Reduction in disialyl-T antigen levels in mice deficient for both St6galnac3 and St6galnac4 results in blood filling of lymph nodes.

Sialic acid (SA) is present at the terminal ends of carbohydrate chains in glycoproteins and glycolipids and is involved in various biological phenomena. The biological function of the disialyl-T (SAα2-3Galß1-3(SAα2-6)GalNAcα1-O-Ser/Thr) structure is largely unknown. To elucidate the role of disialyl-T structure and determine the key enzyme from the N-acetylgalactosaminide α2,6-sialyltransferase (St6galnac) family involved in its in vivo synthesis, we generated St6galnac3- and St6galnac4-deficient mice. Both single-knockout mice developed normally without any prominent phenotypic abnormalities. However, the St6galnac3::St6galnact4 double knockout (DKO) mice showed spontaneous hemorrhage of the lymph nodes (LN). To identify the cause of bleeding in the LN, we examined podoplanin, which modifies the disialyl-T structures. The protein expression of podoplanin in the LN of DKO mice was similar to that in wild-type mice. However, the reactivity of MALII lectin, which recognizes disialyl-T, in podoplanin immunoprecipitated from DKO LN was completely abolished. Moreover, the expression of vascular endothelial cadherin was reduced on the cell surface of high endothelial venule (HEV) in the LN, suggesting that hemorrhage was caused by the structural disruption of HEV. These results suggest that podoplanin possesses disialyl-T structure in mice LN and that both St6galnac3 and St6galnac4 are required for disialyl-T synthesis.

[A Case of Gastric Cancer Invading the Pancreas with Pathological Complete Response Treated by Preoperative S-1 plus Oxaliplatin Therapy].

A 66-year-old man with severe anemia was diagnosed with gastric cancer. CT examination revealed primary gastric tumor, which involved the pancreas body, with regional lymph nodes that were enlarged(T4b[panc], cN2, cM0, cStage ⅣA). He received three courses of preoperative S-1 plus oxaliplatin therapy. Primary tumor and metastatic lymph nodes were reduced remarkably. We performed a curative distal gastrectomy(D2)without pancreas resection. Histopathological examination revealed Grade 3 pathological complete response in both primary tumor and metastatic lymph nodes.

Data on long-term survival of the NOD/Shi-scid IL-2Rγnull (NOG) mouse in two facilities.

The objective of this study was to investigate an appropriate observation period for an evaluation of tumorigenicity in NOD/Shi-scid IL-2 Rγnull (NOG) mice. At SNBL, 19 male and 19 female NOG mice were observed the general condition from 7 weeks old up to 68 weeks old and at FBRI, 7 male and 16 female NOG mice were observed the general condition throughout the lifespan from 7 weeks old. The survival rate started to decline rapidly around 54 to 56 weeks of age in both facilities without a facility difference. Based on these survival data, it seems reasonable to terminate a tumorigenicity study at 52 weeks of age.

Monkeys mutant for PKD1 recapitulate human autosomal dominant polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) caused by PKD1 mutations is one of the most common hereditary disorders. However, the key pathological processes underlying cyst development and exacerbation in pre-symptomatic stages remain unknown, because rodent models do not recapitulate critical disease phenotypes, including disease onset in heterozygotes. Here, using CRISPR/Cas9, we generate ADPKD models with PKD1 mutations in cynomolgus monkeys. As in humans and mice, near-complete PKD1 depletion induces severe cyst formation mainly in collecting ducts. Importantly, unlike in mice, PKD1 heterozygote monkeys exhibit cyst formation perinatally in distal tubules, possibly reflecting the initial pathology in humans. Many monkeys in these models survive after cyst formation, and cysts progress with age. Furthermore, we succeed in generating selective heterozygous mutations using allele-specific targeting. We propose that our models elucidate the onset and progression of ADPKD, which will serve as a critical basis for establishing new therapeutic strategies, including drug treatments.

[A Case of Intraductal Pancreatic Mallignant Tumor with Difficulty in Differentiating between IPMN and ITPN].

The patient was a male in his early 60s. Diabetes had aggravated 6 months earlier, and the patient was referred to our hospital for close examination. On contrast CT, enhanced mass shadows filling the lumen of the main pancreatic duct, which was dilated throughout the pancreas, were observed, and the mass was diagnosed as an adenocarcinoma on EUS-FNA. Based on these findings, main-duct IPMN was suspected and total pancreatectomy was performed. On macroscopic observation of the resected specimen, outgrowth of a solid tumor was observed in the main pancreatic duct, whereas only low-level mucus retention was noted in the pancreatic duct. Histopathological examination revealed a papillary/tubular tumor growth, suggesting interstitial infiltration throughout the pancreas. On immunostaining, the tumor was partially positive for MUC5AC, based on which the patient was diagnosed with an intraductal pancreatic mallignant tumor, with difficulty in differentiating between IPMC and ITPC. Clinicopathologically, many aspects regarding ITPN remain unclear. Further accumulation of such cases and investigation of the tumor pathology are necessary.

Preoperative evaluation of accessory hepatic ducts by drip infusion cholangiography with CT.

BACKGROUND: This retrospective study aimed to investigate the incidence of each type of accessory hepatic duct by drip infusion cholangiography with CT (DIC-CT). METHODS: Five hundred sixty nine patients who underwent preoperative DIC-CT and laparoscopic cholecystectomy were reviewed. Accessory hepatic ducts were classified as follows: type I (accessory hepatic ducts that merged with the common hepatic duct between the confluence of the right and left hepatic ducts and the cystic duct confluence), type II (those that merged with the common hepatic duct at the same site as the cystic duct), type III (those that merged with the common bile duct distal to the cystic duct confluence), type IV (the cystic duct merged with the accessory hepatic duct), and type V (accessory hepatic ducts that merged with the common hepatic or bile duct on the left side). RESULTS: Accessory hepatic ducts were observed in 50 patients. Type I, II, III, IV, and V accessory hepatic ducts were detected in 32, 3, 1, 11, and 3 patients, respectively. Based on their drainage areas, the accessory hepatic ducts were also classified as follows: a posterior branch in 22 patients, an anterior branch in 9 patients, a combination of posterior and anterior branches in 16 patients, a left-sided branch in 2 patients, and a caudate branch in 1 patient. None of the patients with accessory hepatic ducts suffered bile duct injuries. CONCLUSION: There are a number of variants of the accessory hepatic duct. DIC-CT is useful to detect the accessory hepatic duct.

[Metastatic Pancreatic Carcinoma from Renal Cell Carcinoma Indistinguishable from Primary Pancreatic Endocrine Tumor - A Case Report].

A hypervascularized tumor was detected in a 65-year-old man who had underwent a nephrectomy for a right renal cell carcinoma at the age of 55 years. We diagnosed the tumor as a non-functioning pancreatic neuroendocrine tumor or a metastatic tumor from the renal cell carcinoma. We performed distal pancreatectomy with splenectomy and lymph node dissection. The tumor was histopathologically diagnosed as metastatic renal cell carcinoma.

Idiopathic pulmonary hemosiderosis complicated by Down syndrome.

We report the case of a 9-year-old girl with Down syndrome (DS) diagnosed with idiopathic pulmonary hemosiderosis (IPH). Although acute pneumonia complicated by hemolytic anemia was suspected, IPH was finally diagnosed on bronchoscopy. Treatment with prednisolone achieved good clinical response. An association between IPH and DS was not able to be identified, but immunological issues in DS may contribute to the onset of IPH. Recurrent and intractable respiratory symptoms with marked infiltrative shadows in the bilateral lungs and complicated by severe anemia in patients with DS should suggest IPH.

True left-sided gallbladder with variations of bile duct and cholecystic vein.

A left-sided gallbladder without a right-sided round ligament, which is called a true left-sided gallbladder, is extremely rare. A 71-year-old woman was referred to our hospital due to a gallbladder polyp. Computed tomography (CT) revealed not only a gallbladder polyp but also the gallbladder located to the left of the round ligament connected to the left umbilical portion. CT portography revealed that the main portal vein diverged into the right posterior portal vein and the common trunk of the left portal vein and right anterior portal vein. CT cholangiography revealed that the infraportal bile duct of segment 2 joined the common bile duct. Laparoscopic cholecystectomy was performed for a gallbladder polyp, and the intraoperative finding showed that the cholecystic veins joined the round ligament. A true left-sided gallbladder is closely associated with several anomalies; therefore, surgeons encountering a true left-sided gallbladder should be aware of the potential for these anomalies.

Effect of cod liver oil supplementation on the stearoyl-CoA desaturase index in obese children: a pilot study.

To investigate the effects of n-3 polyunsaturated fatty acids on stearoyl-CoA desaturase (SCD) activity, we treated 10 obese children (mean age: 12.9 years) with cod liver oil once daily for 12 weeks. The effects of cod liver oil supplementation on SCD activity, as estimated by the palmitoleate/palmitate ratio, depended on the docosahexaenoic acid (DHA) contents at baseline. Baseline DHA contents were negatively correlated with baseline SCD activity. After the treatment, baseline DHA contents were found to be significantly associated with the reduction of SCD activity. Cod liver oil supplementation may be a complementary treatment for obese children with low baseline contents of DHA.

Rapid development of atherosclerosis in the world's smallest Microminipig fed a high-fat/high-cholesterol diet.

AIM: Experimental studies of human atherogenesis require an appropriate animal model that mimics human physiology and pathology. Because swine physiology is similar to human physiology, we developed a hyperlipidemia-induced atherosclerosis model using the recently developed world's smallest Microminipig(TM). METHODS: These animals weigh only 5kg at 3months of age, much smaller than any other miniature pig. We found that the administration of a high-fat/high-cholesterol diet containing at least 0.2% cholesterol without cholic acid for as little as eight weeks induces hypercholesterolemia and subsequent atherosclerosis in these animals. RESULTS: The serum levels of low-density lipoprotein cholesterol(LDL-C) and the percent distribution of cholesterol in the LDL fractions were markedly increased. The hepatic expression of LDL receptor and hydroxymethylglutaryl-CoA reductase was coordinately decreased. The cholesteryl ester transfer protein activity, which plays a role in reverse cholesterol transport, was detected in the serum of the Microminipigs. Niemann-Pick C1-like 1 protein was expressed in both the liver and small intestine; however, hepatic apoB mRNA editing enzyme was not expressed. As in humans, and in contrast to that observed in mice, most of the hepatic lipase activity was localized in the liver. These results suggest that the hyperlipidemia-induced gene expression profile linked to cholesterol homeostasis and atherogenesis is similar in Microminipigs and humans. CONCLUSION: We conclude that the characteristics of the Microminipig, including its easy handling size, make it an appropriate model for studies of atherosclerosis and related conditions.

Plasma hepatocyte growth factor elevation may be associated with early metastatic disease in primary lung cancer patients.

PURPOSE: Hepatocyte growth factor (HGF), a ligand of the c-met proto-oncogene, exhibits activating effects on human lung cancer both in vitro and in vivo. However, few studies have reported the correlations between concentration changes of blood HGF and postsurgical prognosis. METHODS: We evaluated whether surgery-related blood HGF elevation has prognostic significance in patients with surgically resected non-small cell lung cancer. We examined blood HGF concentration, c-met expression, and postoperative prognosis of 25 cases of primary resected, non-small cell lung cancer. RESULTS: We divided the patients into 2 groups according to receiver operating characteristics curve analysis using 7.2 ng/mL as the cut-off value of blood HGF concentration. Survival curve analysis revealed that patients with a high level of HGF (over the cutoff value) exhibited a poor prognosis of metastatic disease, compared to those in the low-level group after curative surgery (log rank test, P = 0.020; Wilcoxon test, P = 0.016). CONCLUSION: Elevation of HGF in plasma may be an important prognostic factor for early metastatic disease in patients with primary lung cancer. Moreover, inhibition of HGF elevation may have therapeutic effects on early distant metastasis of lung cancer.

[Incidental detection of thymic mucoepidermoid carcinoma with massive calcification; report of a case].

31-year-old man who complained of chest pain due to chest wall wound caused by tumble. He took chest X-ray examination and suggested of having mediastinal tumor. Computed tomography (CT) scan revealed 3.0 cm mass in diameter with irregular shaped calcification and lobulated cystic lesion (size : 21 x 17 x 41 mm). The mediastinal tumor was totaly removed by surgical operation. No invasive portion was observed between mediastinal pleura and solid tumor. Pathological examination revealed that tumor was composed of mucoepidermoid carcinoma of the thymus and postoperative adjuvant radiotherapy was performed. The patient have survived without disease for 3 years after surgery.

BCL2L2 is a probable target for novel 14q11.2 amplification detected in a non-small cell lung cancer cell line.

Amplification of chromosomal DNA is thought to be one of the mechanisms that activates cancer-related genes in tumors. In a previous genome-wide screening of DNA copy number aberrations in a panel of non-small cell lung cancer (NSCLC) cell lines using an in-house bacterial artificial chromosome-based array, we identified a novel amplification at 14q11.2 in HUT29 cells derived from human lung adenocarcinoma. To identify the most likely target for the 14q11.2 amplification, we determined the extent of the amplicon by fluorescence in situ hybridization and then analyzed NSCLC cell lines for the expression levels of 28 genes present within the 1-Mb amplified region. Significant overexpression in the HUT29 cell line with amplification, relatively frequent overexpression in additional NSCLC cell lines compared with an immortalized normal lung epithelial cell line, and reported information about the function of each candidate gene prompted us to characterize the BCL2-like2 (BCL2L2) gene, a prosurvival member of the BCL2 family, as the most likely target for the 14q11.2 amplicon. Immunohistochemical analysis of 61 primary cases of lung adenocarcinoma demonstrated that BCL2L2 overexpression was significantly associated with tumor stage and differentiation status, and tended to be associated with a poorer prognosis. Downregulation of BCL2L2 expression using small interfering RNA dramatically inhibited the growth of HUT29 cells, but showed no effect on anticancer reagent-induced cell death of the same cell line. These findings demonstrate that overexpressed BCL2L2, through amplification or other mechanisms, promotes the growth of NSCLC, especially the adenocarcinoma subtype, and might be a therapeutic target.

Frequent silencing of the candidate tumor suppressor PCDH20 by epigenetic mechanism in non-small-cell lung cancers.

Protocadherins are a major subfamily of the cadherin superfamily, but little is known about their functions and intracellular signal transduction. We identified a homozygous loss of protocadherin 20 (PCDH20, 13q21.2) in the course of a program to screen a panel of non-small-cell lung cancer (NSCLC) cell lines (1 of 20 lines) for genomic copy number aberrations using an in-house array-based comparative genomic hybridization. PCDH20 mRNA was expressed in normal lung tissue but was not expressed in the majority of NSCLC cell lines without a homozygous deletion of this gene (10 of 19 lines, 52.6%). Expression of PCDH20 mRNA was restored in gene-silenced NSCLC cells after treatment with 5-aza 2'-deoxycytidine. The DNA methylation status of the PCDH20 CpG-rich region correlated inversely with the expression of the gene and a putative target region for methylation showed clear promoter activity in vitro. Methylation of this PCDH20 promoter was frequently observed in primary NSCLC tissues (32 of 59 tumors, 54.2%). Among our primary NSCLC cases, the methylated PCDH20 seemed to be associated with a shorter overall survival (P = 0.0140 and 0.0211 in all and stage I tumors, respectively; log-rank test), and a multivariate analysis showed that the PCDH20 methylation status was an independent prognosticator. Moreover, restoration of PCDH20 expression in NSCLC cells reduced cell numbers in colony formation and anchorage-independent assays. These results suggest that epigenetic silencing by hypermethylation of the CpG-rich promoter region of PCDH20 leads to loss of PCDH20 function, which may be a factor in the carcinogenesis of NSCLC.

Frequent silencing of DBC1 is by genetic or epigenetic mechanisms in non-small cell lung cancers.

Genome-wide screening of DNA copy number aberrations in 27 cell lines derived from non-small cell lung cancers (NSCLCs), using a custom-made comparative genomic hybridization (CGH)-array, identified a homozygous deletion of the deleted in bladder cancer 1 gene (DBC1) in one cell line. Homozygous deletion of DBC1, located at 9q33.1, was also observed in two of 53 primary NSCLC tumors examined. Moreover, 21 of the other 26 cell lines showed complete loss of DBC1 expression, although normal lung tissues express this gene, and treatment with 5-aza-2'-deoxycytidine restored expression of DBC1. Hypermethylation in part of a CpG island around the exon 1 of DBC1 has been reported in urothelial cancers, but the potential association between methylation and expression status was never clarified in that disease. In our experiments, a different part of the same CpG island showed promoter activity in vitro and was frequently methylated in our cell lines and primary tumors of NSCLC, where methylation status correlated inversely with gene expression. Among our primary NSCLC cases, methylation of the DBC1 promoter occurred more frequently in men, elderly patients and smokers than in women, younger patients and nonsmokers respectively, but it was not correlated with tumor stage or histology. Exogenous overexpression of DBC1 in NSCLC cell lines lacking its expression inhibited cell growth. Our results provide the first evidence that DBC1 is a likely tumor suppressor for NSCLC; silencing of the gene through homozygous deletion or methylation of its promoter region may be associated with progression of this disease.

Membrane cholesterol but not putative receptors mediates anandamide-induced hepatocyte apoptosis.

The endogenous cannabinoid anandamide, a lipid mediator, induces various physiologic events such as vascular relaxation, inhibition of gap-junctions formation, tumor proliferation, neurologic analgesia, and apoptosis. Although increased concentration of anandamide in plasma has been implicated in pathophysiologic states including endotoxin-induced hypotension, the effects of anandamide on hepatocytes still remain unclear. In this study, we present evidence that plasma anandamide concentration is highly increased in severe hepatitis and cirrhosis patients. In addition, concentrations of anandamide within the pathophysiologic range potently induced apoptosis of hepatoma cell line (Hep G2) and primary hepatocytes, suggesting a possible link between increased anandamide level and hepatocyte damage. Anandamide-induced cell death was preceded by G0/G1 cell-cycle arrest, activation of proapoptotic signaling (i.e., p38 MAPK and JNK), and inhibition of antiapoptotic signaling (i.e., PKB/Akt) pathways. Moreover, anandamide increased susceptibility to oxidative stress-induced hepatocyte damage. In this context, methyl-beta-cyclodextrin (MCD), a membrane cholesterol depletor, or mevastatin, an HMG-CoA reductase inhibitor, or N-acetyl cysteine, an antioxidant, potently inhibited the anandamide-induced proapoptotic events and cell death, whereas putative cannabinoid receptor antagonists did not exhibit an inhibitory effect on anandamide-induced cell death. Furthermore, binding assay using polymyxin beads revealed that anandamide could interact with cholesterol. In conclusion, our data suggest that cholesterol present in the cell membrane determines the fate of hepatocytes exposed to anandamide, possibly functioning as an anandamide receptor.