Kosuke Matsui

Natural antisense transcript stabilizes inducible nitric oxide synthase messenger RNA in rat hepatocytes

During inflammation, inducible nitric oxide synthase (iNOS) is induced to generate the important mediator nitric oxide (NO). Interleukin 1β (IL‐1β) induces iNOS messenger RNA (mRNA), iNOS protein, and NO in rat hepatocytes. We found that the stability of iNOS mRNA changed during the induction and that the antisense (AS) strand corresponding to the 3′‐untranslated region (3′UTR) of iNOS mRNA was transcribed from the iNOS gene. Expression levels of the iNOS AS transcript correlated with those of iNOS mRNA. The 1.5‐kilobase region 3′‐flanking to iNOS gene exon 27 was involved in IL‐1β induction. Knockdown experiments suggest that sense oligonucleotides to iNOS mRNA significantly reduced iNOS mRNA levels in the hepatocytes by blocking the interaction between iNOS mRNA and the AS transcript. Overexpression of iNOS AS transcript stabilized the reporter luciferase mRNA through the fused iNOS mRNA 3′UTR. These results together with the data in a yeast RNA‐hybrid assay suggested that the iNOS AS transcript interacted with iNOS mRNA and stabilized iNOS mRNA. The iNOS mRNA colocalized with the AU‐rich element‐binding protein HuR, a human homolog of embryonic lethal‐abnormal visual protein, and heterogeneous nuclear ribonucleoprotein L (hnRNP L) in the cytoplasm of rat hepatocytes. Interaction assays further revealed that the iNOS AS transcript interacted with HuR, which interacted with hnRNP L, suggesting that iNOS mRNA, the AS transcript, and the RNA‐binding proteins may mutually interact. Conclusion: The natural AS transcript of the iNOS gene interacts with iNOS mRNA and may play an important role in the stability of iNOS mRNA. This RNA‐RNA interaction may be a new therapeutic target for NO‐mediating inflammatory diseases. (HEPATOLOGY 2008.)

Predictors of microvascular invasion before hepatectomy for hepatocellular carcinoma.

Microvascular invasion (MVI) is difficult to detect before resection of hepatocellular carcinoma (HCC).

Intraoperative indocyanine green fluorescent imaging for prevention of bile leakage after hepatic resection

BACKGROUND Bile leakage is a common complication of hepatectomy, and is associated with an increase in sepsis and liver failure. There are no standard preventive methods against bile leakage after hepatic surgery. The aim of the present randomized clinical trial was to evaluate the application of indocyanine green (ICG) fluorescent cholangiography for preventing postoperative bile leakage. METHODS 102 patients who underwent hepatic resection without biliary reconstruction were divided into 2 groups. The control group (n = 50) underwent a leak test with ICG dye alone, and the experimental group underwent a leak test with ICG dye, followed by ICG fluorescent cholangiography using the Photodynamic Eye (PDE group, n = 52). RESULTS Among 42 patients with fluorescence in the PDE group, 25 patients had insufficient closure of bile ducts on the cut surface of the liver, which were closed by suture or ligation. There were 5 patients who developed postoperative bile leakage in the control group versus no bile leakage in the PDE group (10% vs 0%, P = .019). CONCLUSION ICG fluorescent cholangiography could detect insufficiently closed bile ducts that could not be identified by a standard bile leak test. ICG fluorescent cholangiography may have useful potential for prevention of bile leakage after hepatic resection.

Hepatic resection for hepatocellular carcinoma in the elderly

Aging of the population has significantly increased the number of elderly patients undergoing surgery for hepatocellular carcinoma (HCC). We aimed to compare the results of hepatectomy for HCC in patients ≥70 years old with those for younger patients.

Risk factors and outcome of early recurrence after resection of small hepatocellular carcinomas

BACKGROUND This study aimed to clarify risk factors for early recurrence and examine the subsequent outcome in patients undergoing potentially R0 resection of small hepatocellular carcinomas (HCCs) (<or=2 cm in greatest dimension). METHODS Eighty-nine patients were divided into 2 groups as follows: 26 patients suffering from recurrence within 2 years of surgery (early recurrence group) and 63 patients who were disease-free for at least 2 years (disease-free 2Y group). RESULTS Only 7 of 63 patients (11%) from the group that was disease-free for at least 2 years died during the 5-year period after surgery, whereas 13 of 26 patients (50%) from the early recurrence group died. Multivariate analysis showed that the preoperative maximum removal rate of technetium-99m-diethylenetriamine pentaacetic acid-galactosyl human serum albumin and microscopic vascular invasion were independent predictors of the early recurrence of small HCC. CONCLUSIONS Early recurrence of small HCC is the leading cause of death within 5 years after R0 resection. The preoperative hepatic functional reserve influences early recurrence, even in patients with small tumors.

Timing of Resection for Synchronous Liver Metastases from Colorectal Cancer

BackgroundThis study aimed to compare the surgical outcome and long-term survival between simultaneous and delayed resection of liver metastases from colorectal cancer (LM), and to identify the factors influencing hepatic disease-free survival in patients with synchronous LM.MethodsSeventy-four patients with LM were divided into two groups, i.e., 32 patients who underwent hepatectomy at the time of colorectal surgery (simultaneous group) and 42 patients who underwent delayed liver resection (delayed group).ResultsThe hepatic disease-free survival rates of patients from the delayed group with either ≥3 or <3 liver metastases were significantly better than that of the simultaneous group. Multivariate analysis showed that simultaneous resection was one of three independent prognostic indicators with an influence on hepatic disease-free survival. In 13 of the 42 (31%) patients from the delayed group, new metastatic lesions were found in the same and/or different segments after re-evaluation during the interval between operations. These patients had a higher incidence of poorly differentiated or mucinous adenocarcinoma, shorter interval between procedures, and larger tumors than patients without tumor progression.ConclusionsTumor progression could be recognized and occult metastases were detected during the interval between operations. Delayed resection of synchronous LM may be useful to reduce the risk of rapid recurrence in the remnant liver. Patients with poorly differentiated or mucinous adenocarcinoma and those with larger tumors who undergo delayed liver resection should receive neoadjuvant chemotherapy during the interval between operations.

Clinicopathologic characteristics of patients with non-B non-C hepatitis virus hepatocellular carcinoma after hepatectomy.

BACKGROUND A substantial population of hepatocellular carcinoma (HCC) patients is negative for markers of hepatitis B virus and hepatitis C virus (HCV) infection (non-B non-C hepatitis virus [NBC]). METHODS Clinicopathologic data and outcomes were compared retrospectively for HCC patients with hepatitis B virus, HCV, and NBC who had undergone hepatectomy. RESULTS The TNM stage was significantly higher, and the prevalence of cirrhosis was significantly lower, in the NBC group compared with the HCV group. Among patients with a maximum tumor diameter of 5 cm or less, the survival rates were significantly higher in the NBC group than in the HCV group. Multivariate analysis revealed that preoperative serum des-gamma-carboxy prothrombin (DCP) level was a prognostic factor for survival in NBC-HCC patients. The DCP/tumor size ratio was significantly higher in NBC-HCC patients with normal liver histology than in patients with hepatitis or cirrhosis. CONCLUSIONS NBC-HCC patients had more advanced tumors compared with HCV-HCC patients, but significantly higher survival rates. Measurement of DCP potentially is significant for early diagnosis of NBC HCC, which may increase the chance of curative therapy without recurrence.

Dexamethasone inhibits the induction of iNOS gene expression through destabilization of its mRNA in proinflammatory cytokine-stimulated hepatocytes.

In the inflamed liver, proinflammatory cytokines including TNF-&agr;, IL-1&bgr;, and IFN-&ggr; stimulate the induction of iNOS gene expression, leading to excess production of NO and resulting in liver injury. The induction of iNOS is regulated by transactivation of the iNOS promoter with transcription factors such as nuclear factor &kgr;B and by posttranscriptional modifications such as mRNA stabilization. The synthetic glucocorticoid dexamethasone has been reported to inhibit iNOS induction, which may contribute to its inflammation-reducing effects. The objective was to investigate the mechanisms involved in the down-regulation of iNOS gene expression by dexamethasone. Primary cultured rat hepatocytes were treated with IL-1&bgr; (1 nM) in the presence or absence of dexamethasone. The induction of iNOS and its signal were analyzed. Dexamethasone (10-250 nM) inhibited the expression of iNOS mRNA and protein dose and time dependently, resulting in decreases in NO production. However, dexamethasone did not inhibit the up-regulation of type I IL-1 receptor stimulated by IL-1&bgr;. Dexamethasone also had no effect on the degradation of I&kgr;B proteins and on the activation of nuclear factor &kgr;B. Transfection experiments with iNOS promoter-luciferase constructs revealed that dexamethasone had no effect on the transactivation of the iNOS promoter but decreased the stabilization of iNOS mRNA. In support of the latter observation, dexamethasone inhibited the expression of an iNOS gene antisense transcript, which stabilizes iNOS mRNA by interacting with its 3&vprime;-untranslated region and 3&vprime;-untranslated region-binding proteins. Dexamethasone may inhibit the induction of iNOS gene expression at the step of mRNA stabilization rather than promoter activation and may provide useful therapeutic effects in iNOS induction involved in liver injuries.ABBREVIATIONS-iNOS-inducible nitric oxide synthase; IL-1&bgr;-interleukin 1&bgr;; NF-&kgr;B-nuclear factor &kgr;B; IL-1RI-type I interleukin 1 receptor; 3&vprime;-UTR-3&vprime;-untranslated region

Protective effect of FR183998, a Na+/H+ exchanger inhibitor, and its inhibition of iNOS induction in hepatic ischemia-reperfusion injury in rats.

Recent evidence indicates that inhibition of the Na+/H+ exchanger improves heart and brain injuries induced by I/R. Studies were performed to investigate whether FR183998, a Na+/H+ exchanger inhibitor, has protective effects on hepatic I/R injury in rats. Male Sprague-Dawley rats were subjected to 70% hepatic ischemia by occluding the hepatic artery, portal vein, and bile duct associated with the left and median liver lobes with a microvascular clip for 2 h. FR183998 (1 mg/kg) was administered i.v. 10 min before the hepatic ischemia. Hepatic I/R increased the serum levels of aspartate transaminase, alanine transaminase, and lactate dehydrogenase, which peaked at 9 h after reperfusion. FR183998 reduced these injury markers and recovered liver functions. Histopathologic analysis revealed that FR183998 prevented the incidences of hepatic necrosis, apoptosis, and neutrophil infiltration at 6 and 9 h (P < 0.05). FR183998 reduced the increases in proinflammatory cytokines such as TNF-&agr; (1 - 6 h), IL-6 (1 - 12 h), interferon-&ggr; (6 - 12 h), IL-1&bgr; (1 - 3 h), and cytokine-induced neutrophil chemoattractant 1 (1-3 h), but enhanced the anti-inflammatory cytokine IL-10 (1 h). FR183998 inhibited the hepatic I/R-induced activation of the transcription factor nuclear factor-&kgr;B at 1 to 6 h and reduced the induction of iNOS at 6 to 12 h, followed by inhibition of nitric oxide production. Furthermore, FR183998 decreased the expression of the iNOS gene antisense transcript, which is involved in the stability of iNOS messenger RNA, at 9 to12 h in the liver of hepatic I/R rats. These results demonstrate that FR183998 reduces the induction of proinflammatory cytokines and iNOS at least in part through inhibition of nuclear factor-&kgr;B activation and iNOS antisense transcript expression, thereby preventing hepatic I/R injury.ABBREVIATIONS - NHE-Na+/H+ exchanger; I/R-ischemia/reperfusion; NO-nitric oxide; iNOS-inducible nitric oxide synthase; TNF-&agr;-tumor necrosis factor-&agr;; CINC-1-cytokine-induced neutrophil chemoattractant 1; IFN-&ggr;-interferon-&ggr;; IL-interleukin; NF-&kgr;B-nuclear factor-&kgr;B

Usefulness of Tc-99m-GSA scintigraphy for liver surgery

Postoperative mortality remains high after hepatectomy compared with other types of surgery in patients who have cirrhosis or chronic hepatitis. Although there are several useful perioperative indicators of liver dysfunction, no standard markers are available to predict postoperative liver failure in patients with hepatocellular carcinoma (HCC) undergoing hepatectomy. The best preoperative method for evaluating the hepatic functional reserve of patients with HCC remains unclear, but technetium-99m diethylenetriamine pentaacetic acid galactosyl human serum albumin (99mTc-GSA) scintigraphy is a candidate. 99mTc-GSA is a liver scintigraphy agent that binds to the asialoglycoprotein receptor, and can be used to assess the functional hepatocyte mass and thus determine the hepatic functional reserve in various physiological and pathological states. The maximum removal rate of 99m Tc-GSA (GSA-Rmax) calculated by using a radiopharmacokinetic model is correlated with the severity of liver disease. There is also a significant difference of GSA-Rmax between patients with chronic hepatitis and persons with normal liver function. Regeneration of the remnant liver and recurrence of hepatitis C virus infection in the donor organ after living donor liver transplantation have also been investigated by 99mTc-GSA scintigraphy. This review discusses the usefulness of 99mTc-GSA scintigraphy for liver surgery.