Risk factors for Clostridium difficile-associated diarrhea and the effectiveness of prophylactic probiotic therapy.

Measures for prevention of Clostridium difficile-associated diarrhea, a common nosocomial infection, in hospital settings are urgently needed. This study was conducted to identify the risk factors contributing to C. difficile-associated diarrhea and to evaluate the clinical benefit of probiotics in its prevention. The study included 2716 patients at least 20 years old who received an injected antibiotic at any time between February 2010 and February 2011; a total of 2687 patients (98.9%) were assigned to the non-C. difficile-associated diarrhea group, and 29 patients (1.1%) were assigned to the C. difficile-associated diarrhea group. Univariate analysis revealed a significant difference between the two groups for the following factors: antibiotic therapy for > or = 8 days; enteral nutrition; intravenous hyperalimentation; fasting; proton pump inhibitor use; H2 blocker use; and serum albumin < or = 2.9g/dL (p<0.05). Multivariate logistic regression analysis revealed a significant difference between the two groups for several factors. Antibiotic therapy for > or = 8 days, intravenous hyperalimentation, proton pump inhibitor use, and H2 blocker use were therefore shown to be risk factors for C. difficile-associated diarrhea. Prophylactic probiotic therapy was not shown to suppress the occurrence of C. difficile-associated diarrhea.

Risk factors contributing to urinary protein expression resulting from bevacizumab combination chemotherapy.

Proteinuria following administration of bevacizumab is reported to be a specific adverse effect, but the risk factors for proteinuria have not been elucidated. In this study, the risk factors for urinary protein expression resulting from bevacizumab combination chemotherapy were investigated. The subjects were 47 patients aged > or = 20 years who had received bevacizumab combination chemotherapy at Gifu Municipal Hospital between February 2010 and February 2011. A total of 13 patients were excluded based on exclusion criteria; of the remaining 34 patients, 24 (70.6%) were assigned to the urinary protein non-expression group, and 10 (29.4%) were assigned to the urinary protein expression group. The results of multivariate logistic regression analysis revealed a significant difference in systolic blood pressure (> or =130 mmHg) between the two groups (OR: 14.499, 95%CI: 1.326-158.577, p=0.028). This finding shows that systolic blood pressure (> or =130 mmHg) is a risk factor for urinary protein expression resulting from bevacizumab combination chemotherapy.

Inflammatory pseudotumor of the liver diagnosed by needle liver biopsy under ultrasonographic tomography guidance.

Inflammatory pseudotumor of the liver is a rare benign lesion, but exploratory laparotomy and a hepatectomy are often performed unnecessarily after various misdiagnoses, including liver abscess, hepatocellular carcinoma, metastatic liver tumor, and cholangiocarcinoma. We present a case of hepatic inflammatory pseudotumor in a 17-year-old man in whom diagnosis was confirmed by liver needle biopsy under ultrasonographic tomography (UST) guidance. He had complained of fever and right hypochondralgia 2 months after being operated for appendicitis. He was admitted to our hospital because of the persistence of these symptoms and the presence of a hepatic mass lesion detected by UST. He had hepatomegaly, with tenderness; leukocytosis and elevated erythrocyte sedimentation rate and C-reactive protein level were noted. UST showed a hypoechoic mass in the liver and pre-contrast computerized tomography (CT) revealed a low-density area with an ill defined margin, which was barely enhanced by the contrast medium. On the basis of the patient's clinical symptoms and the laboratory data and imaging studies, the presence of a liver abscess was suspected and antibiotics were administered. One month after the initiation of the antibiotic therapy, UST demonstrated that the portal vein had dilated serpiginously and penetrated into the mass. As the heterogeneous appearance displayed by post-enhanced CT indicated the need for a differential diagnosis of the hepatic mass lesion to rule out hepatocellular carcinoma, percutaneous needle biopsy was performed, under UST guidance. Histopathological examination demonstrated marked infiltration of plasma cells and fibrosis, findings which were consistent with those of hepatic inflammatory pseudotumor. There was a spontaneous reduction of the hepatic pseudotumor without continuous antibiotics and this reduction was documented on follow-up UST and CT.

Hepatocellular carcinoma with sarcomatous change arising in primary biliary cirrhosis.

We report an autopsy case of hepatocellular carcinoma (HCC) with sarcomatous change arising in the context of primary biliary cirrhosis (PBC) in a 79-year-old man. Primary biliary cirrhosis was diagnosed (stage I according to Scheuer's classification) by findings on blood biochemical analysis, laparoscopy, and liver biopsy at age 69 years. Five years later, (at age 74 years), a mass lesion was detected in the S6 region of the liver by abdominal ultrasonography, and target biopsy revealed well differentiated HCC. Blood biochemistry, ultrasonography, and computed tomography findings showed that the PBC had progressed to stage IV (cirrhotic stage). Percutaneous ethanol injection therapy (PEIT) was administered to the HCC several times over a 5-year period; however, the patient died of liver failure in February, 1994 (at age 79 years). Viral markers for hepatitis B and C were negative during the course, and hepatitis C virus RNA was not detected by polymerase chain reaction. Autopsy findings showed liver cirrhosis and diffuse involvement of spindle-shaped sarcomatoid cells in the liver, particularly in the S6 region, associated with several nodules of trabecular HCC cells. A zone of transition between the sarcomatoid cells and the trabecular hepatocellular carcinoma cells was observed. The sarcomatoid cells were diffusely disseminated in the peritoneal cavity and had metastasized to multiple organs. Immunohistochemically, the cells were positive for fibrinogen, as were the coexisting trabecular hepatocellular carcinoma cells. The HCC had been treated several times with PEIT. Of interest, PEIT may be an important factor in this type of tumor progression.

Liver cell adenoma in a young man with elevated serum PIVKA-II level.

A 21-year-old man was referred to our hospital because of a liver mass lesion detected by abdominal ultrasonography. He had received no hormonal treatment. Physical examinations revealed no abnormalities, and laboratory data, including hepatic function test results, were within normal ranges, with the exception of elevated levels of serum protein induced by vitamin K absence or antagonist (PIVKA)-II (2.2 AU/ml). Abdominal ultrasonography revealed a hyperechoic mass lesion measuring 10 x 10 cm, with hypoechoic areas located in the right posterior segment of the liver. A low-density area and a hypervascular area were detected in the right posterior segment of the liver by computed tomography and celiac angiography, respectively. As hepatocellular carcinoma could not be completely excluded, the tumor was resected. The tissue consisted of sheets of tumor cells with eosinophilic cytoplasm and round nuclei showing a thin trabecular pattern, and these histological findings indicated liver cell adenoma. After resection of the tumor, serum PIVKA-II returned to the normal level.

Ultrastructural study of TPA-induced cell motility: human well-differentiated rectal adenocarcinoma cells move as coherent sheets via localized modulation of cell-cell adhesion.

We previously found that 12-O-tetradecanoylphorbol-13-acetate (TPA)-enhanced invasion of Matrigel was associated with augmentation of cell motility but not with metalloproteinase activity in a highly metastatic variant (L-10) of human rectal adenocarcinoma cell line RCM-1. In a two-dimensional cell motility assay, TPA induced active L-10 cell locomotion with characteristic morphology; the cells moved outwards from the cell islands mainly as a localized coherent sheet of cells. The leading cells showed locomotor morphologies with fan-shaped leading lamellae while the following cells had cell contacts on all sides and appeared to lack leading lamellae. In the present ultrastructural study, the following cells frequently showed tapering cytoplasmic protrusions and leading lamella-like processes underlapping a preceding cell, indicating that the locomotion mechanism is almost the same for both the leading and following cells. For this type of locomotion as a coherent sheet we propose that localized modulation of cell-cell adhesion was induced such that wide intercellular gaps occurred at the lower portion of the cells to allow the cells to extend the tapering cytoplasmic processes and leading lamellae while close cell-cell contacts remained at the upper portion of the cells. These TPA-induced changes took place predominantly in the cells at the periphery of the cell islands, while the cells in the middle of the cell islands maintained close cell-cell contacts including complex interdigitation all around the cells, suggesting the modulation of TPA action by cell-cell interaction. Additionally, consistent with the evidence for junctional complexes between the cells moving outwards, the Lucifer-yellow dye transfer studies showed some, limited cell-cell coupling, suggesting the presence of at least some gap junctional intercellular communication in the moving cell sheets.

Smoldering adult T-cell leukemia with B-cell lymphoma and early gastric cancer.

We report a case of smoldering adult T-cell leukemia (ATL) with B-cell lymphoma and early gastric cancer. A 64-year-old man was admitted to our hospital because of proteinuria and hypergammaglobulinemia. Systemic lymphadenopathy, "flower cells" in peripheral white blood cells, and hypergammaglobulinemia with monoclonal gammopathy (IgA, lambda type) were found. As Southern blot analysis revealed monoclonal integration of human T-lymphotrophic virus type I proviral DNA in peripheral blood mononuclear cells, he was diagnosed as having smoldering ATL. The tissue specimen of an inguinal lymph node showed proliferation of abnormal lymphocytes which were stained with anti-lambda antibody, indicating B-cell lymphoma. A polypoid lesion in the stomach was histologically diagnosed as early gastric cancer.

A two-dimensional model of cell movement. Well differentiated human rectal adenocarcinoma cells move as coherent sheets upon TPA stimulation.

We previously found that 12-O-tetradecanoylphorbol-13-acetate (TPA)-enhanced invasion of Matrigel was associated with augmentation of cell motility but not with metalloproteinase activity in a highly metastatic variant (L-10) of human rectal adenocarcinoma cell line RCM-1. In the present study, with a two-dimensional cell motility assay, we investigated morphology of TPA-induced motility and biochemical pathways that may be involved in the induction of such a motile response to TPA. TPA induced active cell locomotion in L-10 cells with characteristic morphology: the cells moved outwards from the cell islands mainly as a localized coherent sheet of cells with few single moved out cells, but not cell proliferation. The front cells showed locomotor morphologies with front-tail polarity and well-spread leading lamella. Thus, this TPA-induced L-10 cell spreading and motility system seems to be a good model to investigate how well-differentiated adenocarcinoma cells move as cohesive cell nests. Agents which selectively modulate the adenylate cyclase or G protein-related pathways, e.g., 2',5'-dideoxyadenosine and pertussis toxin, had negligible effect upon motility. In contrast, the membrane-permeable synthetic diacylglycerol 1-oleoyl-2-acetyl-glycerol, which has been reported to activate protein kinase C (PKC) directly, could induce cell spreading and motility. Unexpectedly, PKC inhibitors staurosporine and H-7 enhanced TPA-induced cell spreading and motility. Staurosporine itself could induce cell spreading and motility. Taken together, these observations suggested possible involvement of PKC in TPA-induced L-10 cell spreading and motility and that staurosporine might have PKC agonist effect on induction of the spreading and motility.

Stimulation of TIMP-1 and metalloproteinase production in co-cultures of human tumor cells and human fibroblasts.

The co-cultures of five different human tumor cell lines with human normal fibroblasts significantly stimulated the production of tissue inhibitors of metalloproteinases-1 (TIMP-1) when compared to cultures of individual cells. In the co-culture of T24 human urinary bladder carcinoma cells and CCD18 human fibroblasts, production of both TIMP-1 and metalloproteinases was stimulated, and the stimulatory effects were dependent on the cellular ratio between the fibroblasts and carcinoma cells. On day 6 of culture, collagenase and stromelysin were stimulated at a ratio of CCD18 fibroblasts to T24 cells of 1:0.1, while the maximum TIMP-1 production occurred at a ratio of 1:1. Thus, the cellular ratio in the interaction of carcinoma cells with host fibroblasts affects the production of TIMP-1 and metalloproteinases and hence modulates the balance between them.

TPA-enhanced invasion of Matrigel associated with augmentation of cell motility but not metalloproteinase activity in a highly metastatic variant (L-10) of human rectal adenocarcinoma cell line RCM-1.

We previously found that the enhanced activity to invade Matrigel upon stimulation with 12-O-tetradecanoylphorbol-13-acetate (TPA) was one of the major properties of a highly metastatic variant (L-10) of a human rectal adenocarcinoma cell line RCM-1. To clarify the mechanism of this enhancement, we examined the effect of TPA on 2 major biological factors involved in tumor cell invasion: cell motility and matrix-degrading metalloproteinase activity. The enhanced invasiveness was inhibited by protein-kinase-C inhibitors. TPA markedly enhanced both haptotactic response to type-IV collagen and motility on tissue-culture glass substrate of L-10 cells in a dose-response manner quite similar to that of TPA-enhanced invasion of Matrigel. On the other hand, TPA showed little enhancement of metalloproteinase production, which was assessed by gelatin- and casein-zymography, and of type-IV collagenolytic activity. Addition of TIMP (tissue inhibitors of metalloproteinase)-I inhibited TPA-enhanced invasion of Matrigel by only up to 13%. Thus, TPA treatment of L-10 cells enhanced invasion of Matrigel in association with augmentation of cell motility but did not enhance metalloproteinase activity.

Characteristics of a metastatic variant to the liver of human rectal adenocarcinoma cell line RCM-1.

The interaction of human rectal adenocarcinoma cell line RCM-1 cells with extracellular matrix components, was studied to elucidate the key steps in the liver metastasis of colorectal carcinomas. Highly metastatic variant L-10 cells selected from the metastatic foci of the liver after intrasplenic implantation in nude mice and its parental L-0 cells were used. L-10 cells showed a greater ability to adhere to laminin, fibronectin, and type I and type IV collagens than did L-0 cells but less haptotactic activity than that of L-0 cells to type I or type IV collagen, possibly due to the formation of cellular aggregates. In vitro invasion activities of both cell lines to basement membrane components (Matrigel) or type I collagen were minimal but enhanced by the addition of 12-O-tetradecanoylphorbol-13-acetate (TPA). L-10 cells showed greater ability to invade Matrigel than did L-0 cells, while L-0 cells exhibited higher activity in the invasion of type I collagen than did L-10 cells. TPA did not increase the production of metalloproteinases by both cells when analyzed by gelatin zymography. Based on the differences between the two cell lines, we postulated the following: (1) the high metastatic potential of L-10 cells was due to a greater capacity to attach to and cross the basement membrane; (2) TPA directly enhanced tumor cell invasiveness, not via the increased secretion of metalloproteinases; and (3) haptotactic migration had no significant correlation with the increased metastatic potential of L-10 cells.

Solitary retrovesical neurofibroma extending to the perineal region.

Herein a case of solitary retrovesical neurofibroma extending to the perineal region in a 69-year-old man is reported. This tumor commonly occurs in the skin and subcutaneous tissue but has been rarely reported to originate from the retrovesical area.

Correlation between urokinase-type plasminogen activator production and the metastatic ability of human rectal cancer cells.

The correlation between the production of plasminogen activators (PA), especially urokinase-type PA (u-PA), by cancer cells and their metastatic potential was studied. For this purpose, cells from the human rectal adenocarcinoma tumor line (RCM-l/nu) originally maintained by serial passage in nude mice as the solid subcutaneous tumor, were injected into the spleen. Cancer cells from liver metastatic foci were suspended and then injected into the spleen. After 10 cycles of this selection, a highly metastatic liver tumor line termed L-10 was obtained. The amount of u-PA in the supernatant of the tumor homogenate of L-10 was larger than that of RCM-l/nu. Using an in vitro culture system, the media conditioned by L-10 cells had a higher PA activity and a higher u-PA antigen level than by RCM-l/nu cells. The apparent difference in u-PA activity and antigen levels of these two lines was not due to the difference in the production of plasminogen activator inhibitor (PAI), because PAI antigen level and PAI activity in the culture media were almost equal between them. No tissue-type PA production was detectable in these tumor lines. From these results we deduce that u-PA may play an important role in tumor metastasis.

New human colorectal carcinoma cell lines that secrete proteinase inhibitors in vitro.

Two new human cell lines, RCM-1 and CoCM-1, have been established from primary colorectal adenocarcinomas. Both cell lines were unique in that the cultures secreted trypsin inhibitors in vitro. The activities of these inhibitors were accumulated in serum-free media of both cell lines over a period of several days. Two inhibitors (PI-1 and PI-2) were isolated from serum-free conditioned medium in which RCM-1 was grown by anion-exchange and gel filtration high-performance liquid chromatography. PI-1 inhibited trypsin and chymotrypsin strongly, and pancreatic elastase weakly. Its molecular weight was about 57 kilodaltons (Kd) as determined by gel filtration chromatography. It cross-reacted with the antiserum elicited against human alpha 1-antitrypsin in double immunodiffusion. PI-1 corresponding to alpha 1-antitrypsin was also demonstrated immunohistochemically in both cell lines. PI-2 inhibited trypsin strongly, and chymotrypsin, kallikrein and plasmin weakly. It had higher molecular weight (200-300 Kd) than that of PI-1, and did not cross-react with antisera against human alpha 1-antitrypsin, alpha 2-macroglobulin, alpha 1-antichymotrypsin, alpha 2-plasmin inhibitor, inter-alpha-trypsin inhibitor and urinary trypsin inhibitor. RCM-1 and CoCM-1 are the first colorectal adenocarcinoma cell lines that secrete functionally active trypsin inhibitors, including alpha 1-antitrypsin in vitro, and are useful for the study of tumor-cell derived proteinase inhibitors.

Occurrence and formation of vertebral anomalies in hatchery reared ayu, Plecoglossu altivelis.

The influence of low food supply on the time and order of formation of vertebral column and the occurrences of vertebral anomalies in hatchery-reared ayu, Plecoglossus altivelis, were described and reported. The larvae reared with low food supply, "lack of living micro-crustaceans," grew considerably more slowly than the well fed larvae. The order of cartilaginification and ossification of the vertebral column seemed unaffected by growth rate, but time of formation varied according to the growth rate. The frequencies of abnormal cartilaginous arch and spine, abnormal arch and spine, compressed (shortened) vertebrae and fused vertebrae in fish reared with low food supply were clearly higher than those for fish with high food supply. The slow space of cartilaginification and ossification of vertebral column and the high incidence of vertebral anomalies may be associated with lack of living micro-crustaceans during the larval stage.

Electrophoretic patterns of proteins from the dentary, vertebra and scale in the ayu fish, Plecoglossus altivelis, Temminck et Schlegel.

The electrophoretic patterns of the proteins extracted from dentary, vertebra and scale with the phosphate buffer, 0.1 NHcl and SDS solution were studied to investigate their molecular aspects. The banding patterns and relative amounts of the proteins from dentary, vertebra and scale showed a basic similarity.