Toru Mizuguchi

Long-term and perioperative outcomes of laparoscopic versus open liver resection for hepatocellular carcinoma with propensity score matching: A multi-institutional Japanese study

The aim of this study was to compare the long‐term outcomes and perioperative outcomes of laparoscopic liver resection (LLR) with those of open liver resection (OLR) for hepatocellular carcinoma (HCC) between well‐matched patient groups.

Alteration of expression of liver-enriched transcription factors in the transition between growth and differentiation of primary cultured rat hepatocytes

In the present study, we showed the role of the liver‐enriched transcription factors in the transition during which proliferating hepatocytes become quiescent. We used primary rat hepatocytes cultured in modified L‐15 medium. The cells proliferated and, after the addition of 2% dimethyl sulfoxide (DMSO) from day 4, they stopped growing and gradually differentiated. During hepatic proliferation, expression of hepatocyte nuclear factors (HNF)1α, HNF4, C/EBPα, and C/EBPβ mRNAs was depressed, whereas that of HNF3α and HNF3β transcripts was enhanced. After the addition of DMSO, the expression of HNF1α, HNF3γ, and HNF4 returned to the level in isolated cells and HNF1β mRNA expression gradually increased. However, expression of C/EBPα and C/EBPβ mRNAs was partially recovered. The mitoinhibitory agents, IL‐1β, IL‐6, TGF‐β, and activin A, were examined to determine whether they could induce differentiation of proliferating hepatocytes as shown in cells treated with DMSO. Although these factors inhibited cell growth, the cells did not differentiate. The expression pattern of HNF3γ mRNA was quite different in the cells cultured with DMSO and those cultured with cytokines. Therefore, hepatic differentiation requires not only inhibition of DNA synthesis but also induction of appropriate transcription factors. Thus, expression of HNF3γ, C/EBPα, and C/EBPβ may be necessary for hepatocytes to acquire highly differentiated functions in addition to coexpression of certain amounts of transcripts of HNF1α, HNF1β, HNF3α, HNF3β, and HNF4 as well as suppression of C/EBPδ. J. Cell. Physiol. 174:273–284, 1998.

Gene expression analysis identifies over-expression of CXCL1, SPARC, SPP1, and SULF1 in gastric cancer

To elucidate gene expression signatures associated with gastric carcinogenesis, we performed a genome‐wide expression analysis of 46 Finnish and 20 Japanese gastric tissues. Comparative analysis between Finnish and Japanese datasets identified 58 common genes that were differentially expressed between cancerous and non‐neoplastic gastric tissues. Twenty‐six of these genes were up‐regulated in cancer and 32 down‐regulated. Of these genes, 64% were also differentially expressed in another unrelated publicly available dataset. The expression levels of four of the up‐regulated genes, CXCL1, SPARC, SPP1 and SULF, were further analyzed in 82 gastric tissues using quantitative real‐time RT‐PCR. This analysis validated the results from the microarray analysis as the expression of these four genes was significantly higher in the cancerous tissue compared with the normal tissue (fold change 3.4–8.9). Over‐expression of CXCL1 also positively correlated with improved survival. To conclude, irrespective of the microarray platform or patient population, a common gastric cancer gene expression signature of 58 genes, including CXCL1, SPARC, SPP1, and SULF, was identified. These genes represent potential biomarkers for gastric cancer.

Long-term and perioperative outcomes of laparoscopic versus open liver resection for colorectal liver metastases with propensity score matching: A multi-institutional Japanese study

The aim of the present study was to clarify the surgical outcome and long‐term prognosis of laparoscopic liver resection (LLR) compared with conventional open liver resection (OLR) in patients with colorectal liver metastases (CRLM).

Growth and maturation of small hepatocytes.

Proliferation of adult rat hepatocytes is observed in serum‐free Dulbeccos modified Eagles medium (DMEM) supplemented with 10 mmol/L nicotinamide and 10 ng/mL epidermal growth factor (EGF). The proliferating cells are mainly mononucleate and form small cell colonies surrounded by mature hepatocytes. Although these cells in focal colonies have a less‐differentiated appearance, immunocytochemically and ultrastructurally they possess hepatic characteristics. The size of small hepatocytes is one‐third to half that of mature hepatocytes. Therefore, we call the cells forming a colony, small hepatocytes. The small hepatocytes can be subcultured for several passages. Furthermore, the cells are rich in the supernatant following 50 g centrifugation for 1 min after collagenase liver perfusion. When the cells are cultured in DMEM supplemented with 10% foetal bovine serum, 10 mmol/L nicotinamide, 1 mmol/L ascorbic acid 2‐phosphate, 10 ng/mL EGF and 1% dimethyl sulphoxide, each small hepatocyte can clonally proliferate for more than 3 months. A small hepatocyte divides to form a colony and the number of cells reaches more than 100 within 20 days. With time in culture, cells with a large cytoplasm appear within a colony. They have many mitochondria and large peroxisomes with crystalline nucleoids and are typical, mature hepatocytes. Immunoreactivity to connexin 32 and well‐developed bile canaliculus structures are often observed in the cell‐cell borders. Thus, we suggest that small hepatocytes may be considered to be ‘committed progenitor cells’ that can further differentiate into mature hepatocytes.

Proliferation of hepatocyte progenitor cells isolated from adult human livers in serum-free medium.

Rat small hepatocytes (SHs) are committed progenitor cells that can differentiate into mature hepatocytes and can selectively proliferate in serum-free medium when they are cultured on hyaluronic acid (HA)-coated dishes. In this study we examined the separation of human SHs from adult human livers. We obtained liver tissues from the resected liver of 16 patients who underwent hepatic resections. Extracted liver specimens were clearly separate from the tumor regions with sufficient margins. Hepatic cells were isolated using the modified method of two-step collagenase perfusion. A low-speed centrifugation was performed and cells in the supernatant were finally cultured on HA-coated dishes in serum-free DMEM/F12 medium including nicotinamide, EGF, and HGF. Small-sized hepatocytes selectively proliferated to form colonies and many colonies continued growing for more than 3 weeks. The average number of cells in a colony was 38.6 ± 18.0, 79.0 ± 54.0, and 101.5 ± 115.7 at day 7, 14, and 21, respectively. About 0.04% of plated cells could form an SH colony. Immunocytochemistry showed that the cells forming a colony were positive for albumin, transferrin, keratin 8, and CD44. The results of RT-PCR showed that colony-forming cells expressed albumin, transferrin, α1-antitrypsin, fibrinogen, glutamine synthetase, many cytochrome P450s, and liver-enriched transcription factors (HNF3α, HNF4α, C/EBPα, and C/EBPβ). Furthermore, the cells expressed not only the genes of hepatic differentiated functions but also those of both hepatic stem cell marker (Thy1.1, EpCAM, AFP) and SH marker (CD44, D6.1A, BRI3). Albumin secretion into culture medium was also observed. Our results demonstrate the existence of hepatocyte progenitor cells in human adult livers, and the cells can grow in a serum-free medium on HA-coated dishes. Human SHs may be a useful source for cell transplantation as well as pharmaceutical and toxicological investigations.

Laminin α5 mediates ectopic adhesion of hepatocellular carcinoma through integrins and/or Lutheran/basal cell adhesion molecule

Laminins are a diverse group of alpha/beta/gamma heterotrimers formed from five alpha, three beta and three gamma chains; they are major components of all basal laminae (BLs). One laminin chain that has garnered particular interest due to its widespread expression pattern and importance during development is laminin alpha 5. Little is known, however, about the expression and function of laminins containing the alpha 5 chain in human hepatocellular carcinoma (HCC). Here, using a specific antibody, we examined the expression of laminin alpha 5 in normal liver and in HCCs. In normal liver, although laminin alpha 5 was observed in hepatic BLs underlying blood vessels and bile ducts, it was absent from the parenchyma, which may be the origin of HCC. On the other hand, laminin alpha 5 deposition was observed throughout all HCCs tested, regardless of tumor grade. In well-differentiated HCCs, it localized along the trabecules of the tumor. In poorly-differentiated HCCs, it was present in surrounding tumor nodules. In HCC cell lines, laminin alpha 5 heterotrimerized with beta and gamma chains and was secreted into the culture media. To attempt to understand the function of laminins containing alpha 5, the expression of its receptors in HCCs was also determined. In this regard, alpha 3 beta 1/alpha 6 beta 1 integrins and Lutheran/basal cell adhesion molecule (Lu/B-CAM) were expressed in HCC cells. In vitro studies showed that HCC cells readily attached to laminin containing the alpha 5 chain, more so than did primary hepatocytes. In addition to alpha 3 beta 1/alpha 6 beta 1 integrins and Lu/B-CAM, laminin alpha 5 was recognized by integrin alpha 1 beta 1, which also was expressed in HCC cells. These results suggest that laminins containing alpha 5 serve as functional substrates regulating progression of HCC.

Current Status of Surgery for Pancreatic Cancer

Background: In Japan the annual incidence of pancreatic cancer has increased over the last decade, but no advancement has been made in the long-term prognosis after resection. The significant differences in the surgical procedures between Western countries and Japan have been discussed. Therefore, an adequate comparison and analysis of the data from Japan, Europe and the USA is required. This review evaluates many important published reports from Japan which influence surgical procedure. Methods: Several important highlights and controversies regarding the concept of surgical treatment and surgical procedure are discussed comparing the results in Japan with those in Western countries. Results: No significant difference in diagnostic strategy using various imaging methods was observed between Japan and Europe. The stage classification for pancreatic cancer by the Japanese Pancreatic Society (JPS) seems to be superior to others, because the results on long-term prognosis after pancreatectomy of cases with pancreatic head cancer, diagnosed as tubular adenocarcinoma, has been arranged logically. Pancreatectomy with extended radical dissection is recommended in Japan, but several clinical studies from Europe and the USA suggest that this is ineffective. The basic concepts of this controversy have recently come closer altogether. Scientific clinical trials for instance on the necessity of adjuvant treatment, etc., are now on-going. Conclusion: The characteristics on diagnosis and treatment of pancreatic cancer in Japan are described. The JPS registration system for pancreatic cancer can provide much more information, i.e. dependency on diagnostic methods, highly frequent sites of lymph node and of distant metastases, the prognosis of small pancreatic cancers, etc. The indication for any surgical treatments should be limited to cases with the possibility of cancer free margins.

Laparoscopic hepatectomy: A systematic review, meta-analysis, and power analysis

PurposeA previous meta-analysis study demonstrated that bleeding and the duration of the hospital stay following laparoscopic hepatectomy (Lap) were significantly smaller and shorter, respectively, than for patients undergoing an open approach (Op). The aim of the present study was to re-evaluate perioperative variables and adverse outcomes in patients undergoing Lap versus (vs) Op after 2000.MethodsA PubMed and Ovid Medline search identified clinical studies that compared the outcomes of Lap vs Op patients after 2000. A meta-analysis and power analysis were performed.ResultsOperative time was not significantly different between the two approaches (95% confidence interval [CI]: −0.063 to 0.992). Patient bleeding in the Lap group was significantly lower than in the Op group (95% CI: −1.027 to −0.390). Complications with Lap patients were significantly less frequent (95% CI: 0.231–0.642), and the duration of the hospital stay for Lap patients was significantly shorter (95% CI: −0.950 to −0.530) than for Op patients. Only one paper presented 80% power with 0.05 α-errors in all four outcomes, whereas four studies did not have sufficient statistical power.ConclusionsThe clinical benefits of Lap include a smaller incidence of complications and a shorter duration of hospital stay at the current time. Several studies had too few cases to sufficiently evaluate these factors, although other studies were appropriately analyzed.

Serum Hyaluronate Level for Predicting Subclinical Liver Dysfunction after Hepatectomy

The serum hyaluronate (HA) level reflects sinusoidal endothelial cell function correlated with liver function. We have reviewed multiple liver function indicators from 37 patients who underwent hepatectomy for various liver diseases. The serum HA level was well correlated with the indocyanine green retention rate at 15 minutes (ICGR15), lectin-cholesterol (LCAT), hepatocyte growth factor (HGF), liver uptake ratio of technetium-99m galactosyl human serum albumin (99mTc-GSA) at 15 minutes (HH15), prealbumin, and hepatic uptake ratio of 99mTc-GSA at 15 minutes (LHL15). In addition, the model for end-stage liver disease (MELD) score at 7 days after operation was well correlated with serum HA, ICGR15, HH15, and LHL15. In patients who showed serum an HA level of = 100 ng/ml before hepatectomy, the MELD score had significantly deteriorated by 7 days after hepatectomy. Of the 20 patients who showed a serum HA level < 100 ng/ml before hepatectomy, 11 had high serum HA after hepatectomy. The bilirubin level 7 days after operation in this group was much higher than that for patients who maintained a serum HA level < 100 ng/ml after hepatectomy. In addition, the serum HGF level before hepatectomy in this group was significantly lower. We concluded that the serum HA level is a reliable indicator when evaluating liver function and predicting liver dysfunction after hepatectomy. Furthermore, patients with a significantly low HGF level who have a normal HA level are susceptible to liver dysfunction after hepatectomy.