Attitudes toward overtime work and self-training: A survey on obstetricians and gynecologists in Japan.

AIM: The Ministry of Health, Labour, and Welfare of Japan proposed a regulation of overtime work as a reform in work style. However, the regulation may deteriorate the quality of medical services due to the reduction in training time. Thus, the study aimed to reveal perceptions in terms of generation gaps in views on self-training and overtime work, among members of the Japan Society of Obstetrics and Gynecology (JSOG). METHODS: A web-based, self-administered questionnaire survey was conducted among members of the JSOG. In total, 1256 respondents were included in the analysis. Data were collected on age, sex, experience as a medical doctor, location of workplace, work style, the type of main workplace, and number of full-time doctors in the main workplace. The study examined the attitudes of the respondents toward overtime work and self-training. The respondents were categorized based on experience as a medical doctor. RESULTS: According to years of experience, 112 (8.9%), 226 (18.0%), 383 (30.5%), 535 (42.6%) doctors have been working for ≤5, 6-10, 11-19, and ≥ 20 years, respectively. Although 54.5% of doctors with ≤5 years of experience expected the regulation on working hours to improve the quality of medical services, those with ≥20 years of experience expressed potential deterioration. After adjusting for covariates, more years of experience were significantly related with the expectation of deterioration in the quality of medical services. CONCLUSIONS: The study revealed a generation gap in the views about self-training and overtime work among obstetricians and gynecologists in Japan.

Novel biomarker candidates for the diagnosis of ovarian clear cell carcinoma.

Ovarian clear cell carcinoma can arise from endometriosis; however, it is distinct from other types of epithelial ovarian carcinoma in terms of its clinicopathological and molecular features. Cancer antigen 125 lacks the sensitivity and specificity required for accurate clinical diagnosis of clear cell carcinoma. Therefore, the aim of the current review was to identify novel biomarker candidates for the immunohistochemical and serological diagnosis of clear cell carcinoma. A search of the relevant English language literature published between 1966 and 2014 was conducted using the PubMed MEDLINE online database. High-throughput tissue microarray technology and proteomic screening combined with mass spectrometry may provide additional information regarding diagnostic biomarker candidates for ovarian clear cell carcinoma. The present review summarizes the characteristics of potential genomic alterations that activate cancer signaling pathways and, thus, contribute to carcinogenesis. The major signaling pathways activated in clear cell carcinoma are associated with cell cycle regulation (hepatitis A virus cellular receptor 1 and tumor protein D52), growth factor signaling (insulin-like growth factor binding protein 1; KiSS-1 metastasis-suppressor; erb-b2 receptor tyrosine kinase 2; and fibroblast growth factor receptor 2), anti-apoptosis and survival pathways [sialidase 3 (membrane sialidase)], metabolism (γ-glutamyltransferase 1), chemoresistance (napsin A aspartic peptidase, glutathione peroxidase 3; and aldehyde dehydrogenase 1 family, member A1), coagulation [coagulation factor III (thromboplastin, tissue factor); and tissue factor pathway inhibitor 2], signaling (lectin, galactoside-binding and soluble, 3), and adhesion and the extracellular matrix [cadherin 1, type 1, E-cadherin (epithelial); versican; and laminin, α 5]. The present review of the relevant literature may provide a basis for additional clinical investigation of the ovarian clear cell carcinoma serum biomarker candidate proteins identified herein.

Extensive screening system using suspension array technology to detect mitochondrial DNA point mutations.

We established an extensive and rapid system using suspension array to detect 61 representative mitochondrial DNA (mtDNA) heteroplasmic or homoplasmic point mutations (29 for Series A and 32 for Series B) in 22 genes: 1 each in MT-RNR1, -TV, -ND1, -TQ, -TW, -TC, and -TH genes; 2 each in MT-TN, -TG, -ND4, -TL2, -TE, and -CYB genes; 3 each in MT-ATP6, -ND3, and -ND5 genes; 4 each in MT-CO1 and -TK genes; 5 each in MT-TI, -TS1, and -ND6 genes; and 10 in the MT-TL1 gene. We carefully selected 5'-biotinylated primers and pooled primers for use in two sets of multiplex-PCR amplifications. To detect both mutant and wild-type mtDNA, even when polymorphisms were present near the target mutation sites, we designed specific oligonucleotide probes. By using the mtDNA point mutation detection system of Series A (29 mutations) and Series B (32 mutations), we screened a total of 3103 mutant sites in 107 DNA samples for Series A and 13,101 mutant sites in 397 DNA samples for Series B. We succeeded in determining 99.4% (Series A) and 99.6% (Series B) of the targeted mutant sites by use of the system. The 22 samples with the m.3243A>G heteroplasmic mutation revealed positive signals with both mutant- and wild-type-specific probes in this detection system with a detection limit of approximately 2%. This genetic screening platform is useful to reach a definitive diagnosis for mitochondrial diseases.

TRH therapy in a patient with juvenile Alexander disease.

Alexander disease is a rare disorder of the central nervous system caused by a de novo mutation in the glial fibrillary acidic protein (GFAP) gene. Unlike the much more common infantile form, the juvenile form is slowly progressive with bulbar, pyramidal and cerebellar signs. Herein, we report a 9-year old Japanese girl suffering from frequent vomiting, slurred speech and truncal ataxia. Juvenile Alexander disease was diagnosed by genetic analysis, which detected a novel GFAP mutation, D360V. We also describe our clinical success in treating this patient with thyrotropin releasing hormone (TRH).

High bone turnover of type I collagen depends on fetal growth.

The bone metabolic processes of proliferation and differentiation in preterm and term newborns have yet to be fully elucidated. Seventy-four umbilical cord blood samples were collected from preterm and term newborns delivered at 27 to 42 gestational weeks (GWs). Carboxy-terminal propeptide of type I procollagen (PICP), pyridinoline cross-linked telopeptide domain of type I collagen (ICTP), alkaline phosphatase (ALP), and bone-specific alkaline phosphatase (BAP) were measured. Calcitonin (CT), estrogen (E2), intact parathyroid hormone, and insulin-like growth factor-I (IGF-I) were also examined in 20 or 23 randomly selected samples. We conducted cross-sectional regression analyses for bone metabolic markers, fetal growth markers including GWs, birth weight (BW), height (BH) and head circumference (HC), and bone related hormones. PICP and ICTP activities were very high, but decreased significantly with fetal growth based on GWs, BW, BH, and HC changes (GWs, BW, and BH to both PICP and ICTP, P < 0.0001; HC to ICTP, P < 0.0001; HC to PICP, P < 0.05), while BAP and ALP did not change significantly. E2 and CT both showed a significant positive correlation with Ca (P < 0.05), but neither hormone had any apparent correlation with PICP, ALP, BAP, or ICTP. These results suggest very active bone formation and resorption of type I collagen to be dependent on fetal growth and that fetal osteoblasts dominate the proliferation phase of development rather than the maturation phase. However, factors contributing to high bone turnover in the fetus remain to be elucidated.

Platelet mitochondrial evaluation during cytochrome c and dichloroacetate treatments of MELAS.

We hypothesized that serial changes in platelet (PLT) mitochondrial enzyme (ME) activities might correspond to the effects of medications for mitochondrial encephalomyopathy and stroke-like episodes (MELAS). Cytochrome c and sodium dichloroacetate (DCA) were given to a 7-year-old girl with MELAS who had an A3243G mitochondrial DNA mutation. The effects were evaluated with whole PLT-ME assays, developed by our group, using a microplate-reader. During cytochrome c treatment, complex II+III (II+III), complex IV (IV) and citrate synthase (CS) activities showed gradual but statistically significant decrease. II+III activity dropped below normal. II+III/CS activity was initially below normal, followed by a transient improvement, then decreased again before the appearance of central nervous system symptoms. II+III, IV, II+III/CS and IV/CS activities reached their lowest levels in association with a stroke-like episode, then increased with DCA treatment. Our results suggest that progressive mitochondrial dysfunction may occur before the stroke-like episodes in MELAS and that DCA treatment may increase mitochondrial activities. Our whole PLT-ME assay system may be useful for serially evaluating mitochondrial functions in relation to clinical symptoms.

Positive contribution of pathogenic mutations in the mitochondrial genome to the promotion of cancer by prevention from apoptosis.

The role of mitochondrial dysfunction in cancer has been a subject of great interest and much ongoing investigation. Although most cancer cells harbor somatic mutations in mitochondrial DNA (mtDNA), the question of whether such mutations contribute to the promotion of carcinomas remains unsolved. Here we used trans-mitochondrial hybrids (cybrids) containing a common HeLa nucleus and mtDNA of interest to compare the role of mtDNA against the common nuclear background. We constructed cybrids with or without a homoplasmic pathogenic point mutation at nucleotide position 8,993 or 9,176 in the mtDNA ATP synthase subunit 6 gene (MTATP6) derived from patients with mitochondrial encephalomyopathy. When the cybrids were transplanted into nude mice, the MTATP6 mutations conferred an advantage in the early stage of tumor growth. The mutant cybrids also increased faster than wild type in culture. To complement the mtDNA mutations, we transfected a wild-type nuclear version of MTATP, whose codons were converted to the universal genetic codes containing a mitochondrial target sequence, into the nucleus of cybrids carrying mutant MTATP6. The restoration of MTATP slowed down the growth of tumor in transplantation. Conversely, expression of a mutant nuclear version of MTATP6 in the wild-type cybrids declined respiration and accelerated the tumor growth. These findings showed that the advantage in tumor growth depended upon the MTATP6 function but was not due to secondary nuclear mutations caused by the mutant mitochondria. Because apoptosis occurred less frequently in the mutant versus wild-type cybrids in cultures and tumors, the pathogenic mtDNA mutations seem to promote tumors by preventing apoptosis.

[The problems with pediatric home care].

It is obvious that we are facing serious problems with development of home medical care for children who have severe physical difficulties. One of the problems is a shortage of pediatricians who have a sufficient and specialized knowledge to take care those patients. In this paper, we report some actual problems with the care of the child suffering from mitochondrial encephalomyopathy at home TPN.

[A two-year-old infant with a myopathic form of very-long-chain Acyl-CoA dehydrogenase deficiency].

A two-year-three-month old girl was hospitalized for detailed examination following repeated hyper-creatine kinasemia and cervical muscle cramps induced by pyrexia and persistent hypertonicity of the cervical muscles. Physical examination showed mild hypotonia but no muscle weakness. Induction of symptoms by continuous cervical muscular exercise and the appearance of dicarboxylic aciduria during the fasting test indicated a disorder of fatty acid oxidation. Free fatty acid and acyl carnitine analyses using dried blood spots, and acyl-CoA dehydrogenase activity assays using cultured skin fibroblasts established a diagnosis of very-long-chain acyl-CoA dehydrogenase (VLCAD) deficiency. Currently VLCAD deficiency has been divided into three phenotypes; a severe childhood form, a milder childhood form, and an adult form. However, we suggest that the severe and milder childhood forms would be better described as a systemic form, and the adult form and our infant case as a myopathic form. An early onset of the myopathic form within the first year of life, as well as its diagnosis in early infancy, has never been described in the literature.

Continuous culture of novel mitochondrial cells lacking nuclei.

We isolated stable cell lines, designated as mitochondrial cells, from cybrids obtained by fusing mitochondria-less HeLa cells with platelets from patients with Leigh syndrome, a subtype of mitochondrial encephalomyopathy. The cells contain a pathogenic point mutation, T9176C, in the mitochondrial DNA. Hematoxylin-eosin staining, confocal fluorescent microscopy and flow cytometry in fixed or living cells showed that the majority of these mitochondrial cells lack nuclear DNA and nuclei, but contain active mitochondria. Despite the absence of nuclear DNA, these cells can be continuously generated in culture. Therefore, it is likely that they arise from the minority of cells which possess a nucleus.