baroreflex gain, norepinephrine (NE) plasma concentration (high-performance liquid chromatography), anaesthetic depth (Narcotrend(®) EEG monitoring), and vital parameters were analysed in vivo during Xe mono-anaesthesia in human volunteers (n=8). In vitro, NE transporter (NET) expressing HEK293 cells and SH-SY5Y neuroblastoma cells were pre-treated with ketamine, MK-801, NMDA/glycine, or vehicle. Subsequently, cells were incubated with or without Xe (65%). NE uptake was measured by using a fluorescent NET substrate (n=4) or [(3)H]NE (n=6). RESULTS: In vivo, Xe anaesthesia increased mean (standard deviation) arterial pressure from 93 (4) to 107 (6) mm Hg and NE plasma concentration from 156 (55) to 292 (106) pg ml(-1), P<0.01. MSA and baroreflex gain were unaltered. In vitro, ketamine decreased NET activity (P<0.01) in NET-expressing HEK293 cells, while Xe, MK-801, and NMDA/glycine did not. Xe reduced uptake in SH-SY5Y cells expressing NET and NMDA-Rs (P<0.01). MK-801 (P<0.01) and ketamine (P<0.01) also reduced NET activity, but NMDA/glycine blocked the effect of Xe on [(3)H]NE uptake. CONCLUSIONS: In vivo, Xe anaesthesia does not alter sympathetic activity and baroreflex gain, despite increased mean arterial pressure. In vitro, Xe decreases the uptake of NE in neuronal cells by the inhibition of NET. This inhibition might be related to NMDA-R antagonism and explain increased NE concentrations at the synaptic cleft and in plasma, contributing to cardiovascular stability during Xe anaesthesia.

Ketamine induces apoptosis via the mitochondrial pathway in human lymphocytes and neuronal cells.

BACKGROUND: Ketamine has been shown to have neurotoxic properties, when administered neuraxially. The mechanism of this local toxicity is still unknown. Therefore, we investigated the mechanism of cytotoxicity in different human cell lines in vitro. METHODS: We incubated the following cell types for 24 h with increasing concentrations of S(+)-ketamine and racemic ketamine: (i) human Jurkat T-lymphoma cells overexpressing the antiapoptotic B-cell lymphoma 2 protein, (ii) cells deficient of caspase-9, caspase-8, or Fas-associated protein with death domain and parental cells, and (iii) neuroblastoma cells (SHEP). N-Methyl-d-aspartate (NMDA) receptors and caspase-3 cleavage were identified by immunoblotting. Cell viability and apoptotic cell death were evaluated flowcytometrically by Annexin V and 7-aminoactinomycin D double staining. Mitochondrial metabolic activity and caspase-3 activation were measured. RESULTS: Ketamine, in a concentration-dependent manner, induced apoptosis in lymphocytes and neuroblastoma cell lines. Cell lines with alterations of the mitochondrial pathway of apoptosis were protected against ketamine-induced apoptosis, whereas alterations of the death receptor pathway did not reduce apoptosis. S(+)-Ketamine and racemic ketamine induced the same percentage of cell death in Jurkat cells, whereas in neuroblastoma cells, S(+)-ketamine was slightly less toxic. CONCLUSIONS: Ketamine at millimolar concentrations induces apoptosis via the mitochondrial pathway, independent of death receptor signalling. At higher concentrations necrosis is the predominant mechanism. Less toxicity of S(+)-ketamine was observed in neuroblastoma cells, but this difference was minor and therefore unlikely to be mediated via the NMDA receptor.

Disease-discordant twin structural MRI studies on affective disorders.

Magnetic resonance imaging (MRI) studies have identified neural structures implicated in the pathophysiology of mood disorders, especially bipolar disorder (BD) and major depressive disorder (MDD). However, the role of genetic and environmental influences on such brain deficits is still unclear. In this context, the present review summarizes the current evidence from structural MRI and Diffusion Tensor Imaging (DTI) studies on twin samples concordant or discordant for BD or MDD, with the aim of clarifying the role of genetic and environmental risk factors on brain alterations. Although the results showed a complex interplay between gene and environment in affective disorders, the evidence seem to underline that both genetic and environmental risk factors have an impact on brain areas and vulnerability to MDD and BD. However, the precise mechanism of action and the interaction between these factors still needs to be unveiled. Therefore, future larger studies on concordant or discordant twins should be encouraged, because this population provides a unique opportunity to probe separately genetic and environmental markers of disease vulnerability.

Effects of haemofiltration and mannitol treatment on cardiopulmonary-bypass induced immunosuppression.

Cardiac surgery using cardiopulmonary bypass (CPB) causes a systemic inflammatory response. Additionally, an impairment of the responsiveness of peripheral blood mononuclear cells (PBMC) to further immunological stimuli has been observed. The aim of our present study was to evaluate the ability of antioxidant therapy with mannitol or haemofiltration during CPB to modulate this immunosuppression after CPB. Forty-five patients undergoing elective heart-surgery were prospectively enrolled and randomized into three groups (control, mannitol, haemofiltration). Blood samples were taken after induction of anaesthesia (T1), 20 min after CPB (T2) and 24 h post-operatively (T3). Expression density of the monocytic surface receptor CD14, HLA-DR expression and cytokine release (TNF-alpha and IL10) after lipopolysaccharide-stimulation were evaluated. At T2, the CD14(dim) cell population was maintained in both intervention groups while in the control group there was a decrease of this proinflammatory monocytic phenotype. No significant differences regarding HLA-DR expression or cytokine release could be demonstrated. This study shows that the suppression of the stimulated immune response after CPB can potentially be alleviated by mannitol or haemofiltration in an experimental in-vitro setting. In the light of data showing that this depression of the immune response might affect the post-operative course of patients, these results could have a potential clinical relevance.

Internalization via Antennapedia protein transduction domain of an scFv antibody toward c-Myc protein.

We constructed a single-chain variable fragment miniantibody (G11-scFv) directed toward the transactivation domain of c-Myc, which is fused with the internalization domain Int of Antennapedia at its carboxyl terminus (a cargo-carrier construct). In ELISA experiments, an EC(50) for binding saturation was achieved at concentrations of G11-scFv-Int(-) of approximately 10(-8) M. Internalization of a fluoresceinated Fl-G11-scFv-Int(+) construct was observed in intact human cultured cells with confocal microscopy. After 5 h of incubation in medium containing 1 microM Fl-G11-scFv-Int(+) or Fl-G11-scFv-Int(-), fluorescence intensity was determined in individual cells, both for cytoplasmic and nuclear compartments: concentration levels of Fl-G11-scFv-Int(+), relative to the extracellular culture medium concentration, were 4-5 times higher in the cytoplasm, 7-8 times higher in the nucleus, and 10 times higher in the nucleoli. In the same experimental conditions, the Fl-G11-scFv-Int(-) construct was 3-4 times more concentrated outside of the cells than inside. Cell membranes kept their integrity after 5 h of incubation. The antiproliferative activity of our miniantibody was studied on HCT116 cells. Incubation with 4 microM G11-scFv-Int(+) for 4 days induced very significant statistical and biological growth inhibition, whereas Int alone was completely inactive. Miniantibodies capable of penetrating cell membranes dramatically broaden the potential for innovative therapeutic agents and attack of new targets.

Apoptosis induction by different local anaesthetics in a neuroblastoma cell line.

BACKGROUND: Local anaesthetics are known to induce apoptosis in clinically relevant concentrations. Hitherto, it is unknown what determines the apoptotic potency of local anaesthetics. Therefore, we compared apoptosis induction by local anaesthetics related to their physicochemical properties in human neuronal cells. METHODS: Neuroblastoma cells (SHEP) were incubated with eight local anaesthetics, two of the ester and six of the amide types. At least, five concentrations of each local anaesthetic were evaluated. After incubation for 24 h, rates of cells in early apoptotic stages and overall cell death were evaluated by annexin V and 7-amino-actinomycin D double staining by flow cytometry. The concentrations that led to half-maximal neurotoxic effects (LD50) were calculated and compared for all local anaesthetics. RESULTS: All local anaesthetics were neurotoxic in a concentration-dependent manner. All drugs induced similar rates of early apoptotic cell formation at low concentrations, whereas at high concentrations, late apoptotic or necrotic cell death predominated. Comparison of LD50 values of the different local anaesthetics resulted in the following order of apoptotic potency from high to low toxicity: tetracaine>bupivacaine>prilocaine=mepivacaine=ropivacaine>lidocaine>procaine=articaine. The toxicity correlated with octanol/buffer coefficients and also with experimental potency of the local anaesthetic, but was unrelated to the structure (ester or amide type). CONCLUSIONS: All commonly used local anaesthetics induce neuronal apoptosis in clinically used concentrations. The neurotoxicity correlates with lipid solubility and thus with the conduction blocking potency of the local anaesthetic, but is independent of the chemical class (ester/amide).

Inhibition of hemoxygenase-1 improves survival after liver resection in jaundiced rats.

BACKGROUND: The role of hemoxygenase (HO)-1 after partial liver resection (PLR) in jaundiced animals has yet to be defined. We therefore investigated: (1) the acute effects of bile duct ligation (BDL) on bilirubin accumulation and hepatocellular integrity after PLR; (2) how BDL and PLR affect HO-1 protein expression; (3) how functional HO-1 blockade affects survival and liver regeneration after BDL and PLR. METHODS: Male Sprague-Dawley rats were subjected to BDL or a sham operation. After 3 days, a 70% hepatectomy was performed. In a second set of experiments, BDL animals received either Sn(IV) mesoporphyrin IX dichloride (SnMP) for HO-1 blockade or a vehicle. Three days later, PLR was performed and survival of the animals was observed for 7 days. RESULTS: PLR, BDL and both together cause a hepatocellular injury and HO-1 expression. Inhibition of HO-1 with SnMP in jaundiced rats that underwent PLR was associated with improved survival, attenuated postoperative weight loss and better liver synthesis. CONCLUSION: The present findings add further evidence that the protective properties of increased HO-1 expression largely depend on the model used, and that HO-1 overexpression in the model of liver resection during acute cholestasis may also be detrimental.

A new unbalanced chromosomal abnormality in 1q31.1 to 1q32 without phenotypic consequences.

A familial duplication in the long arm of one chromosome 1 was detected due to recurrent abortions in a couple. The duplication was present in the male partner of the couple and in his mother, both clinically healthy. By reverse FISH, the duplication was determined to be located in 1q31. Multicolor banding (MCB) and application of locus-specific probes narrowed down the breakpoints to 1q31.1 and 1q32. The duplication spans a region of 13.9 Mb. None of the two breakpoints was colocalized with a known fragile site in 1q31.2, which, however, was duplicated. To the best of our knowledge, this is the first report of an unbalanced chromosome abnormality in this region that is not correlated with any clinical consequences.

Structure and spectroscopic characteristics of 4-tert-butylphenoxyacetylhydrazones of arylaldehydes.

By reaction of 4-tert-butylphenoxyacetylhydrazide with aromatic aldehydes and acetone a new series of 4-tert-butylphenoxyacetylhydrazones was synthesized. The structural peculiarities of the investigated molecules have been determined by means of X-ray analysis and IR spectroscopy. The diagnostically important IR spectral criteria required for the conformational analysis of acethylhydrazones have been considered. It was established that the 4-tert-butylphenoxyacetylhydrazide in condensed phase exists only as a ZN-C(O)-conformer. Its derivatives exist as EN-C(O) and ZN-C(O)-forms. As a rule, the prevalence of ZN-C(O)-form has been observed in CCl4 solutions. The structure of investigated compounds is also determined by a system of inter- and intramolecular hydrogen bonds. The energy of hydrogen bonds was estimated.

[Influence of gender on stimulated cytokine response in patients with severe sepsis]. / Einfluss des Geschlechts auf die stimulierbare Zytokinantwort bei Patienten mit schwerer Sepsis.

AIM: Studies suggest that female mice have lower mortality rates than males after sepsis or trauma-hemorrhage. This study investigated the impact of gender and disease severity on monocyte hyporesponsiveness in severe human sepsis. METHODS: We prospectively investigated 49 (male n=28, female n=21) consecutive patients with severe sepsis. Tumor necrosis factor-alpha (TNF-alpha) and interleukin-10 (IL-10) were assayed by ELISA in unstimulated whole blood cultures or after stimulation with lipopolysaccharide (LPS; E. coli 0111:B4) or Staph. aureus Cowan strain I (SAC-I) lysate at days 1, 2, 3, 4, and 8 after enrollment. Testosterone and estradiol levels were quantified by electrochemoluminescence immunoassays. RESULTS: Mortality was similar for males (35.7%) and females (42.9%). While disease severity was also comparable, septic patients showed a substantial suppression in stimulated TNF-alpha response compared to healthy controls who recovered within 8 days in surviving patients. Stimulated cytokine response recovered in female non-surviving patients, while it remained suppressed in non-surviving male patients and was significantly different compared to female non-surviving patients. Testosterone levels were substantially suppressed in male but not female septic patients compared to normal values but did not differ between surviving and non-surviving patients. Estradiol levels were elevated in female and male septic patients. Addition of different concentrations of testosterone and estradiol to whole blood obtained from younger (<35 years old) and older (>60 years old) male as well as from younger (proestrous premenopausal) and older (postmenopausal) female non-septic volunteers revealed no effect on LPS-stimulated TNF-alpha and IL-10 release. CONCLUSION: Severe sepsis leads to a substantial suppression of stimulated cytokine response. Prolonged suppression may serve as a marker of unfavourable outcome in male but not in female individuals suffering from severe sepsis. Furthermore, our data suggest that gender differences in cellular immunity described for young, sexually mature animals obviously persist in typical postmenopausal intensive care unit patients, although a direct interaction between testosterone or estradiol and LPS-stimulated cytokine response could not be demonstrated.

[Elimination of anatomical and esthetic lesions in patients with rough infected defects of tissue].

The reconstructive plastic operations were made in 217 patients with infected tissue defects with continuous regional infusions or with a simultaneous sanitizing operation, or within 7-15 days. Skin plasty, catheterization of the main artery (210), secondary surgical treatment and sequest-necrectomy (185), osteosynthesis (106), bone plasty (69) and others (1118 operations in all) were carried on in all the patients, i.e. 5.1 operations per each patient with artery catheterization taken into account. The method of plastic replacement was chosen individually depending on localization, depth and spread of the defect. Good and satisfactory results were obtained in all the patients with no esthetic impairments.

5-Fluorouracil/irinotecan induced lethal toxicity as a result of a combined pharmacogenetic syndrome: report of a case.

Combination cancer chemotherapy induced toxicity can be associated with combined pharmacogenetic syndromes. Dihydropyrimidine dehydrogenase (DPD) is the principal enzyme involved in the catabolic detoxification of 5-fluorouracil (5FU). A heterozygous G > A transition at the 5' splicing donor consensus sequence in intron 14 leading to exon 14 skipping (IVS14+1 G > A, DPYD*2A) with partial loss of enzyme activity may be partly responsible for 5FU induced toxicity, whereas irinotecan associated toxicity may in part be explained by an aberrant UGT1A1 promoter (TA)(n) genotype underlying Gilbert's syndrome with reduced liver glucuronidation activity. This report describes a 44 year old white woman who suffered from severe gastrointestinal and haematological toxicity while undergoing 5FU(24h)/folinic acid/irinotecan treatment for adenocarcinoma of the sigmoid colon. Despite appropriate supportive treatment, her condition rapidly deteriorated and led to death. Molecular analysis revealed a hitherto undescribed combined pharmacogenetic syndrome, consisting of heterozygosity for the DPD IVS14+1 G > A mutation and UGT1A1 (TA)(6/7) heterozygosity, which probably contributed to the fatal outcome in this patient.

Changes of salivary estrogen levels for detecting the fertile period.

OBJECTIVES: Is the saliva test, Geratherm ovu control, as accurate as the established urinary luteinizing hormone (LH) test for detecting ovulation and the following the fertile period? STUDY DESIGN: The voluntary participants were 74 healthy women with regular menstrual cycles and not using any hormonal contraceptives. The women used Geratherm ovu control, a small plastic hand-held microscope, for detecting the fertile period. A drop of saliva from sublingual was put onto the lens of the microscope. Three results were possible: non-fertile (dot pattern), transitional and fertile (ferning pattern). The participants performed the saliva test from the 5th till the 22nd day of the menstrual cycle and noted the respective result in a table. In addition to Geratherm ovu control, the EXACTO test for determining urinary LH concentration and the time of peak fertility was also performed. RESULTS: Positive LH shows a sharp increase beginning on the 10th cycle day with a maximum on the 17th cycle day. The curve for positive saliva and questionable positive saliva (one curve) is almost parallel with the curve for positive LH, reaching a maximum on the 16th cycle day. There is a high level of conformity for the same test results from the 5th (100%) till the 14th (84%) cycle day and from the 18th (80%) till the 22nd (96%) cycle day which corresponds to the pre- and post-ovulatory period. CONCLUSION: The saliva and the LH test both detect the fertile window of a menstrual cycle. Caused by the different hormones (estrogen for the saliva and LH for the LH test) leading to the respective positive test results, saliva turns positive 24h before LH. Consequently, the saliva test can be used as an ovulation test and help women maximize their chances of conceiving. There is also a high congruence between LH and saliva in the pre- and post-ovulatory period, indicating that the saliva test can also be used for contraception purposes.

Severity of polymicrobial sepsis modulates mitochondrial function in rat liver.

Mitochondrial dysfunction is assumed to be an important contributor to multi organ dysfunction syndrome. Here, the effects of varying degrees of sepsis on hepatic mitochondrial function were investigated. Moderate or more severe sepsis was induced in rats using a colon ascendens stent peritonitis (CASP)-model (16 G and 14 G stent respectively). Respiratory control ratio (RCR) was significantly higher in the 16 G-group and unchanged in the 14 G-group compared with healthy controls. The ADP/O ratio was similar in all groups. Our results indicate that different severities of sepsis differently influence the mitochondrial function, which could be a sign of adaptive reaction.

Caught in the trio trap? Potential selection bias inherent to association studies using parent-offspring trios.

During the last years, the validity of classic case control studies in psychiatric genetic research has been increasingly under question due to the risk of population stratification problems inherent to this type of association study. By consequence, the application of family-based association studies using parent-offspring trios has been strongly advocated. Recently, however, in a study comparing clinical characteristics between index patients from parent-offspring trios and singleton patients with bipolar affective disorder, the question was raised whether a systematic neglect of case control association studies could lead to a selection bias of susceptibility genes. In a similar approach, we compared demographic and clinical characteristics of 122 singleton bipolar patients with those of 54 bipolar patients derived from parent-offspring trios. The singleton patients did not only present with a higher age of onset, but also with a higher frequency of suicidal behavior and a higher familial loading for suicidality. These findings suggest that the genetic mechanism for disease might be different between trio-based and classic case control samples, where patients are examined whose parents are not available for genetic studies. Thus, giving up case control designs for the sake of family-based association studies could be at the risk of selecting against several genetically determined factors.

Endothelin-1 as a regulator of hepatic microcirculation: sublobular distribution of effects and impact on hepatocellular secretory function.

Endothelin-1 is a potent vasoconstrictor in the portal circulation causing sinusoidal constriction as well as presinusoidal resistance changes. Using in vivo videomicroscopy we studied the sublobular differences in dose response characteristics of sinusoid constriction at 10-13 min into continuous infusion of 5, 1.0, 3.0, and 5.0 pmol of endothelin-1 (ET-1)/100 g body weight/min in pentobarbital anesthetized rats. In addition, bile flow was monitored to estimate parenchymal secretory function. ET-1 evoked a profound constrictor effect in both sublobular regions studied. However, the maximal decrease of sinusoidal width in periportal inflow region (zone 1; 1 pmol/100 g/min: 4.8 +/- .1 microns) was reached at slightly lower concentrations of the peptide than in pericentral outflow region (zone 3; 3 pmol/100 g/min: 6.2 +/- .3 microns) compared to respective baseline values (zone 1: 7.1 +/- .1 microns; zone 3: 10.2 +/- .1 microns), suggesting upstream binding and clearance of the peptide. The constrictor response in zone 1 was biphasic and at higher concentrations of the peptide (5 pmol/100 g/min: 5.5 +/- .2 microns) sinusoidal widths increased again compared to the maximal response with the 1 pmol. Secretory function as reflected by the bile flow was maintained or even slightly increased with the lower doses (.5 and 1 pmol) of ET-1 and during the first 10 min into infusion of the higher doses (3 and 5 pmol). Subsequently, an approximately 20-25% decrease in bile flow accompanied the infusion of higher doses of ET-1.(ABSTRACT TRUNCATED AT 250 WORDS)

Chronic ethanol consumption exacerbates liver injury following hemorrhagic shock: role of sinusoidal perfusion failure.

Although the deleterious effect of chronic ethanol consumption on subsequent stressful events has long been recognized, the pathophysiological mechanisms are incompletely understood. This study tested whether chronic ethanol consumption in doses that increase sinusoidal contractility increases susceptibility to hepatic microvascular failure and liver injury after hemorrhagic shock. Liver microcirculation was assessed by in vivo microscopy during hemorrhage and up to 24 h after onset of resuscitation and was compared with liver histology and serum enzyme levels. Mean sinusoidal blood flow was neither impaired by chronic ethanol feeding at baseline nor during hemorrhage and early resuscitation. However, failure of individual sinusoids to conduct flow was observed more frequently after fluid resuscitation in ethanol-fed animals (e.g. at 1 h after onset of volume therapy: 26% of sinusoids) than in controls (11%), reflecting substantial flow heterogeneity. Failing sinusoids had substantially smaller diameters than sinusoids conducting flow with a more profound and sustained response in ethanol-fed rats. At 24 h marked pericentral necrosis and increase in serum alanine aminotransferase levels were observed in six of nine surviving ethanol-fed animals but only in 1 of 10 pair fed controls and correlated with microvascular failure. These data suggest that early as well as late microvascular failure in this model of hemorrhagic shock and resuscitation is primarily mediated at the level of individual sinusoids. Chronic ethanol feeding exacerbates microvascular and hepatocellular injury after shock/resuscitation, probably involving increased sinusoidal contractile responsiveness.

Role of reactive oxygen species for hepatocellular injury and heme oxygenase-1 gene expression after hemorrhage and resuscitation.

Reactive oxygen species (ROS) generated during hemorrhage and subsequent resuscitation (H/R) may contribute to cellular injury but may also regulate an adaptive cellular response to stress. Heme oxygenase (HO)-1 has been recognized as an important stress-inducible gene conferring protection after H/R. The aim of this study was to determine the contribution of ROS to hepatocellular injury and to induction of HO-1 in parenchymal and nonparenchymal cells after H/R. Anesthetized Sprague-Dawley rats were subjected to reversible H/R with or without coadministration of the potent antioxidant Trolox (6 mg/kg body wt). HO-1 gene expression was determined at baseline, at the end of hemorrhagic hypotension, and after 1, 3, and 5 h of resuscitation on the messenger ribonucleic acid (mRNA) and protein level. Assessment of hepatocellular injury by alpha-glutathione-S-transferase serum levels showed a significant increase after H/R that was attenuated by Trolox (sham: 38 (26-42); H/R: 286 (150-696); Trolox: 14 (2-227) microg/L; median (25th/75th percentile) P<0.05). Injury correlated with induction of HO-1 mRNA (r2 = 0.97) on the whole organ level and with the expression pattern of HO-1-immunoreactive protein in pericentral hepatocytes after H/R. Trolox attenuated H/R-induced increase of HO-1 in hepatocytes. In contrast, nonparenchymal cells showed high constitutive levels of HO-1 mRNA and protein that were increased by sham operation and H/R to a similar extent. HO-1 steady-state transcripts in nonparenchymal cells were not modulated by Trolox. These results suggest a differential regulation of HO-1 gene expression in hepatocytes and nonparenchymal cells. ROS formation seems to contribute to early hepatocellular injury but also serves as an important trigger for HO-1 gene expression in parenchymal cells, which confers delayed protection after H/R.