The Altered Metabolic Molecular Signatures Contribute to the RAD001 Resistance in Gastric Neuroendocrine Tumor.

Although the inhibition of mTOR is a promising treatment for neuroendocrine tumors, several questions are still open for cell specificity and resistance. With the newly characterized gastric neuroendocrine tumor mouse model (CEA424-SV40 T antigen transgenic mice), the anti-tumor efficiency of RAD001 (Everolimus) was tested both in vitro and in vivo. Tumor samples were analyzed for the expression of RNA by cDNA microarrays and also signaling pathways to get more details on the local surviving or selected cells. RAD001 treatment dramatically slowed down tumor growth and prolonged the animals' survival. This inhibitory effect has a preference for tumor cells since gastrointestinal hormone and neuroendocrine tumor specific markers were more reduced than the epithelial ones. While phosphorylation of p70S6K was almost completely blocked both in vitro and in vivo, the phosphorylation of 4EBP1 was only partially inhibited in vitro and unaffected in vivo. RAD001 treatment induced feedback activation of metabolism related pathways like PI(3)K-Akt-mTOR and MEK/ERK signalings. An induction of senescence as well as differential expression of genes responsible for metabolism was also observed, which highlighted the contribution of metabolic molecular signatures to the escape of the tumor cells from the treatment. Together, our data revealed efficient anti-tumor ability of RAD001 in a new gastric neuroendocrine tumor mouse model system and offered new insights into the clinical aspects of the incomplete elimination of tumor cells in patients treated.

Platelet GPIIb supports initial pulmonary retention but inhibits subsequent proliferation of melanoma cells during hematogenic metastasis.

Platelets modulate the process of cancer metastasis. However, current knowledge on the direct interaction of platelets and tumor cells is mostly based on findings obtained in vitro. We addressed the role of the platelet fibrinogen receptor glycoprotein IIb (integrin αIIb) for experimental melanoma metastasis in vivo. Highly metastatic B16-D5 melanoma cells were injected intravenously into GPIIb-deficient (GPIIb-/-) or wildtype (WT) mice. Acute accumulation of tumor cells in the pulmonary vasculature was assessed in real-time by confocal videofluorescence microscopy. Arrest of tumor cells was dramatically reduced in GPIIb-/- mice as compared to WT. Importantly, we found that mainly multicellular aggregates accumulated in the pulmonary circulation of WT, instead B16-D5 aggregates were significantly smaller in GPIIb-/- mice. While pulmonary arrest of melanoma was clearly dependent on GPIIb in this early phase of metastasis, we also addressed tumor progression 10 days after injection. Inversely, and unexpectedly, we found that melanoma metastasis was now increased in GPIIb-/- mice. In contrast, GPIIb did not regulate local melanoma proliferation in a subcutaneous tumor model. Our data suggest that the platelet fibrinogen receptor has a differential role in the modulation of hematogenic melanoma metastasis. While platelets clearly support early steps in pulmonary metastasis via GPIIb-dependent formation of platelet-tumor-aggregates, at a later stage its absence is associated with an accelerated development of melanoma metastases.

Expression of a neuroendocrine gene signature in gastric tumor cells from CEA 424-SV40 large T antigen-transgenic mice depends on SV40 large T antigen.

BACKGROUND: A large fraction of murine tumors induced by transgenic expression of SV40 large T antigen (SV40 TAg) exhibits a neuroendocrine phenotype. It is unclear whether SV40 TAg induces the neuroendocrine phenotype by preferential transformation of progenitor cells committed to the neuroendocrine lineage or by transcriptional activation of neuroendocrine genes. METHODOLOGY/PRINCIPAL FINDINGS: To address this question we analyzed CEA424-SV40 TAg-transgenic mice that develop spontaneous tumors in the antral stomach region. Immunohistology revealed expression of the neuroendocrine marker chromogranin A in tumor cells. By ELISA an 18-fold higher level of serotonin could be detected in the blood of tumor-bearing mice in comparison to nontransgenic littermates. Transcriptome analyses of antral tumors combined with gene set enrichment analysis showed significant enrichment of genes considered relevant for human neuroendocrine tumor biology. This neuroendocrine gene signature was also expressed in 424GC, a cell line derived from a CEA424-SV40 TAg tumor, indicating that the tumor cells exhibit a similar neuroendocrine phenotype also in vitro. Treatment of 424GC cells with SV40 TAg-specific siRNA downregulated expression of the neuroendocrine gene signature. CONCLUSIONS/SIGNIFICANCE: SV40 TAg thus appears to directly induce a neuroendocrine gene signature in gastric carcinomas of CEA424-SV40 TAg-transgenic mice. This might explain the high incidence of neuroendocrine tumors in other murine SV40 TAg tumor models. Since the oncogenic effect of SV40 TAg is caused by inactivation of the tumor suppressor proteins p53 and RB1 and loss of function of these proteins is commonly observed in human neuroendocrine tumors, a similar mechanism might cause neuroendocrine phenotypes in human tumors.

Matrix metalloproteinase-9 promotes neutrophil and T cell recruitment and migration in the postischemic liver.

Matrix metalloproteinases-2 and -9 (MMP-2/9) are critically involved in degradation of extracellular matrix, and their inhibition is discussed as a promising strategy against hepatic ischemia-reperfusion (I/R) injury. Here, we analyzed the role of MMP-2 and -9 for leukocyte migration and tissue injury in sham-operated mice and in mice after I/R, treated with a MMP-2/9 inhibitor or vehicle. Using zymography, we show that the MMP-2/9 inhibitor abolished I/R-induced MMP-9 activation, whereas MMP-2 activity was not detectable in all groups. As demonstrated by intravital microscopy, MMP-9 inhibition attenuated postischemic rolling and adherence of total leukocytes in hepatic postsinusoidal venules, CD4+ T cell accumulation in sinusoids, and neutrophil transmigration. These effects were associated with reduction of plasma tumor necrosis factor alpha (TNF-alpha) levels and endothelial expression of CD62P. Motility of interstitially migrating leukocytes was assessed by near-infrared reflected light oblique transillumination microscopy in the postischemic cremaster muscle. Upon MMP-9 blockade, leukocyte migration velocity and curve-line and straight-line migration distances were reduced significantly as compared with the vehicle-treated I/R group. Postischemic sinusoidal perfusion failure, hepatocellular apoptosis, and alanine aminotransferase activity were only slightly reduced after MMP-9 inhibition, whereas aspartate aminotransferase activity and mortality were significantly lower. In conclusion, MMP-9 is involved in the early recruitment cascades of neutrophils and CD4+ T cells, promotes neutrophil and T cell transmigration during hepatic I/R, and is required for motility of interstitially migrating leukocytes. MMP-9 blockade is associated with an attenuation of TNF-alpha release and endothelial CD62P expression, weakly protects from early microvascular/hepatocellular I/R damage, but improves postischemic survival.

Synergistic effects of brain death and liver steatosis on the hepatic microcirculation.

BACKGROUND: The routine transplantation of steatotic livers could potentially mitigate the donor shortage, but so far is associated with a high rate of graft dysfunction. Steatosis and brain death have been perceived as independent risk factors, but they may synergistically target the hepatic microcirculation. This study compares the effects of brain death on the microcirculation of steatotic and normal livers. METHODS: Brain death was induced in obese and lean Zucker rats. Lean and obese sham-operated animals served as controls. Liver microcirculation was investigated using intravital fluorescence microscopy. Serum liver enzyme and reduced glutathione, expression of P-selectin, ICAM-1 and VCAM-1 mRNA in the liver were determined. The ultrastructural alterations were compared by electron microscopy. RESULTS: In nonbrain-dead animals, liver steatosis was associated with smaller sinusoidal diameters, but did not impair sinusoidal perfusion. During brain death, sinusoidal diameter and perfusion were reduced in normal and, to a greater extent, in steatotic livers. Also, more leukocytes were recruited to the microvasculature of steatotic livers than to normal livers in brain-dead state. The highest liver enzyme activities and the lowest hepatic GSH concentrations were measured in brain-dead animals with steatotic livers; only in these organs was endothelial cell swelling regularly observed. In brain-dead state, only the P-selectin mRNA expression was increased in steatotic livers as compared to normal livers. CONCLUSIONS: Brain death amplifies the adverse effects of steatosis on the hepatic microcirculation. Our results underline the need for therapeutic intervention in brain-dead state when steatotic livers are to be used for transplantation.

Junctional adhesion molecule-A deficiency increases hepatic ischemia-reperfusion injury despite reduction of neutrophil transendothelial migration.

The endothelial receptors that control leukocyte transmigration in the postischemic liver are not identified. We investigated the role of junctional adhesion molecule-A (JAM-A), a receptor expressed in endothelial tight junctions, leukocytes, and platelets, for leukocyte transmigration during hepatic ischemia-reperfusion (I/R) in vivo. We show that JAM-A is up-regulated in hepatic venular endothelium during reperfusion. I/R-induced neutrophil transmigration was attenuated in both JAM-A-/- and endothelial JAM-A-/- mice as well as in mice treated with an anti-JAM-A antibody, whereas transmigration of T cells was JAM-A independent. Postischemic leukocyte rolling remained unaffected in JAM-A-/- and endothelial JAM-A-/- mice, whereas intravascular leukocyte adherence was increased. The extent of interactions of JAM-A-/- platelets with the postischemic endothelium was comparable with that of JAM-A+/+ platelets. The I/R-induced increase in the activity of alanine aminotransferase (ALT)/aspartate aminotransferase (AST) and sinusoidal perfusion failure was not reduced in JAM-A-/- mice, while the number of terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL)-positive hepatocytes was significantly higher. Thus, we show for the first time that JAM-A is up-regulated in hepatic venules and serves as an endothelial receptor of neutrophil transmigration, but it does not mediate leukocyte rolling, adhesion, or platelet-endothelial cell interactions. JAM-A deficiency does not reduce I/R-induced microvascular and hepatocellular necrotic injury, but increases hepatocyte apoptosis, despite attenuation of neutrophil infiltration.

A new animal model to assess angiogenesis and endocrine function of parathyroid heterografts in vivo.

BACKGROUND: It is still a matter of investigation how angiogenesis and restoration of gland perfusion determine graft function after free parathyroid autotransplantation. We provide a new animal model allowing simultaneous and repetitive in vivo assessment of angiogenesis and endocrine function of parathyroid transplants. METHODS: Fresh human parathyroid tissue from patients with secondary hyperparathyroidism was grafted into dorsal skinfold chamber preparations of athymic nude mice (CD1-nu; n=8). Equivalent pieces of the same human donor specimens were heat-inactivated and served as control grafts (n=7). RESULTS: In all animals receiving parathyroid transplants, intact human parathyroid hormone levels were detectable by species-specific enzyme-linked immunosorbent assay analysis of plasma samples on day 5 after transplantation and increased by 2.5-fold over the observation period (19 days) in contrast with controls. Plasma Ca levels revealed no differences between the groups. On day 5 after transplantation, intravital fluorescence microscopy revealed murine angiogenic microvessels sprouting along nonperfused human donor vessels, and 1 week later functional microvasculature was established in all parathyroid transplants. Histologic analysis revealed well-vascularized endocrine tissue. In contrast, control grafts were necrotic and partly resorbed; they exhibited no angiogenic activity or well-vascularized fat cells indicating fatty degeneration. In addition, species-specific Western blot analysis revealed vascular endothelial growth factor expression of parathyroid transplants rather than functional vessel density as the functional parameter of angiogenesis determining transplant function in vivo. CONCLUSION: This model may serve to understand mechanisms associated with specific parathyroid transplant angiogenesis and its significance for transplant function to optimize clinical success of autotransplantation in therapy-resistant patients.

Mycoplasma haemocanis infection--a kennel disease?

Mycoplasma haemocanis (formerly Haemobartonella canis) is a red blood cell parasite that causes disease mainly in immunosuppressed and splenectomized dogs. Clinical outbreak of the disease resulted in failure of a large experimental project. We aimed to identify whether M. haemocanis has increased prevalence in kennel-raised dogs. In a prospective study, we compared the prevalence of M. haemocanis in whole blood (anti-coagulated by use of EDTA) collected from pet dogs (University of Illinois, Urbana Champaign, Ill.; n = 60) with that in blood from dogs raised in three distinct kennels in western Europe (WE; n = 23), eastern Europe (EE; n = 20), and North America (NA; n = 20). Screening included antibody testing and microscopy of blood smears. The presence of M. haemocanis was identified using a polymerase chain reaction (PCR) assay for specific DNA of the organism. None of the pet dogs (0%) was test positive for M. haemocanis DNA. Mycoplasma haemocanis was found in dogs tested at all of the kennels. Infection rate in the three kennels was 30, 35, and 87%, respectively (all P < 0.001 versus control, chi2-test). Latent infection with M. haemocanis was not a single observation in kennel-raised dogs. Prevalence may be higher than that in a pet dog population. The potential exists for these latent infections to adversely affect or confound research results.

5-Aminoisoquinolinone, a novel inhibitor of poly(adenosine disphosphate-ribose) polymerase, reduces microvascular liver injury but not mortality rate after hepatic ischemia-reperfusion.

OBJECTIVE: The aim of this study was to investigate the impact of the novel, potent, water-soluble inhibitor of poly(adenosine diphosphate-ribose) polymerase (PARP) 5-aminoisoquinolinone (5-AIQ) on hepatic microcirculation, hepatocellular injury, and survival in a murine model of hepatic ischemia-reperfusion. DESIGN: Randomized animal study. SETTING: Research laboratory. SUBJECTS: C57BL6 mice were subjected to warm either partial (90 mins) or total (75 mins) ischemia of the liver. INTERVENTIONS: Either PARP inhibitor 5-AIQ (3 mg/kg) or vehicle was administered to mice intravenously immediately before the start of reperfusion. Sham-operated animals served as controls. MEASUREMENTS AND MAIN RESULTS: As shown by intravital fluorescence microscopy after 30-60 mins of reperfusion, ischemia-reperfusion significantly enhanced platelet- and leukocyte-endothelial cell interactions in hepatic microvessels and impaired sinusoidal perfusion. Hepatocellular injury was characterized by an increase in the number of necrotic and apoptotic cells, dramatic elevation of aspartate aminotransferase/alanine aminotransferase serum activity, and lipid peroxidation in liver tissue. 5-AIQ treatment attenuated ischemia-reperfusion-induced increases in the numbers of adherent platelets and leukocytes as well as of necrotic and apoptotic cells and ameliorated perfusion failure. Furthermore, PARP inhibition prevented the increase in aspartate aminotransferase activity after ischemia-reperfusion but did not affect postischemic alanine aminotransferase release. However, no protective impact of 5-AIQ on postischemic oxidative stress was observed. Although PARP inhibition did not alter the survival percentage after ischemia-reperfusion (22% in both groups), this approach prolonged survival from 1 to 24 hrs (ischemia-reperfusion + vehicle) up to 48-72 hrs in the treated group. CONCLUSIONS: PARP inhibition with 5-AIQ during hepatic ischemia-reperfusion attenuates microvascular injury and reduces the extent of necrotic/apoptotic cell damage but does not protect from oxidative injury and does not improve postoperative survival rate.

Impact of intraischemic temperature on oxidative stress during hepatic reperfusion.

This study was designed to investigate the influence of intraischemic liver temperature on oxidative stress during postischemic normothermic reperfusion. In C57BL/6 mice, partial hepatic ischemia was induced for 90 min and intraischemic organ temperature adjusted to 4 degrees C, 15 degrees C, 26 degrees C, 32 degrees C, and 37 degrees C. As detected by electron spin-resonance spectroscopy, plasma/blood concentrations of hydroxyl and ascorbyl radicals were significantly increased in all groups after ischemia/reperfusion independent of the intraischemic temperature. In tissue, however, postischemic lipid peroxidation was attenuated after organ cooling down to 32 degrees C-26 degrees C and not detectable after ischemia at 15 degrees C-4 degrees C. mRNA expression of superoxide dismutase-1 and heme oxygenase-1, measured during reperfusion, was significantly elevated in the group at 37 degrees C as compared to the hypothermic groups at 4 degrees C-32 degrees C. The reduction of radical generation was associated with a prevention of adenosine monophosphate hydrolysis during ischemia in the hypothermic groups. In conclusion, ischemia-reperfusion-induced oxidative stress in the liver tissue is non-linearly-dependent on intraischemic temperature, whereas the plasma/blood concentration of radicals is not affected by organ cooling. Oxidative stress is reduced through mild hypothermia at 32 degrees C-26 degrees C and inhibited completely at 15 degrees C. Reduction of initial intracellular radical generation and prevention of secondary oxidant-induced tissue injury are possible mechanisms of this protection.

Heat-shock preconditioning protects fatty livers in genetically obese Zucker rats from microvascular perfusion failure after ischemia reperfusion.

Reduced tolerance of steatotic livers to ischemic injury is considered to correlate with impaired microcirculation. The aim of this study was to investigate the impact of heat-shock preconditioning (HSPC) on microcirculatory failure after ischemia/reperfusion (I/R) in steatotic livers by means of intra-vital fluorescence microscopy. Obese Zucker rats were used. In the HS group, rats underwent whole-body hyperthermia followed by 60-min partial liver ischemia. In group IR, rats were exposed only to ischemia. Microcirculation parameters (sinusoidal perfusion rate, sinusoidal diameter, leukocyte-endothelial interaction) were significantly better preserved in the HS group than in the IR group. Liver enzymes, oxygenated glutathione/reduced glutathione (GSSG/GSH) ratio, and electron microscopy showed less damage in the HS group. A marked expression of heat shock protein 72 (HSP72) and heme oxygenase (HO-1) was found only in the livers of group HS. HSPC mitigated the I/R injury of steatotic livers by preventing post-ischemic failure of microcirculation. This beneficial effect was found to be associated with the induction of HSP72 and HO-1.

Up-regulation of inducible nitric oxide synthase in Helicobacter pylori-associated gastritis may represent an increased risk factor to develop gastric carcinoma of the intestinal type.

The enzyme inducible nitric oxide synthase (iNOS) is part of the host innate defense system against bacterial infection. During chronic inflammation, like that seen with a Helicobacter pylori infection, constant nitric oxide production may lead to tissue and DNA damage, thus increasing the patient's risk for developing cancer. Several investigations on iNOS expression in H. pylori-associated gastritis have resulted in conflicting data. Therefore, we investigated the association between chronic H. pylori infection and iNOS expression in samples from stomach carcinoma patients as well as in antral biopsies from patients with H. pylori-associated gastritis. iNOS expression was analyzed by means of reverse transcriptase (RT)-PCR and quantified by competitive RT-PCR. To study in situ localization of iNOS in biopsy samples, immunohistochemistry was performed. iNOS enzyme activity was quantified using an arginine/citrulline assay. A significant increase in iNOS mRNA signal was only present in one-third of the analyzed patient biopsies with H. pylori-associated gastritis. These biopsies showed a 90% association with intestinal metaplasia and a 100% association with CagA-positive H. pylori. Intestinal metaplasia is discussed to be one step in the carcinogenesis of stomach cancer. Quantitation of iNOS transcripts and iNOS enzyme activity in non-cancerous mucosa of gastric cancer patients revealed a significant increase in iNOS transcripts and iNOS activity only in the mucosa of patients with stomach cancer of the intestinal type but not in the diffuse type. Our results support the hypothesis that CagA-positive H. pylori strains are associated with the expression and activity of iNOS, and therefore might contribute to the development of intestinal metaplasia leading to gastric cancer of the intestinal type.

P-selectin mediates platelet-endothelial cell interactions and reperfusion injury in the mouse liver in vivo.

Platelets are suggested to participate in the pathogenesis of hepatic ischemia-reperfusion (I/R) injury. This study was designed to analyze platelet-endothelial cell interactions in the postischemic mouse liver in vivo and to define the role of endothelial versus platelet P-selectin for these interactions. Platelet-endothelial cell interactions were quantitatively analyzed using intravital fluorescence microscopy after lobar hepatic I/R in C57BL/6 wild-type and P-selectin-deficient mice after infusion of ex vivo rhodamine-6G-labeled wild-type and P-selectin-deficient platelets. Reperfusion injury and apoptosis were assessed by established methods. In wild-type animals, hepatic I/R caused significantly enhanced platelet-endothelial cell interactions in terminal arterioles and postsinusoidal venules as well as platelet stagnation in sinusoids. Concomitantly, transaminase and caspase-3 activities were elevated and sinusoidal perfusion was impaired. In contrast, platelet-endothelial cell interactions were nearly absent in arterioles and venules of mice lacking endothelial P-selectin, irrespective of the presence of P-selectin on infused platelets, but still significantly elevated in sinusoids. Simultaneously, sinusoidal perfusion failure was ameliorated, and transaminase- and caspase-3 activities were significantly reduced in P-selectin-deficient mice as compared with wild-type animals. The present intravital microscopic study provides, for the first time, quantitative analyses of platelet-endothelial cell interactions in the postischemic hepatic microcirculation. Our in vivo data show that endothelial P-selectin is critical for postischemic platelet-endothelial cell interactions within hepatic presinusoidal arterioles and postsinusoidal venules. P-selectin deficiency prevents microvascular injury and apoptosis after warm hepatic I/R.

Platelet adhesion mediated by fibrinogen-intercelllular adhesion molecule-1 binding induces tissue injury in the postischemic liver in vivo.

BACKGROUND: Platelets are thought to be involved in the induction of hepatic ischemia-reperfusion (I/R) injury. The mechanisms of platelet adhesion in the hepatic microvasculature and the role of platelets in the pathogenesis of I/R-induced liver damage in vivo remain unclear. METHODS: In C57BL/6 mice, platelet- and leukocyte-endothelial cell interactions were quantitatively analyzed using intravital fluorescence microscopy in sham-operated animals, after warm lobar hepatic I/R (90/20 min) in wild-type and intercellular adhesion molecule (ICAM)-1-deficient mice, and after I/R in wild-type mice treated with an antifibrinogen antibody. Fibrinogen deposition on the endothelium was detected by intravital microscopy and by immunostaining. Reperfusion injury was assessed by measurement of liver enzyme and caspase-3 activities and of lipid peroxidation. RESULTS: Hepatic I/R induced fibrinogen deposition on hepatic endothelium, followed by a dramatic increase in the number of firmly adherent platelets in the liver microvasculature. Simultaneously, the number of adherent leukocytes in postsinusoidal venules and the aspartate aminotransferase/alanine aminotransferase and caspase-3 activities were elevated. Although ICAM-1 deficiency attenuated postischemic adherence of both platelets and leukocytes, the application of an antifibrinogen antibody selectively reduced the number of adherent platelets but did not influence leukocyte adhesion. The selective blockade of platelet adherence significantly prevented the postischemic increase in liver enzyme and caspase-3 activities. Furthermore, sinusoidal perfusion failure and lipid peroxidation were attenuated in the treated group. CONCLUSIONS: These in vivo data show that platelet adhesion mediated through fibrinogen deposition on ICAM-1 expressed on the endothelium of postischemic hepatic microvessels induces microvascular injury and hepatocellular apoptosis after I/R of the liver during early reperfusion.

Mechanism of antigen presentation after hypertonic loading of soluble antigens.

Hypertonic loading of proteins into cells has been used to introduce soluble proteins into the major histocompatibility complex class I pathway of antigen presentation followed by cytotoxic T-lymphocyte (CTL) induction. The precise mechanism for this pathway is not completely understood. The antigen is either processed and presented by/on the same cell or by professional antigen-presenting cells (APC) after taking up the antigen from damaged or apoptotic cells. After loading labelled ovalbumin (OVA), it could be co-precipitated with the proteasome complex, supporting the role of this pathway for antigen processing. The processing speed however, appeared to be slow since intact OVA could be detected inside the cells even after 18 hr. This corresponded well with the processing of OVA by isolated proteasomes. On the other hand, enough peptides for recognition of target cells by CTLs were generated in this reaction. One reason for the low level of processing might be that hypertonic loading may damage the cells and inhibit direct processing. In fact, at least 50% of the cells became positive for Annexin V binding after hypertonic loading which indicates severe membrane alterations usually associated with the progress of apoptosis. Annexin V binds to phosphatidylserine residues which also serve as ligand for CD36 expressed on monocytes and some immature dendritic cells. This may direct the phagocytic pathway to hypertonically loaded cells and thus enable professional APCs to present OVA-peptides. Therefore, in addition to the direct processing of OVA, CTLs can be primed by professional APC after uptake of apoptotic, OVA-loaded cells.

Poly(ADP-ribose) polymerase triggers the microvascular mechanisms of hepatic ischemia-reperfusion injury.

Activation of poly(ADP-ribose) polymerase (PARP) mediates oxidative stress-induced cell injury. We tested the hypothesis that PARP contributes to ischemia-reperfusion (I/R) damage of the liver by triggering the mechanisms of microcirculatory failure. Leukocyte- and platelet-endothelial cell interactions as well as sinusoidal perfusion were analyzed by intravital fluorescence microscopy after lobar hepatic I/R (90 min/30 min) in C57BL/6 x 129/Sv wild-type (PARP+/+) and PARP-deficient (PARP-/-) mice. Hepatic I/R induced leukocyte/platelet-endothelial cell interactions and tissue injury in PARP+/+ mice, as indicated by impaired sinusoidal perfusion and increased alanine aminotransferase (ALT)/aspartate aminotransferase (AST) serum activities. In PARP-/- mice, however, the postischemic increase in the numbers of rolling/adherent leukocytes and platelets was significantly lower. In addition, I/R-induced translocation of CD62P as well as mRNA expression of CD62E, CD54, and CD106 were attenuated. The degree of perfusion failure was reduced and the increase in the ALT/AST activities was lower in PARP-/- mice compared with PARP+/+ mice. We conclude that PARP contributes to hepatic microvascular injury by triggering the expression/translocation of adhesion molecules and modulating leukocyte/platelet-endothelial cell interactions.