Tomohisa Inoue

Vicinal dithiol-binding agent, phenylarsine oxide, inhibits inducible nitric-oxide synthase gene expression at a step of nuclear factor-kappaB DNA binding in hepatocytes.

Inflammatory cytokine interleukin 1β induces inducible nitric-oxide synthase (iNOS) mRNA and its protein, which are followed by increasing the production of nitric oxide, in primary cultures of rat hepatocytes. Nuclear factor-κB (NF-κB), an important transcription factor for iNOS gene expression, is also activated and translocated to the nucleus. In the present study, we found that vicinal dithiol-binding agent, phenylarsine oxide (PAO), inhibited the induction of iNOS protein and mRNA as well as the release of nitrite (nitric oxide metabolite) into the culture medium. Simultaneous addition of a vicinal dithiol compound, 2,3-dimercaptopropanol, with PAO completely abolished these inhibitions. PAO could not prevent either degradation of an inhibitory protein, IκB, of NF-κB or translocation of NF-κB to the nucleus. However, electrophoretic mobility shift assay demonstrated that PAO decreased the interaction between NF-κB and its binding consensus oligonucleotide. Transfection experiments with iNOS promoter-luciferase construct revealed that PAO inhibited NF-κB binding to DNA. These results indicate that PAO inhibits iNOS gene expression at a step of NF-κB binding to DNA by modifying its vicinal dithiol moiety, which may play a crucial role for the iNOS regulation in hepatocytes.

An enhancement of nitric oxide production regulates energy metabolism in rat hepatocytes after a partial hepatectomy

BACKGROUND/AIMS Infection after a liver resection often results in hepatic failure. Nitric oxide is one of the candidates which has been suspected to cause cellular dysfunction during infection in the liver. We have previously reported that the inflammatory cytokine interleukin-1beta (IL-1beta) induced the expression of the inducible nitric oxide synthase gene in primary cultured rat hepatocytes. We hypothesized that an enhancement of nitric oxide production after the resection was implicated in a change in liver energy metabolism, thus resulting in liver dysfunction. METHODS In this study, we performed a 70% hepatectomy or a sham operation in rats, and then isolated hepatocytes from the remnant liver by collagenase perfusion. The cultured hepatocytes were treated with cytokines including IL-1beta. The effects on nitric oxide induction, the ATP content and ketone body ratio (acetoacetate/beta-hydroxybutyrate) were then compared between the partial hepatectomized (PH) and sham-operated (control) rats. RESULTS IL-1beta augmented the induction of nitric oxide production two-fold in hepatocytes from the PH rats as compared to the control rats. IL-1beta markedly decreased the ATP content in the PH rats, although IL-1beta also decreased the ATP content in the control rats, but to a lesser extent. IL-1beta also decreased the ketone body ratio in both groups. The addition of L-arginine further stimulated the inhibition of the ATP levels and the ketone body ratio concomitantly with increased nitric oxide production in the PH rats. N(G)-monomethyl-L-arginine, an inhibitor of nitric oxide synthase, abolished the effects of IL-1beta on the ATP levels and ketone body ratio, as well as on the nitric oxide production. CONCLUSIONS These results demonstrate that the decreased ATP content observed in PH rats resulted from an increase in nitric oxide production. The decrease in ketone body ratio indicates that nitric oxide-induced mitochondrial dysfunction contributes significantly to ATP attenuation in hepatocytes. Therefore, the regulation of nitric oxide induction may be crucial for preventing liver failure after a hepatic resection.

Nitric oxide production and hepatic dysfunction in patients with postoperative sepsis.

1. Although hepatic function is well known to deteriorate following bacterial infection, the underlying mechanisms remain poorly understood. We have previously reported that nitric oxide (NO) radical leads to a decrease in the ketone body ratio (KBR) and in ATP content due to the inhibition of mitochondrial electron transport in primary cultured rat hepatocytes.

Acceptance of Skin Allografts in Pigs by Portal Venous Injection of Donor Bone Marrow Cells

OBJECTIVE To confirm in pigs whether a new method for organ allografts, originally established in mice by the authors, might be applicable to humans. SUMMARY BACKGROUND DATA The authors recently established a new method for organ allografts in mice that includes the injection of donor bone marrow cells (BMCs) using the portal vein (PV), followed by the administration of cyclosporin A (CsA) on days 2 and 5, and the intravenous injection of BMCs on day 5. In the present study, they modify this method (a single-day protocol) and apply it to pigs. METHODS Allogeneic BMCs of donor pigs were injected using the PV (a superior mesenteric vein). The skin grafting was carried out on the day of the PV injection. The recipient pigs received donor grafts, autologous grafts, and third-party grafts at the same time. In addition, an open wound was made as the epithelized control. Full-thickness skin grafts were harvested from the dorsal wall of the donors. CsA (10 mg/kg) was injected intramuscularly into recipient pigs on days 2 and 5 after the PV injection. RESULTS One hundred percent of skin grafts survived for >300 days when donor BMCs were injected using the PV (n = 6). However, the skin grafts of the three pigs that had received BMCs using the intravenous route were rejected within 3 to 4 weeks after transplantation. The third-party skin grafts showed necrotic changes on day 21 after transplantation. CONCLUSIONS One hundred percent of skin allografts can be obtained, even in pigs, by injecting donor BMCs using the PV, carrying out skin allografts, and administering CsA on days 2 and 5. This single-day protocol would be of great advantage for human organ transplantation.

Ruptured Dissecting Aneurysm in Bilateral Iliac Arteries Caused by Ehlers-Danlos Syndrome Type IV: Report of a Case

Abstract Ehlers-Danlos syndrome (EDS) is an inherited disorder of connective tissue characterized by hyperextensible skin, hypermobile joints, and ab-normalities of the cardiovascular system. Ten types and several subtypes of EDS have so far been recognized based on genetic, clinical, and biochemical characteristics. The spectrum of the disorder varies from mild to life-threatening vascular complications. EDS type IV is a particularly dangerous form with a lethal spontaneous rupture of the major arteries and aneurysmal formation. We present herein a case of a ruptured dissecting aneurysm in the bilateral iliac arteries caused by EDS type IV. A previously healthy 33-year-old man without any physical features of this connective tissue disorder experienced a metachronous vascular rupture two times. Successful synthetic bypass grafting was performed with great difficulty. The diagnosis of EDS type IV was made afterwards based on an electrophoresis analysis of a skin biopsy specimen which revealed a lack of type III collagen. Surgical intervention in cases of arterial complications in EDS type IV patients have been reported to be both difficult and frequently unsuccessful. The early clinical recognition of this syndrome is therefore of great importance due to the hazards of such surgical therapies.