Familial hemophagocytic lymphohistiocytosis (FHL) is a potentially lethal genetic disorder of immune dysregulation that requires prompt and accurate diagnosis to initiate life-saving ...immunosuppressive therapy and to prepare for hematopoietic stem cell transplantation. In the present study, 85 patients with hemophagocytic lymphohistiocytosis were screened for FHL3 by Western blotting using platelets and by natural killer cell lysosomal exocytosis assay. Six of these patients were diagnosed with FHL3. In the acute disease phase requiring platelet transfusion, it was difficult to diagnose FHL3 by Western blot analysis or by lysosomal exocytosis assay. In contrast, the newly established flow cytometric analysis of intraplatelet Munc13-4 protein expression revealed bimodal populations of normal and Munc13-4–deficient platelets. These findings indicate that flow cytometric detection of intraplatelet Munc13-4 protein is a sensitive and reliable method to rapidly screen for FHL3 with a very small amount of whole blood, even in the acute phase of the disease.
Sagittal PET image of (−)-11Cgalanthamine in the ddY mouse.
Improved radiopharmaceuticals for imaging cerebral acetylcholinesterase (AChE) are needed for the diagnosis of Alzheimer’s disease (AD). ...Thus, 11C-labeled (−)-galanthamine and its enantiomers were synthesized as novel agents for imaging the localization and activity of AChE by positron emission tomography (PET). C-11 was incorporated into (−)- and (+)-11Cgalanthamine by N-methylation of norgalanthamines with 11Cmethyl triflate. Simple accumulation of 11C in the brain was measured in an in vivo biodistribution study using mice, whilst donepezil was used as a blocking agent in analogous in vivo blocking studies. In vitro autoradiography of rat brain tissue was performed to investigate the distribution of (−)-11Cgalanthamine, and confirmed the results of PET studies in mice. The radiochemical yields of N-methylation of (−)- and (+)-norgalanthamines were 13.7% and 14.4%, respectively. The highest level of accumulation of 11C in the brains of mice was observed at 10min after administration (2.1% ID/g). Intravenous pretreatment with donepezil resulted in a 30% decrease in accumulation of (−)-11Cgalanthamine in the striatum; however, levels in the cerebellum were unchanged. In contrast, use of (+)-11Cgalanthamine led to accumulation of radioactivity in the striatum equal to that in the cerebellum, and these levels were unaffected by pretreatment with donepezil. In in vitro autoradiography of regional radioactive signals of brain sections showed that pretreatment with either (−)-galanthamine or donepezil blocked the binding of (−)-11Cgalanthamine to the striatum, while sagittal PET imaging revealed accumulation of (−)-11Cgalanthamine in the brain. These results indicate that (−)-11Cgalanthamine showed specific binding to AChE, whereas (+)-11C-galanthamine accumulated in brain tissue by non-specific binding. Thus, optically pure (−)-11Cgalanthamine could be a useful PET tracer for imaging cerebral AChE.
Autoinflammatory diseases occupy one of a group of primary immunodeficiency diseases that are generally thought to be caused by mutation of genes responsible for innate immunity, rather than by ...acquired immunity. Mutations related to autoinflammatory diseases occur in 12 genes. For example, low-level somatic mosaic NLRP3 mutations underlie chronic infantile neurologic, cutaneous, articular syndrome (CINCA), also known as neonatal-onset multisystem inflammatory disease (NOMID). In current clinical practice, clinical genetic testing plays an important role in providing patients with quick, definite diagnoses. To increase the availability of such testing, low-cost high-throughput gene-analysis systems are required, ones that not only have the sensitivity to detect even low-level somatic mosaic mutations, but also can operate simply in a clinical setting. To this end, we developed a simple method that employs two-step tailed PCR and an NGS system, MiSeq platform, to detect mutations in all coding exons of the 12 genes responsible for autoinflammatory diseases. Using this amplicon sequencing system, we amplified a total of 234 amplicons derived from the 12 genes with multiplex PCR. This was done simultaneously and in one test tube. Each sample was distinguished by an index sequence of second PCR primers following PCR amplification. With our procedure and tips for reducing PCR amplification bias, we were able to analyze 12 genes from 25 clinical samples in one MiSeq run. Moreover, with the certified primers designed by our short program-which detects and avoids common SNPs in gene-specific PCR primers-we used this system for routine genetic testing. Our optimized procedure uses a simple protocol, which can easily be followed by virtually any office medical staff. Because of the small PCR amplification bias, we can analyze simultaneously several clinical DNA samples with low cost and can obtain sufficient read numbers to detect a low level of somatic mosaic mutations.
The efficacy of sorafenib against hepatocellular carcinoma (HCC) has been extensively reported. However, there is little information available about the use of sorafenib for HCC patients with ...end‐stage renal failure. We herein report the safe introduction of sorafenib therapy for a HCC patient on hemodialysis. A 63‐year‐old man had received multidisciplinary treatments, including transarterial chemoembolization (TACE) and radiofrequency ablation, for HCC since 1996, and had been undergoing hemodialysis since 2005. He also underwent TACE for multiple liver recurrence of HCC in 2011. Sorafenib therapy (200 mg/day) started 8 days after the TACE. The pharmacokinetic parameters of sorafenib and its active metabolite, M‐2, were within the reference levels observed in patients with normal renal function 8 and 9 days after the initiation of sorafenib. The dose of sorafenib was reduced to 200 mg every other day on day 154 due to hypertension and general fatigue. Because of the progression of disease after 5 months, sorafenib was withdrawn on day 180. He was admitted to the emergency department because of a high fever during hemodialysis on day 201, and died of septic shock induced by Staphylococcus lugdunensis on day 203. Sorafenib was well tolerated at an initial dose of 200 mg/day for a HCC patient undergoing hemodialysis, thus indicating that renal failure is not necessarily a contraindication for sorafenib therapy.
「ABSTRACT」 Anhidrotic ectodermal dysplasia with immunodeficiency (EDA-ID) is characterized according to its various manifestations, which include ectodermal dysplasia, vascular anomalies, ...osteopetrosis, and diverse immunological abnormalities such as susceptibility to pathogens, impaired antibody responses to polysaccharides, hypogammaglobulinemia, hyper-IgM syndrome, impaired natural killer cell cytotoxicity, and autoimmune diseases. Two genes responsible for EDA-ID have been identified: nuclear factor-κB (NF-κB) essential modulator (NEMO) for X-linked EDA-ID (XL-EDA-ID) and IκBα for autosomal-dominant EDA-ID (AD-EDA-ID). Both genes are involved in NF-κB activation, such that mutations or related defects cause impaired NF-κB signaling. In particular, NEMO mutations are scattered across the entire NEMO gene in XL-EDA-ID patients, which explains the broad spectrum of clinical manifestations and the difficulties associated with making a diagnosis. In this review, we focus on the pathophysiology of EDA-ID and different diagnostic strategies, which will be beneficial for early diagnosis and appropriate treatment.
Telaprevir (TVR) is a protease inhibitor used in combination with pegylated interferon alfa-2b and ribavirin for hepatitis C, and TVR strongly inhibits CYP3A4 and CYP3A5. We reported successful TVR ...treatment of liver transplant patients with recurrence of hepatitis C during receiving immunosuppressive therapy. Before initiation of triple therapy, all patients switched from tacrolimus to cyclosporine, which has a lower inhibitory effect on CYP3A4 and CYP3A5 than tacrolimus. To avoid graft failure, we measured the cyclosporine blood concentrations at 0, 2, and 6 h after administration to maintain the target level (150–200 ng/mL) within 1 week after initiation of TVR and adjusted the dose of cyclosporine. The dose of cyclosporine was decreased 0.24–0.40 fold in all patients after initiation of TVR treatment. In 3 patients, the dose of TVR was decreased two-thirds of starting dose because of adverse effects, including anorexia and skin rash. However, the HCV RNA level rapidly decreased to undetectable levels within 1 month. Furthermore, all patients completed the TVR therapy in 12 weeks and did not experience liver graft rejection. In addition, we found the rapid elimination of inhibitory effect of TVR on the disposition of cyclospirne in the all four cases and therefore, rapid increase in the dosage of cyclosporine would be required immediately after the end of TVR administration. These results suggest that frequent measurement of cyclosporine levels was important for successful TVR triple therapy and prevention of rejection.
•CSDS alters the gut microbiota composition in mice.•The supplementation of heat-sterilized B. breve M-16V prevents social impairment induced by CSDS.•The supplementation of heat-sterilized B. breve ...M-16V suppresses the CSDS-induced increase of IL-1β.
Major depressive disorder (MDD) is a common and serious psychiatric disease that involves brain inflammation. Bifidobacterium breve is commonly used as a probiotic and was shown to improve colitis and allergic diseases by suppressing the inflammatory response. Heat-sterilized B. breve has beneficial effects on inflammation. We hypothesize, therefore, that this probiotic might reduce depression symptoms. We tested this is a mouse model of social defeat stress. C57BL/6J mice exposed to chronic social defeat stress (CSDS) for five consecutive days developed a mild depression-like behavior characterized by a social interaction impairment. CSDS also altered the gut microbiota composition, such as increased abundance of Bacilli, Bacteroidia, Mollicutes, and Verrucomicrobiae classes and decreased Erysipelotrichi class. The prophylactic effect of heat-sterilized B. breve as a functional food ingredient was evaluated on the depression-like behavior in mice. The supplementation started two weeks before and lasted two weeks after the last exposure to CSDS. Two weeks after CSDS, the mice showed deficits in social interaction and increased levels of inflammatory cytokines, including interleukin-1β (IL-1β) in the prefrontal cortex (PFC) and hippocampus (HIP). Heat-sterilized B. breve supplementation significantly prevented social interaction impairment, suppressed IL-1β increase in the PFC and HIP, and modulated the alteration of the gut microbiota composition induced by CSDS. These findings suggest that heat-sterilized B. breve prevents depression-like behavior and IL-1β expression induced by CSDS through modulation of the gut microbiota composition in mice. Therefore, heat-sterilized B. breve used as an ingredient of functional food might prevent MDD.
Stressful life events contribute to the onset of major depressive disorder (MDD). We recently demonstrated abnormalities in ubiquitination in the pathophysiology of MDD. However, the underlying ...molecular mechanisms remain unclear. We investigated the involvement of the ubiquitination system-mediated glutamatergic dysfunction in social impairment induced by chronic social defeat stress (CSDS). Adult C57BL/6J mice were exposed to aggressor ICR male mice for 10 consecutive days. Social impairment was induced by CSDS in the social interaction test 1 days after the last stress exposure. In terms of brain microdialysis, CSDS reduced depolarization-evoked glutamate release in the prefrontal cortex (PFC), which was reversed by a glutamate transporter 1 (GLT-1) inhibitor. Interestingly, the expression of ubiquitinated, but not total GLT-1, was decreased in the PFC of mice exposed to CSDS. The expression of neural precursor cells expressing developmentally downregulated gene 4-like (Nedd4L: E3 ligase for GLT-1), and ubiquitin-conjugating enzyme E2D2 (Ube2d2: E2 ubiquitin-conjugating enzyme for Nedd4L) was also reduced in CSDS mice. Furthermore, the downregulation of the Nedd4L-GLT-1 ubiquitination pathway decreased SIT ratio, but up-regulation increased it even in non-CSDS mice. Taken together, the decrease in GLT-1 ubiquitination may reduce the release of extracellular glutamate induced by high-potassium stimulation, which may lead to social impairment, while we could not find differences in GLT-1 ubiquitination between susceptible and resistant CSDS mice. In conclusion, GLT-1 ubiquitination could play a crucial role in the pathophysiology of MDD and is an attractive target for the development of novel antidepressants.
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