Dynamics of M1 macrophages in oral mucosal lesions during the development of acute graft-versus-host disease in rats.

The role of macrophage infiltrates in oral mucosal acute graft-versus-host disease (AGVHD) remains unclear, although clinical studies suggest that macrophage infiltration correlates directly with the severity of AGVHD. In this study, we investigated the role of M1 macrophage infiltration in the oral mucosa of rats with AGVHD. Lewis rat spleen cells were injected into (Lewis × Brown Norway) F1 rats to induce systemic GVHD. Tongue samples were evaluated using histology, immunohistochemistry, dual immunofluorescence, real-time reverse transcription-polymerase chain reaction, Transwell migration assays and Stamper-Woodruff binding assays. At the onset of oral mucosal AGVHD, dual immunofluorescence and migration assays revealed that M1 macrophages had accumulated in the basement membrane (BM) region via the laminin/CD29 ß1 integrin pathway. Macrophage-secreted matrix metalloproteinase-2 was related to BM degradation. The adhesion of macrophages to the oral epithelium could be inhibited by pretreating macrophages with a CC chemokine receptor 2 (CCR2) antibody and/or pretreating lesion sections with monocyte chemoattractant protein-1 (MCP-1) antibody. Our data show that the migration and adhesion of M1 macrophages are associated with oral mucosal AGVHD, which is mediated in part by both laminin/CD29 ß 1 intern and MCP-1/CCR2 pathways. Therefore, our study provides additional support for the contribution of macrophage infiltrate to the development of oral mucosal AGVHD.

Common variants of the G protein-coupled receptor type 4 are associated with human essential hypertension and predict the blood pressure response to angiotensin receptor blockade.

Non-synonymous GRK4 variants, R65L, A142V and A486V, are associated with essential hypertension in diverse populations. This study replicated the association of GRK4 variants, including GRK4(142V), with human essential hypertension in a Japanese population (n=588; hypertensive, n=486 normotensive controls) and determined whether the presence of GRK4 variants predicted the blood pressure (BP) response to angiotensin receptor blockers (ARBs) in patients with essential hypertension. We analyzed 829 patients and compared the response to ARBs between individuals with no GRK4 variants (n=136) and those with variants at one or any of the three loci (n=693). Carriers of hGRK4(142V) had a greater decrease in systolic BP in response to ARBs than non-carrier hypertensive patients. By contrast, those with variants only at GRK4(486V) were less likely to achieve the BP goal in response to an ARB than those with no variants. These studies showed for the first time the association between GRK4(142V) and a larger decrease in BP with ARBs in hypertensive patients.

Resting salivary flow independently associated with oral malodor.

BACKGROUND: Dryness of the oral cavity is considered one cause of oral malodor. However, it is unclear which of the factors regulating the wetness of the oral cavity are involved in oral malodor development. This study investigated the effects of salivary flow and oral mucosal moisture on oral malodor. METHODS: The study population comprised 119 patients (48 men and 71 women, mean age of 50.6 ± 15.4 years) with complaint of oral malodor. After the oral malodor level had been evaluated by the organoleptic test and gas chromatography, the rates of stimulated saliva and resting saliva and the moisture levels of the tongue and buccal mucosa were measured. The plaque index, bleeding on pocket probing, probing pocket depth, and tongue coating score were also assessed. Strong oral malodor was defined as an organoleptic test score of ≥3. RESULTS: The flow rate of resting saliva in women was significantly lower than in men. The flow rate of resting saliva and the moisture levels of the tongue and buccal mucosa showed significant negative correlations with age. The flow rate of resting saliva was significantly lower in patients with strong oral malodor than in those with no or weak oral malodor. The flow rate of stimulated saliva and the moisture levels of the tongue and buccal mucosa had no relationship with strong oral malodor. Logistic regression analysis showed that a ≥5-mm probing pocket depth with bleeding on pocket probing, an increased tongue coating score, and decreased resting salivary flow were strong explanatory factors in clinical findings for oral malodor. CONCLUSION: This study suggests that the flow rate of resting saliva is a significant modulating factor for oral malodor.

Measles virus selectively blind to signaling lymphocyte activation molecule as a novel oncolytic virus for breast cancer treatment.

Oncolytic viruses hold much promise as novel therapeutic agents that can be combined with conventional therapeutic modalities. Measles virus (MV) is known to enter cells using the signaling lymphocyte activation molecule (SLAM), which is expressed on cells of the immune system. Although human breast cancer cell lines do not express SLAM, we found that a wild-type MV (HL strain) efficiently infected various breast cancer cell lines, causing cell death. Based on this finding, we used reverse genetics to generate a recombinant MV selectively unable to use SLAM (rMV-SLAMblind). The rMV-SLAMblind lacked infectivity for SLAM-positive lymphoid cells, while retaining oncolytic activity against breast cancer cells. We showed that, unlike the MV vaccine strains, rMV-SLAMblind used PVRL4 (polio virus receptor-related 4) as a receptor to infect breast cancer cells and not the ubiquitously expressed CD46. Consistent with this, rMV-SLAMblind infected CD46-positive primary normal human cells at a much-reduced level, whereas a vaccine strain of the Edmonston lineage (rMV-Edmonston) efficiently infected and killed them. The rMV-SLAMblind showed antitumor activity against human breast cancer xenografts in immunodeficient mice. The oncolytic activity of rMV-SLAMblind was significantly greater than that of rMV-Edmonston. To assess the in vivo safety, three monkeys seronegative for MV were inoculated with rMV-SLAMblind, and no clinical symptoms were documented. On the basis of these results, rMV-SLAMblind could be a promising candidate as a novel oncolytic virus for breast cancer treatment.

Protection from non-alcoholic steatohepatitis and liver tumourigenesis in high fat-fed insulin receptor substrate-1-knockout mice despite insulin resistance.

AIMS/HYPOTHESIS: Epidemiological studies have revealed that obesity and diabetes mellitus are independent risk factors for the development of non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma. However, the debate continues on whether insulin resistance as such is directly associated with NASH and liver tumourigenesis. Here, we investigated the incidence of NASH and liver tumourigenesis in Irs1 ( -/- ) mice subjected to a long-term high-fat (HF) diet. Our hypothesis was that hepatic steatosis, rather than insulin resistance may be related to the pathophysiology of these conditions. METHODS: Mice (8 weeks old, C57Bl/6J) were given free access to standard chow (SC) or an HF diet. The development of NASH and liver tumourigenesis was evaluated after mice had been on the above-mentioned diets for 60 weeks. Similarly, Irs1 ( -/- ) mice were also subjected to an HF diet for 60 weeks. RESULTS: Long-term HF diet loading, which causes obesity and insulin resistance, was sufficient to induce NASH and liver tumourigenesis in the C57Bl/6J mice. Obesity and insulin resistance were reduced by switching mice from the HF diet to SC, which also protected these mice against the development of NASH and liver tumourigenesis. However, compared with wild-type mice fed the HF diet, Irs1 ( -/- ) mice fed the HF diet were dramatically protected against NASH and liver tumourigenesis despite the presence of severe insulin resistance and marked postprandial hyperglycaemia. CONCLUSIONS/INTERPRETATION: IRS-1 inhibition might protect against HF diet-induced NASH and liver tumourigenesis, despite the presence of insulin resistance.

Performance of Acoustic Radiation Force Impulse imaging for the staging of liver fibrosis: a pooled meta-analysis.

Acoustic Radiation Force Impulse (ARFI) imaging is a novel ultrasound-based elastography method that is integrated in a conventional ultrasound machine enabling the exact localization of measurement site. It might present an alternative method to transient elastography for the noninvasive assessment of liver fibrosis. At present, studies with small patient population have shown promising results. A systematic review and meta-analysis of pooled patient data were performed to evaluate the overall performance of ARFI for the staging of liver fibrosis. Literature databases were searched up to 10/2010. The authors of the original publication were contacted, and the original patient data were requested. A meta-analysis was performed using a random effect meta-analytic method for diagnostic tests. In addition, available data comparing ARFI with FibroScan with the DeLong test were evaluated. Literature search yielded nine full-paper publications evaluating ARFI while using liver biopsy as reference method. Original patient data were available from eight studies including 518 patients. The mean diagnostic accuracy of ARFI expressed as areas under ROC curves (AUROC) was 0.87 for the diagnosis of significant fibrosis (F ≥ 2), 0.91 for the diagnosis of severe fibrosis (F ≥ 3), and 0.93 for the diagnosis of cirrhosis. ARFI can be performed with good diagnostic accuracy for the noninvasive staging of liver fibrosis.

Hashimoto's encephalopathy presenting with micrographia as a typical feature of parkinsonism.

We describe here a 63-year-old woman who presented with gait disturbance and micrographia. Laboratory tests demonstrated the presence of anti-thyroperoxidase (TPO) antibodies and vitamin B(12) deficiency accompanied by the presence of anti-parietal cell antibodies. Lymphocytosis with increased protein was detected in cerebral spinal fluid (CSF). Serum autoantibodies against the anti-NH(2) terminal of α-enolase (NAE), a specific diagnostic marker for Hashimoto's encephalopathy (HE), were also detected. Since underlying autoimmune conditions were suspected to be associated with Hashimoto's disease, steroid therapy was conducted, and the neurological symptoms improved a few days after the therapy was started. Attention should be given to the possibility that typical parkinsonism showing micrographia is caused by HE.

Association between phospholipids and free cholesterol in high-density lipoprotein and the response to hepatitis C treatment in Japanese with genotype 1b.

Pegylated interferon and ribavirin combination therapy is the standard treatment for patients with chronic hepatitis C (CHC), but treatment failure can be difficult to predict. We and others have reported a relation between lipid values and sustained viral responses in patients with CHC. However, the relationship between lipid values and treatment failure has not been previously reported. The present study investigated the association between the profiles of phospholipids and free cholesterol (FC), the main constitutive ingredients of the surface of lipoprotein, classified according to particle size and hepatitis C treatment, and determined the usefulness of these parameters for predicting the outcome of treatment. Fifty-five patients with CHC (33 men and 22 women) were included in the study. The serum total cholesterol, triglyceride, phospholipids, and FC levels in the lipoprotein subclasses were determined using high-performance liquid chromatography with gel permeation columns, enabling the lipoproteins to be classified into 13 subclasses according to particle size. According to a univariate analysis, the treatment failure group had a significantly higher serum phospholipid level overall in the high-density lipoprotein (HDL) and medium HDL fractions as well as a higher serum FC level in the HDL fraction and all HDL subclass fractions compared with the corresponding values in the non-nonvirological response group. Higher serum phospholipid and FC concentrations in the HDL subclasses were predictive of a failure to respond in patients with genotype 1b.

Association between lipoprotein subfraction profile and the response to hepatitis C treatment in Japanese patients with genotype 1b.

Pegylated interferon and ribavirin combination therapy is the standard treatment for patients with chronic hepatitis C (CHC). Some groups have reported a relation between lipid values and response while others have reported that microsomal triglyceride transfer protein, a key enzyme in the assembly and secretion of lipoproteins, was related to hepatitis C virus (HCV). The aim of this study was to investigate the association between the lipoprotein profiles, classified according to size, and hepatitis C treatment and the usefulness for predicting the outcome of treatment. Forty-four patients with CHC (27 men and 17 women) were included in the study. The serum cholesterol and triglyceride (TG) levels in the lipoprotein subclasses were determined using high-performance liquid chromatography with gel permeation columns, which classified lipoproteins into 20 subfractions based on particle size. According to a univariate analysis, those who achieved an sustained viral response (SVR) had a significantly higher serum total cholesterol level, higher cholesterol levels in the low-density lipoprotein subfraction (25.5 nm in diameter) and the very low-density lipoprotein (VLDL) subfraction (44.5 and 36.8 nm), and a higher serum TG level in the VLDL subfraction (44.5 nm), compared with the corresponding values in the non-SVR group. Higher serum cholesterol and TG concentrations in the lipoprotein subfractions were predictive of an SVR to therapy for HCV infection with genotype 1b prior to the start of interferon treatment.

Involvement of JNK pathway in the promotion of the early stage of colorectal carcinogenesis under high-fat dietary conditions.

BACKGROUND AND AIMS: The molecular mechanisms underlying the promotion of colorectal carcinogenesis by a high-fat diet (HFD) remain unclear. We investigated the role of the insulin-signal pathway and the c-Jun N-terminal kinase (JNK) pathway, which reportedly play crucial roles in insulin resistance, during colorectal carcinogenesis in the presence of hyperinsulinaemia induced by a HFD. METHODS: Azoxymethane-induced aberrant crypt foci formation and cell proliferation in the colonic epithelium were compared between mice fed a normal diet (ND) and mice fed a HFD. A western blot analysis was performed to elucidate the mechanism affecting colorectal carcinogenesis by a HFD. RESULTS: The number of aberrant crypt foci and the colonic epithelial cell proliferative activity were significantly higher in the HFD group than in the ND group. While the plasma insulin level was significantly higher in the HFD group than in the ND group, a western blot analysis revealed the inactivation of Akt, which is located downstream of the insulin receptor, in the colonic epithelia of the HFD group. On the other hand, JNK activity was significantly higher in the HFD group than in the ND group. A JNK specific inhibitor significantly suppressed the increase in epithelial cell proliferation only under a HFD, but not under a ND. CONCLUSIONS: Colonic cell proliferation was promoted via the JNK pathway in the presence of a HFD but not in the presence of a ND. This novel mechanism may explain the involvement of the JNK pathway in the effect of dietary fat intake on colon carcinogenesis.

Wave of stiffness propagating along the surface of the newt egg during cleavage.

In the eggs of the newt, Cynops (Triturus) pyrrhogaster, change in stiffness of the cortex was measured in various regions at the time of the cleavage. Measurements were performed by Mitchison and Swann's cell elastimeter method with a modification, in which two fine pipettes were attached to the surface of one egg at the same time, in order to compare the rigidity of two regions. The stiffness of the cortex changed very little before the start of the first cleavage. However, just before the appearance of the first cleavage furrow, the stiffness increased rapidly at the animal pole region, which later returned to the former level. As the cleavage furrow progressed, a wave of high stiffness travelled meridionally as a belt along the surface from the animal pole region toward the vegetal region. At second cleavage, the cycle of change in stiffness was repeated.

Aperitif effects on gastric emptying: a crossover study using continuous real-time 13C breath test (BreathID System).

The aim of this study was to determine whether there is a correlation between aperitif and gastric emptying. Ten healthy male volunteers participated in this randomized, two-way crossover study. Under two conditions (after drinking an aperitif versus not), the (13)C breath test was performed for 4 h with a liquid meal (200 kcal/200 ml) containing 100 mg (13)C acetate. We used 50 ml of umeshu as the aperitif. This is a traditional Japanese plum liqueur, and contains 7 ml alcohol (14%). In the aperitif group, T(1/2), T(lag), and T(peak) were significantly delayed [T(1/2) (132: 113-174) versus (112: 92-134) (P = 0.0069); T(lag) (80: 63-94) versus (55: 47-85) (P = 0.0069); and T(peak) (81: 62-96) versus (54: 34-84) (P = 0.0069), (median: range, aperitif versus control, min)]. Gastric emptying was significantly delayed in the aperitif group as compared with the control group. This study revealed that even a small amount of alcohol such as an aperitif may contribute to delayed gastric emptying.

Risk factors for the progression of endoscopic Barrett's epithelium in Japan: a multivariate analysis based on the Prague C & M Criteria.

PURPOSE: To determine the prevalence and progression of Barrett's epithelium and associated risk factors in Japan. METHODS: The study population comprised 869 cases. Endoscopic Barrett's epithelium was diagnosed based on the Prague C & M Criteria. The correlations of clinical factors with the prevalence and progression of endoscopic Barrett's epithelium were examined. RESULTS: Endoscopic Barrett's epithelium was diagnosed in 374 cases (43%), in the majority of which the diagnosis was short-segment Barrett's esophagus. The progression of Barrett's epithelium was identified in 47 cases. In univariate and multiple logistic regression analyses, aging, smoking habit, and erosive esophagitis were significantly associated with the prevalence of Barrett's epithelium, whereas aging and erosive esophagitis, especially severe erosive esophagitis, were significant contributing factors to the progression of Barrett's epithelium. CONCLUSIONS: Forty-three percent of the total study population was diagnosed as having endoscopic Barrett's epithelium. During the follow-up period, 12.6% of the cases with Barrett's epithelium exhibited progression which was associated with aging and severe erosive esophagitis.

Acute on chronic subdural hematoma as a rare complication in a microscopic polyangiitis patient receiving antithrombotic treatment.

We report a 56-year-old man with microscopic polyangiitis (MPA) who developed acute exacerbation of a chronic subdural hematoma (SDH). Laboratory data demonstrated elevation of myeloperoxidase antineutrophil cytoplasmic antibody (MPOANCA) and rapidly progressing renal dysfunction. Renal biopsy showed crescentic glomerulonephritis (GN) with membranous nephropathy (MN). He was treated with corticosteroids, antithrombotic agents, and an immunosuppressant. One month after initiation of treatment, he had a mild headache. One month later, he developed acute SDH. Although he recovered completely after the operation, he finally died of bacterial infection. On autopsy, a scar of vasculitis was confirmed in the leptomeninges as well as in the kidney and lung. Although SDH is a rare complication in MPA, nephrologists must pay more attention to the initial symptoms before a hematoma attack such as headache, especially in patients using antithrombotic agents.

Assessment of reproductive potential in multiple-spawning fish with indeterminate fecundity: a case study of yellow sea bream Dentex hypselosomus in the East China Sea.

This study examined the spawning season, spawning frequency and batch fecundity of yellow sea bream Dentex hypselosomus in the East China Sea to reassess the previously reported reproductive characteristics of the species. Time-course sampling showed that this species had a diurnal ovarian maturation rhythm. Late tertiary yolk-stage oocytes appeared 2 days before spawning, starting the process of germinal vesicle movement and breakdown. On the day of spawning, ovulation and subsequent spawning occurred in the early morning (0400-0800 hours). Postovulatory follicles disappeared from the ovaries within c. 24 h of ovulation. Seasonal changes in the ovarian conditions indicated that this species spawned more or less throughout the year, with the peak ranging from spring to autumn. The compositions of the developing oocytes and degenerating postovulatory follicles in the ovaries suggested that most females spawned repeatedly over 2 to 3 consecutive days during the peak of the spawning season. Somatic body condition did not have a significant effect on batch fecundity, but there was a significant relationship between batch fecundity and fork length according to spawning status. Females spawning on consecutive days were more fecund than those spawning every other day. The findings show that this species has much greater reproductive potential than previously estimated.

Effectiveness of antiplatelet drugs against experimental non-alcoholic fatty liver disease.

OBJECTIVE: No effective drugs have been developed to date to prevent or treat non-alcoholic fatty liver disease (NAFLD), although diet modification and exercise to improve obesity have been attempted. Therefore, development of a novel drug/strategy to treat NAFLD is urgently needed. In the present study, a novel concept is proposed for the treatment of NAFLD. METHODS: Fisher 344 male rats were given a choline-deficient, l-amino acid-defined (CDAA) diet or a high-fat high-calorie (HF/HC) diet with or without the antiplatelet agents, aspirin, ticlopidine or cilostazol for 16 weeks. Liver steatosis, inflammation and fibrosis, and the possible mechanisms involved were investigated. RESULTS: All three antiplatelet drugs, namely aspirin, ticlopidine and cilostazol, significantly attenuated liver steatosis, inflammation and fibrosis in the CDAA diet group. Of the three agents, cilostazol was the most effective, and the drug also suppressed HF/HC diet-induced liver steatosis. Cilostazol appeared to exert its beneficial effect against NAFLD by suppressing mitogen-activated protein kinase activation induced by oxidative stress and platelet-derived growth factor via intercepting signal transduction from Akt to c-Raf. CONCLUSION: Antiplatelet agents, especially cilostazol, offer the promise of becoming key agents for the treatment of NAFLD.

EFFECT ON MICROBIAL PRODUCTS ON CAESIUM ELUTION BEHAVIOUR FROM CLAY MINERALS.

Some microorganisms in the environment make siderophores, which are low molecular chelators, to take up minerals from soil. Eleven bacteria were separated from the root of white clover by chlome azrol S (CAS) assay. Each bacterium was incubated in casamino acid (CAA) culture, and siderophores in CAA culture were purified. These extractions were applied to biotite or vermiculite spiked with Cs. From each clay mineral, 57.1-72.8% (5100 ppm), 55.6-63.8% (920 ppm) and 48.6-54.3% (2300 ppm), 31.6-34.4% (520 ppm) was eluted, respectively. To understand elution behaviour, Cs desorption ratio of each clay was measured every 30 min. The results indicate Cs elution was occurred quickly.

Long-term forskolin stimulation induces AMPK activation and thereby enhances tight junction formation in human placental trophoblast BeWo cells.

BeWo cells, derived from human choriocarcinoma, have been known to respond to forskolin or cAMP analogues by differentiating into multinucleated cells- like syncytiotrophoblasts on the surfaces of chorionic villi of the human placenta. In this study, we demonstrated that long-term treatment with forskolin enhances the tight junction (TJ) formation in human placental BeWo cells. Interestingly, AMPK activation and phosphorylation of acetyl-CoA carboxylase (ACC), a molecule downstream from AMPK, were induced by long-term incubation (>12h) with forskolin, despite not being induced by acute stimulation with forskolin. In addition, co-incubation with an AMPK inhibitor, compound C, as well as overexpression of an AMPK dominant negative mutant inhibited forskolin-induced TJ formation. Thus, although the molecular mechanism underlying AMPK activation via the forskolin stimulation is unclear, the TJ formation induced by forskolin is likely to be mediated by the AMPK pathway. Taking into consideration that TJs are present in the normal human placenta, this mechanism may be important for forming the placental barrier system between the fetal and maternal circulations.