Chronic Pulmonary Disease Caused by Tsukamurella toyonakaense

Unidentified Mycobacterium species are sometimes detected in respiratory specimens. We identified a novel Tsukamurella species (Tsukamurella sp. TY48, RIMD 2001001, CIP 111916^T), Tsukamurella toyonakaense, from a patient given a misdiagnosis of nontuberculous mycobacterial pulmonary disease caused by unidentified mycobacteria. Genomic identification of this Tsukamurella species helped clarify its clinical characteristics and epidemiology.     

Trehalose Suppresses Lysosomal Anomalies in Supporting Cells of Oocytes and Maintains Female Fertility

Supporting cells of oocytes, i.e., cumulus cells, control oocyte quality, which determines fertilization success. Therefore, the transformation of mature and immature cumulus cells (MCCs and ICCs, respectively) into dysmature cumulus cells (DCCs) with dead characteristics deteriorates oocyte quality. However, the molecular basis for this transformation remains unclear. Here, we explored the link between autophagic decline and cumulus transformation using cumulus cells from patients with infertility, female mice, and human granulosa cell-derived KGN cell lines. When human cumulus cells were labeled with LysoTracker probes, fluorescence corresponding to lysosomes was enhanced in DCCs compared to that in MCCs and ICCs. Similarly, treatment with the autophagy inhibitor chloroquine elevated LysoTracker fluorescence in both mouse cumulus cells and KGN cells, subsequently suppressing ovulation in female mice. Electron microscopy analysis revealed the proliferation of abnormal lysosomes in chloroquine-treated KGN cells. Conversely, the addition of an autophagy inducer, trehalose, suppressed chloroquine-driven problematic lysosomal anomalies and ameliorated ovulation problems. Our results suggest that autophagy maintains the healthy state of the supporting cells of human oocytes by suppressing the formation of lysosomes. Thus, our results provide insights into the therapeutic effects of trehalose on female fertility.     

Prognostic determinants of long-term survival in Japanese patients with cardiac sarcoidosis treated with prednisone.

Cardiac involvement is an important prognostic factor in sarcoidosis, but reliable indicators of mortality risk in cardiac sarcoidosis are unstudied in a large number of patients. To determine the significant predictors of mortality and to assess the efficacy of corticosteroids, we analyzed clinical findings, treatment, and prognosis in 95 Japanese patients with cardiac sarcoidosis. Twenty of these 95 patients had cardiac sarcoidosis proven by autopsy; none of these patients had received corticosteroids. We assessed 12 clinical variables as possible predictors of mortality by Cox proportional hazards model in 75 steroid-treated patients. During the mean follow-up of 68 months, 29 patients (73%) died of congestive heart failure and 11 (27%) experienced sudden death. Kaplan-Meier survival curves showed 5-year survival rates of 75% in the steroid-treated patients and of 89% in patients with a left ventricular ejection fraction > or = 50%, whereas there was only 10% 5-year survival rate in autopsy subjects. There was no significant difference in survival curves of patients treated with a high initial dose (> 30 mg) and a low initial dose (> or = 30 mg) of prednisone. Multivariate analysis identified New York Heart Association functional class (hazard ratio 7.72 per class I increase, p = 0.0008), left ventricular end-diastolic diameter (hazard ratio 2.60/10 mm increase, p = 0.02), and sustained ventricular tachycardia (hazard ratio 7.20, p = 0.03) as independent predictors of mortality. In conclusion, the severity of heart failure was one of the most significant independent predictors of mortality for cardiac sarcoidosis. Starting corticosteroids before the occurrence of systolic dysfunction resulted in an excellent clinical outcome. A high initial dose of prednisone may not be essential for treatment of cardiac sarcoidosis.     

Alteration of Annexin IV Expression in Alcoholics

Western blot analysis was performed by using a specific antibody to measure annexin IV in human postmortem brain samples from alcoholic subjects. The analysis showed a significantly augmented expression in the hippocampus compared with controls, whereas the expression in the frontal cortex was equivalent in both groups. Annexin IV expression in the occipital cortex tended to increase in alcoholics. It was shown further that autoantibodies to annexin IV were increased significantly in alcoholic patients compared with controls. Thus, annexin IV may become a novel biological marker for alcoholics.     

Clinicopathological study of diffuse neurofibrillary tangles with calcification. With special reference to TDP-43 proteinopathy and alpha-synucleinopathy

Diffuse neurofibrillary tangles with calcification (DNTC) is a relatively rare presenile dementia that clinically shows overlapping symptoms of Alzheimer's disease and frontotemporal lobar degeneration (FTLD). DNTC is pathologically characterized by localized temporal or frontotemporal atrophy with massive neurofibrillary tangles, neuropil threads and Fahr's-type calcification without senile plaques. We tried to clarify the molecular basis of DNTC by immunohistochemically examining the appearance and distribution of accumulated alpha-synuclein (aSyn) and TAR DNA-binding protein of 43kDa (TDP-43) in the brains of 10 Japanese autopsy cases. We also investigated the clinically characteristic symptoms from the clinical charts and previous reports, and the correlations with neuropathological findings. The characteristic symptoms were evaluated using the Neuropsychiatric Inventory Questionnaire (NPI-Q). As a result, we confirmed the high frequency of neuronal cytoplasmic accumulation of aSyn (80%) and phosphorylated TDP-43 (90%) in DNTC cases. There was a significant correlation between some selected items of NPI-Q scores and the severity of the limbic TDP-43 pathology. The pathology of DNTC included TDP-43 and aSyn pathology with high frequency. These abnormal accumulations of TDP-43 might be involved in the pathological process of DNTC, having a close relationship to the FTLD-like psychiatric symptoms during the clinical course.     

Heterogeneity and potentiation of AMPA type of glutamate receptors in rat cultured microglia

alpha-amino-hydroxy-5-methyl-isoxazole-4-propionate (AMPA) receptor in rat cultured microglia were analyzed precisely using flop- and flip-preferring allosteric modulators of AMPA receptors, 4-[2-(phenylsulfonylamino)ethylthio]-2,6-difluoro-phenoxyacetamide (PEPA) and cyclothiazide (CTZ), respectively. Glutamate (Glu)- or kainite (KA)-induced currents were completely inhibited by a specific blocker of AMPA receptor, LY300164, indicating that functional Glu-receptors in cultured microglia are mostly AMPA receptor but not KA receptor in many cells. Glu- and KA-induced currents were potentiated by PEPA and CTZ in a concentration-dependent manner. The ratio of the potentiation by PEPA to the potentiation by cyclothiazide varied with cells between 0.1 and 0.9, suggesting cell-to-cell heterogeneity of AMPA receptor subunits expressed in microglia. Quantitative RT-PCR revealed that GluR1-3 mainly occurred in the flip forms, which agreed with the stronger potentiation of receptor currents by CTZ vs. PEPA. Finally, the potentiation of microglial AMPA receptors by PEPA and CTZ inhibited the Glu-induced release of tumor necrosis factor-alpha (TNF-alpha) unpredictably. The increase in TNF-alpha release by Glu or KA required extracellular Na+ and Ca2+ ions but not mitogen-activated protein kinase (MAPK), suggesting the effects of PEPA and CTZ were not due to the inhibition of MAPK. These results suggest that potentiation of microglial AMPA receptors serves as a negative feedback mechanism for the regulation of TNF-alpha release and may contribute to the ameliorating effects of allosteric modulators of AMPA receptors.     

Inactivation of the cystatin E/M tumor suppressor gene in cervical cancer

We have previously localized a cervical cancer tumor suppressor gene to a 300 kb interval of 11q13. Analysis of candidate genes revealed loss of expression of cystatin E/M, a lysosomal cysteine protease inhibitor, in 6 cervical cancer cell lines and 9 of 11 primary cervical tumors. Examination of the three exons in four cervical cancer cell lines, 19 primary tumors, and 21 normal controls revealed homozygous deletion of exon 1 sequences in one tumor. Point mutations were observed in six other tumors. Two tumors contained mutations at the consensus binding sites for cathepsin L, a lysosomal protease overexpressed in cervical cancer. Introduction of these two point mutations using site directed mutagenesis resulted in reduced binding of mutated cystatin E/M to cathepsin L. Although mutations were not observed in any cell lines, four cell lines and 12 of 18 tumors contained promoter hypermethylation. Reexpression of cystatin E/M was observed after 5'aza 2-deoxycytidiene and/or Trichostatin A treatment of cervical cancer cell lines, HeLa and SiHa, confirming promoter hypermethylation. Ectopic expression of cystatin E/M in these two cell lines resulted in growth suppression. There was also suppression of soft agar colony formation by HeLa cells expressing the cystatin E/M gene. Reexpression of cystatin E/M resulted in decreased intracellular and extracellular expression of cathepsin L. Overexpression of cathepsin L resulted in increased cell growth which was inhibited by the reintroduction of cystatin E/M. We conclude, therefore, that cystatin E/M is a cervical cancer suppressor gene and that the gene is inactivated by somatic mutations and promoter hypermethylation.