Induction of HSP70 shows differences in protection against I/R injury derived by ischemic preconditioning and intermittent clamping.

BACKGROUND: Ischemic preconditioning (IP) and intermittent clamping (IC) increase the ischemic tolerance of the liver. The underlying mechanisms are not completely understood. Heat shock proteins protect cellular integrity in stress and have been discussed as mediators in preconditioning. IP and IC in rat livers were compared with respect to HSP induction and postischemic microcirculation. METHODS: All animals were exposed to 70min of partial warm liver ischemia. Different clamping protocols were used: in control animals (C) 70min continuous ischemia was applied. IP was performed by 5min ischemia and 10min reperfusion before the 70min ischemia time. In IC-groups, ischemia time of 70min was divided into four intervals. Each group included 21 animals with 3 different reperfusion intervals; either 30min, 12 or 36h. Intravital microscopy was performed after 30min of reperfusion. AST-levels and HSP induction were analysed 90min, 12 and 36h after reperfusion. RESULTS: IP and IC significantly improved sinusoidal perfusion (IP: 83.4±2.8%; IC: 84.4±4.6% vs. C: 60.4±3.9%; p<0.001) and leucocyte adherence in sinusoids (IP: 51.9±12.0, IC: 40.9±4.7 vs. C: 90.1±17.7/mm(2) liver surface; p<0.001) and postsinusoidal venules. AST-levels were minimized in IP and IC compared to controls (12h after reperfusion: IP: 969±934U/l, IC: 675±562U/l vs. C: 2373±792U/l; p=0.004). In the course of reperfusion HSP70 protein expression doubled between 90min and 12h in IC (0.529±0.227 vs. 0.992±0.246; p<0.05) and control-groups (0.572±0.314 vs. 1.106±0.309; p<0.05) whereas it remained unchanged in the IP-group (0.437±0.383 vs. 0.412±0.439; n.s.). CONCLUSION: Microcirculation is similarly preserved by IP and IC. The early protection derived by IP prevents further induction of HSP70 in opposite to IC. Therefore, IP may offer a more comprehensive protection against I/R on a cellular and transcriptional level.

Effects of hemodilution with a hemoglobin-based oxygen carrier (HBOC-201) on ischemia/reperfusion injury in a model of partial warm liver ischemia of the rat.

BACKGROUND: Ischemia/reperfusion injury is an unavoidable complication in liver surgery and transplantation. Hemodilution with colloids can reduce postischemic injury but limits oxygen transport. Hemoglobin-based oxygen carriers have been evaluated as blood substitute and provide a plasma-derived oxygen transport. It was the aim of our study to evaluate the combined benefits of hemodilution with a better oxygen supply to reperfused liver tissue by the use of HBOC-201 (Hemopure). MATERIAL AND METHODS: A model of partial warm liver ischemia in the rat was used. One group served as untreated control, the other groups were hemodiluted either with Ringer's lactate, Dextran-70, HBOC-201 or a mixture of Dextran and HBOC-201. After reperfusion, intravital microscopy studies were done and tissue pO(2) levels and transaminases measured. Statistical analysis was done by one- and two-way ANOVA, followed by pairwise comparison. RESULTS: Hemodilution with Ringer's lactate did not show any improvement compared to the control group. Dextran and HBOC group were superior to the Ringer and control animals in all parameters studied. Leucocyte adherence in postsinusoidal venules improved from 569.03+/-171.87 and 364.52+/-167.32 in control and Ringer group to 131.68+/-58.34 and 68.44+/-20.31/mm(2) endothelium in Dextran and HBOC group (p<0.001). Concerning tissue pO(2) levels, HBOC (23.4+/-5.0 mmHg) proved to be superior to Dextran (7.9+/-4.4 mmHg; p=0.007). CONCLUSION: HBOC was equivalent to Dextran in reducing I/R injury in the liver, but improved oxygenation of postreperfusion liver tissue.

Quantification of liver perfusion by dynamic magnetic resonance imaging: experimental evaluation and clinical pilot study.

Changes in liver microcirculation are considered essential in assessing ischemia-reperfusion injury, which in turn has an impact on liver graft function and outcome following liver transplantation (LTx). The aim of this study was to introduce dynamic magnetic resonance imaging (dMRI) as a new technique for overall quantification of hepatic microcirculation and compare it to perfusion measured by laser Doppler flowmetry (LDF; hepatic artery/portal vein) and thermal diffusion (TD). The study included 3 groups, measuring hepatic blood flow and microcirculation with the help of TD, LDF, and dMRI. In group I (9 landrace pigs; 26 +/- 5 kg), the native liver before and after partial portal occlusion was studied; in group II (6 landrace pigs; 25.5 +/- 4.4 kg), the liver 24 hours after LTx was studied; and in group III (14 patients), the liver on days 4 to 7 following LTx was studied. A close correlation was found between dMRI measurements and TD (r = 0.7-0.9, P < 0.01) in 4 defined regions of interest. Portal blood flow and partial occlusion of the portal vein were accurately detected by LDF flowmetry and correlated well with dMRI (r = 0.95, P < 0.01). In the clinical setting, representative TD measurements in segment 4b of the transplanted liver correlated well with dMRI analysis in other segments. Quantification of the portal blood flow and imaging of the whole liver could be performed simultaneously by dMRI. In conclusion, dMRI has been proved to be a sensitive modality for the quantification of liver microcirculation and hepatic blood flow in experimental and clinical LTx. It allows for a synchronous, noninvasive assessment of macrocirculation and microcirculation of the liver and could become a valuable diagnostic tool in advanced liver surgery and transplantation.

The influence of selenium substitution on microcirculation and glutathione metabolism after warm liver ischemia/reperfusion in a rat model.

Ischemia/reperfusion (I/R) injury is a variable yet unavoidable complication in liver surgery and transplantation. Selenium-dependent glutathione-peroxidases (GPx) and selenoproteins function as antioxidant defense systems. One target in preventing I/R injury is enhancing the capacity of endogenous redox defense. It was the aim of this study to analyze the effects of selenium substitution on liver microcirculation, hepatocellular injury and glutathione status in a model of partial warm liver ischemia in the rat. Sodium selenite was administered in three different dosages i.v.: 0.125 microg/g, 0.25 microg/g and 0.375 microg/g body weight and compared to an untreated control group (each n=10). Intravital microscopy was performed after 70 min of partial warm liver ischemia and 90 min of reperfusion. Liver tissue and plasma samples were taken at the end of the experiment for laboratory analysis. Microcirculation improved significantly in all therapy groups in contrast to control animals. ALT levels decreased significantly whereas malondialdehyde levels remained unchanged. In liver tissue, selenium supplementation caused an increase in the amount of total and reduced glutathione without changes in oxidized glutathione. This effect is likely mediated by selenite itself and selenoprotein P rather than by modulating GPx activity. We were able to show that selenite substitution has an immediate protective effect on I/R injury after warm hepatic ischemia by acting as a radical scavenger and preserving the antioxidative capacity of the liver.

Laparoscopic sigmoid resections for diverticulitis complicated by abscesses or fistulas.

BACKGROUND AND AIMS: Treatment of choice in recurrent and complicated diverticulitis is surgical resection of the inflamed bowel. Whereas it is accepted that recurrent diverticulitis (RD) can be handled laparoscopically, this is still not generally recommended for complicated diverticulitis (CD). Therefore, we analysed our results of laparoscopic sigmoidectomies concerning intraoperative course, conversion rate, morbidity and hospital stay in RD and CD. MATERIALS AND METHODS: Between 09/2002 and 01/2006, laparoscopic sigmoidectomies were offered to all patients suffering from recurrent or complicated diverticulitis (Hinchey I+II). All resections were performed in a four-port technique with the use of Ultracision and intraabdominal stapler anastomosis. Data were prospectively collected and retrospectively analysed in an intention-to-treat view. RESULTS: Out of 127 laparoscopic colectomies, 58 were performed for diverticulitis (RD 32; CD 26). Eight patients with colovesical and one patient with colovaginal fistula are included. Three patients with abscesses underwent pretreatment by percutaneous drainage. Operative time was longer in CD than in RD (205+/-41 vs 147+/-34 min; p<0.001) and associated with higher blood loss, but conversion rate was low (RD, 2/32 vs CD, 3/26; p=0.64). There was one intraoperative complication in each group; postoperative major complications occurred in 3.13% (RD) vs 11.5% (CD; p=0.316). One anastomotic leakage occurred in the RD group. Length of hospital stay was shorter for RD than for CD (7.1+/-3.4 vs 10.7+/-6.4 days; p=0.02). CONCLUSIONS: Laparoscopic resections should not be limited to recurrent diverticular disease but can be safely applied for complicated diverticulitis.

Liver biochemistry profile, significance and endoscopic management of biliary tract complications post orthotopic liver transplantation.

AIM: To correlate the significance of liver biochemical tests in diagnosing post orthotopic liver transplantation (OLT) biliary complications and to study their profile before and after endoscopic therapy. METHODS: Patients who developed biliary complications were analysed in detail for the clinical information, laboratory tests, treatment offered, response to it, follow up and outcomes. The profile of liver enzymes was determined. The safety, efficacy and outcomes of endoscopic retrograde cholangiography (ERC) were also analysed. RESULTS: 40 patients required ERC for 70 biliary complications. GGT was found to be > 3 times (388.1 +/- 70.9 U/mL vs 168.5 +/- 34.2 U/L, P=0.007) and SAP > 2 times (345.1 +/- 59.1 U/L vs 152.7 +/- 21.4 U/L, P=0.003) the immediate post OLT values. Most frequent complication was isolated anastomotic strictures in 28 (40%). Sustained success was achieved in 26 (81%) patients. CONCLUSION: Biliary complications still remain an important problem post OLT. SAP and GGT can be used as early, non-invasive markers for diagnosis and also to assess the adequacy of therapy. Endoscopic management is usually effective in treating the majority of these biliary complications.

Identification of differentially expressed genes after partial rat liver ischemia/reperfusion by suppression subtractive hybridization.

AIM: To identify potential diagnostic target genes in early reperfusion periods following warm liver ischemia before irreversible liver damage occurs. METHODS: We used two strategies (SSH suppression subtractive hybridization and hybridization of cDNA arrays) to determine early changes in gene expression profiles in a rat model of partial WI/R, comparing postischemic and adjacent nonischemic liver lobes. Differential gene expression was verified (WI/R; 1 h/2 h) and analyzed in more detail after warm ischemia (1 h) in a reperfusion time kinetics (0, 1, 2 and 6 h) and compared to untreated livers by Northern blot hybridizations. Protein expression was examined on Western blots and by immunohistochemistry for four differentially expressed target genes (Hsp70, Hsp27, Gadd45a and IL-1rI). RESULTS: Thirty-two individual WI/R target genes showing altered RNA levels after confirmation by Northern blot analyzes were identified. Among them, six functionally uncharacteristic expressed sequences and 26 known genes (12 induced in postischemic liver lobes, 14 with higher transcriptional expression in adjacent nonischemic liver lobes). Functional categories of the verified marker genes indicate on the one hand cellular stress and tissue damage but otherwise activation of protective cellular reactions (AP-1 transcription factors, apoptosis related genes, heat shock genes). In order to assign the transcriptional status to the biological relevant protein level we demonstrated that Hsp70, Hsp27, Gadd45a and IL-1rI were clearly up-regulated comparing postischemic and untreated rat livers, suggesting their involvement in the WI/R context. CONCLUSION: This study unveils a WI/R response gene set that will help to explore molecular pathways involved in the tissue damage after WI/R. In addition, these genes especially Hsp70 and Gadd45a might represent promising new candidates indicating WI/R liver damage.

Prothrombotic disorders in uremic type-1 diabetics undergoing simultaneous pancreas and kidney transplantation.

BACKGROUND: Although prothrombotic disorders (PTD) are known to increase the risk of graft failure in kidney transplantation only, there are no data on PTD in simultaneous pancreas and kidney transplantation (SPK). METHODS: Forty-seven SPK performed between September 2000 and July 2002 underwent routine screening for PTD. Data were retrospectively analyzed in view of complications (relaparotomy, graft thrombosis, pancreatitis, rejection) and graft function (HbA1c, serum creatinine) 3 months posttransplantation. RESULTS: Twenty-five of forty-seven (53.2%) patients had 30 PTDs. Homozygous mutations of the MTHFR gene (C677T) were found in six, factor-V Leiden mutation (homo- or heterozygous G1691A) in seven, and prothrombin mutation (20210A) in one patient (group 1). Group 2 consists of deficiencies of protein C (n=1), of protein S (n=12), of antithrombin (n=1), and antiphospholipid syndromes (n=2). Overall, PTD had no influence on graft thrombosis (P=0.36) or rejection (P=0.56). In patients with homozygous mutations, relaparotomies were more often necessary than in patients without mutations (42.9% vs. 11.8%, P=0.046). In group 1, there was a trend toward a higher incidence of graft pancreatitis than in patients without mutations (38.5% vs. 14.7%, P=0.075). Three months posttransplantation, HbA1c was 6.0% in patients with and 5.5% in patients without PTD (P=0.023). With regard to serum creatinine, no significant differences were observed. CONCLUSION: PTD are frequent in type-1 diabetics receiving SPK and may have a role in relaparotomies, graft pancreatitis, and pancreas graft function.

Value of donor swabs for intra-abdominal infection in simultaneous pancreas-kidney transplantation.

BACKGROUND: Simultaneous pancreas-kidney transplantation (SPK) has a higher rate of surgical complications compared with other whole organ transplantations. Graft thrombosis and intra-abdominal infections are the most frequent causes for relaparotomy. We evaluated risk factors for abdominal infections after SPK, with emphasis on the value of the routinely taken intraoperative swabs. METHODS: Between June 1994 and December 2000, 177 SPK were performed. Immunosuppression consisted of antithymocyte globulin induction and triple-drug maintenance therapy. Routine swabs were taken from the graft perfusion solutions, from the donors' duodenum, and from the recipients' bladder and jejunum (in case of enteric drainage). RESULTS: A total of 19 (10.7%) of 177 patients underwent 41 relaparotomies as a result of intra-abdominal infections. Positive microbial results from any donor site and positive duodenal swabs were significant risk factors for intra-abdominal infections after SPK (P=0.01, P=0.02). There was a significantly higher incidence of abdominal infections when Candida was found in the donor duodenal swab (P=0.0048). Patient survival was significantly lower in cases with abdominal infection (P=0.02). Survival rates of patients with and without abdominal infection were 89.5% and 97.4% at 1 year and 72.3% and 92.8% at 5 years, respectively. CONCLUSIONS: The results of this study confirm that abdominal infections significantly reduce patient survival and thus jeopardize the success of SPK. Positive donor duodenal swabs have been revealed to be a significant risk factor for a subsequent intra-abdominal infection, especially when Candida was found.