Bar-code (tag) microarrays of yeast gene-deletion collections facilitate the systematic identification of genes required for growth in any condition of interest. Anti-sense strands of amplified bar-codes hybridize with ~10,000 (5,000 each for up- and down-tags) different kinds of sense-strand probes on an array. In this study, we optimized the hybridization processes of an array for fission yeast. Compared to the first version of the array (11 µm, 100K) consisting of three sectors with probe pairs (perfect match and mismatch), the second version (11 µm, 48K) could represent ~10,000 up-/down-tags in quadruplicate along with 1,508 negative controls in quadruplicate and a single set of 1,000 unique negative controls at random dispersed positions without mismatch pairs. For PCR, the optimal annealing temperature (maximizing yield and minimizing extra bands) was 58℃ for both tags. Intriguingly, up-tags required 3× higher amounts of blocking oligonucleotides than down-tags. A 1:1 mix ratio between up- and down-tags was satisfactory. A lower temperature (25℃) was optimal for cultivation instead of a normal temperature (30℃) because of extra temperature-sensitive mutants in a subset of the deletion library. Activation of frozen pooled cells for >1 day showed better resolution of intensity than no activation. A tag intensity analysis showed that tag(s) of 4,316 of the 4,526 strains tested were represented at least once; 3,706 strains were represented by both tags, 4,072 strains by up-tags only, and 3,950 strains by down-tags only. The results indicate that this microarray will be a powerful analytical platform for elucidating currently unknown gene functions.
Oligonucleotides ; Polymerase Chain Reaction ; Schizosaccharomyces ; Yeasts

No abstract available.
Gastrointestinal Microbiome ; Stroke

Incorporation of unique barcodes into fission yeast gene deletion collections has enabled the identification of gene functions by growth fitness analysis. For fine tuning, it is important to examine barcode sequences, because mutations arise during strain construction. Out of 8,708 barcodes (4,354 strains) covering 88.5% of all 4,919 open reading frames, 7,734 barcodes (88.8%) were validated as high-fidelity to be inserted at the correct positions by Sanger sequencing. Sequence examination of the 7,734 high-fidelity barcodes revealed that 1,039 barcodes (13.4%) deviated from the original design. In total, 1,284 mutations (mutation rate of 16.6%) exist within the 1,039 mutated barcodes, which is comparable to budding yeast (18%). When the type of mutation was considered, substitutions accounted for 845 mutations (10.9%), deletions accounted for 319 mutations (4.1%), and insertions accounted for 121 mutations (1.6%). Peculiarly, the frequency of substitutions (67.6%) was unexpectedly higher than in budding yeast (~28%) and well above the predicted error of Sanger sequencing (~2%), which might have arisen during the solid-phase oligonucleotide synthesis and PCR amplification of the barcodes during strain construction. When the mutation rate was analyzed by position within 20-mer barcodes using the 1,284 mutations from the 7,734 sequenced barcodes, there was no significant difference between up-tags and down-tags at a given position. The mutation frequency at a given position was similar at most positions, ranging from 0.4% (32/7,734) to 1.1% (82/7,734), except at position 1, which was highest (3.1%), as in budding yeast. Together, well-defined barcode sequences, combined with the next-generation sequencing platform, promise to make the fission yeast gene deletion library a powerful tool for understanding gene function.
DNA* ; Gene Deletion* ; Mutation Rate ; Open Reading Frames ; Polymerase Chain Reaction ; Saccharomycetales ; Schizosaccharomyces*

BACKGROUND AND OBJECTIVES: Pneumatization of air cells in the mastoid bone is decreased in chronic otitis media (COM). A decrease in the size of the external auditory canal (EAC) is also found frequently in patients with COM, but this has been little studied. We compared the size of affected bony EACs and the contralateral side in patients with single-side COM using high-resolution computed tomography. SUBJECTS AND METHODS: In total, 99 patients with single-side COM were included. Four indicators related to the size of the bony EAC and IAC were measured using high-resolution computed tomography: the axial and coronal lengths of the tympanic membrane, the length of the isthmus, and the area of the bony ear canal. We also compared both internal auditory canals as negative controls. These assessments were made by radiologists who were blinded to the objective of this study. RESULTS: In patients with single-side COM, the axial length of the tympanic membrane was significantly shorter than normal, and the volume of the EAC was also significantly smaller. The length of the isthmus of the EAC was shorter on the affected side, but the difference was not significant. The IAC volume showed no difference between the two sides. CONCLUSIONS: COM affects general temporal bony development, including the bony EAC and mastoid bone. Therefore, whether to correct this should be considered when preparing for COM surgery.
Ear Canal* ; Humans ; Mastoid ; Otitis Media* ; Otitis* ; Tympanic Membrane

BACKGROUND: The present study evaluated the efficacy of a combination of ibandronate and cholecalciferol on the restoration of the levels of 25-hydroxyvitamin D (25[OH]D) and various bone markers in postmenopausal women with osteoporosis. METHODS: This was a randomized, double-blind, active-controlled, prospective 16-week clinical trial conducted in 20 different hospitals. A total of 201 postmenopausal women with osteoporosis were assigned randomly to one of two groups: the IBN group, which received a once-monthly pill containing 150 mg ibandronate (n=99), or the IBN+ group, which received a once-monthly pill containing 150 mg ibandronate and 24,000 IU cholecalciferol (n=102). Serum levels of 25(OH)D, parathyroid hormone (PTH), and various bone markers were assessed at baseline and at the end of a 16-week treatment period. RESULTS: After 16 weeks of treatment, the mean serum levels of 25(OH)D significantly increased from 21.0 to 25.3 ng/mL in the IBN+ group but significantly decreased from 20.6 to 17.4 ng/mL in the IBN group. Additionally, both groups exhibited significant increases in mean serum levels of PTH but significant decreases in serum levels of bone-specific alkaline phosphatase and C-telopeptide of type 1 collagen (CTX) at 16 weeks; no significant differences were observed between the groups. However, in subjects with a vitamin D deficiency, IBN+ treatment resulted in a significant decrease in serum CTX levels compared with IBN treatment. CONCLUSION: The present findings demonstrate that a once-monthly pill containing ibandronate and cholecalciferol may be useful for the amelioration of vitamin D deficiency in patients with postmenopausal osteoporosis. Moreover, this treatment combination effectively decreased serum levels of resorption markers, especially in subjects with a vitamin D deficiency, over the 16-week treatment period.
Alkaline Phosphatase ; Cholecalciferol* ; Collagen Type I ; Female ; Humans ; Osteoporosis* ; Osteoporosis, Postmenopausal ; Parathyroid Hormone ; Prospective Studies ; Vitamin D Deficiency

Dystonia is a movement disorder characterized by involuntary muscle contractions. Patients with dystonia may experience uncontrollable twisting, repetitive movements, or abnormal posture. A 55-year-old man presented with an involuntary left forearm supination, which he had experienced for five years. There was no history of antecedent trauma to the wrist or elbow. Although conventional therapeutic modalities had been performed, the symptoms persisted. When he visited our hospital, electromyography was performed. Reduced conduction velocity was evident at the elbow-axilla segment of the left median nerve. We suspected that there was a problem on the median nerve between the elbow and the axilla. For this reason, we performed an ultrasonography and magnetic resonance imaging study. A spindle-shaped soft tissue mass was observed at the left median nerve that suggested the possibility of neurofibroma. Dystonia caused by traumatic or compressive peripheral nerve injury has often been reported, but focal dystonia due to a neurogenic tumor is extremely rare. Here, we report our case with a review of the literature.
Axilla ; Dystonia ; Dystonic Disorders* ; Elbow ; Electromyography ; Forearm ; Humans ; Magnetic Resonance Imaging ; Median Nerve ; Median Neuropathy ; Middle Aged ; Movement Disorders ; Muscle, Smooth ; Neurofibroma ; Peripheral Nerve Injuries ; Posture ; Supination ; Ultrasonography ; Wrist

OBJECTIVES: The purpose of this study was to classify determinants of cost increases into two categories, negotiable factors and non-negotiable factors, in order to identify the determinants of health care expenditure increases and to clarify the contribution of associated factors selected based on a literature review. METHODS: The data in this analysis was from the statistical yearbooks of National Health Insurance Service, the Economic Index from Statistics Korea and regional statistical yearbooks. The unit of analysis was the annual growth rate of variables of 16 cities and provinces from 2003 to 2010. First, multiple regression was used to identify the determinants of health care expenditures. We then used hierarchical multiple regression to calculate the contribution of associated factors. The changes of coefficients (R2) of predictors, which were entered into this analysis step by step based on the empirical evidence of the investigator could explain the contribution of predictors to increased medical cost. RESULTS: Health spending was mainly associated with the proportion of the elderly population, but the Medicare Economic Index (MEI) showed an inverse association. The contribution of predictors was as follows: the proportion of elderly in the population (22.4%), gross domestic product (GDP) per capita (4.5%), MEI (-12%), and other predictors (less than 1%). CONCLUSIONS: As Baby Boomers enter retirement, an increasing proportion of the population aged 65 and over and the GDP will continue to increase, thus accelerating the inflation of health care expenditures and precipitating a crisis in the health insurance system. Policy makers should consider providing comprehensive health services by an accountable care organization to achieve cost savings while ensuring high-quality care.
Cities ; Health Expenditures/*statistics & numerical data/trends ; Humans ; Regression Analysis ; Republic of Korea

BACKGROUND: Among the various methods for correcting nasal deformity, the composite graft is suitable for the inner and outer reconstruction of the nose in a single stage. In this article, we present our technique for reconstructing the ala and columella using the auricular chondrocutaneous composite graft. METHODS: From 2004 to 2011, 15 cases of alar and 2 cases of columellar reconstruction employing the chondrocutaneous composite graft were studied, all followed up for 3 to 24 months (average, 13.5 months). All of the patients were reviewed retrospectively for the demographics, graft size, selection of the donor site and outcomes including morbidity and complications. RESULTS: The reasons for the deformity were burn scar (n=7), traumatic scar (n=4), smallpox scar (n=4), basal cell carcinoma defect (n=1), and scar contracture (n=1) from implant induced infection. In 5 cases of nostril stricture and 6 cases of alar defect and notching, composite grafts from the helix were used (8.9x12.5 mm). In 4 cases of retracted ala, grafts from the posterior surface of the concha were matched (5x15 mm). For the reconstruction of the columella, we harvested the graft from the posterior scapha (9x13.5 mm). Except one case with partial necrosis and delayed healing due to smoking, the grafts were successful in all of the cases and there was no deformity of the donor site. CONCLUSIONS: An alar and columellar defect can be reconstructed successfully with a relatively large composite graft without donor site morbidity. The selection of the donor site should be individualized according to the 3-dimensional configuration of the defect.
Burns ; Carcinoma, Basal Cell ; Cicatrix ; Congenital Abnormalities ; Constriction, Pathologic ; Contracture ; Demography ; Ear ; Humans ; Necrosis ; Nose ; Retrospective Studies ; Smallpox ; Smoke ; Smoking ; Succinates ; Tissue Donors ; Transplants

Previous studies have demonstrated that rottlerin, a specific PKCdelta inhibitor, potentiates death receptor- mediated apoptosis through a cytochrome c-dependent or -independent pathway. However, its ability to regulate necrotic cell death, as well as the underlying mechanism, remains unknown. We found that in murine fibrosarcoma L929 cells, treatment with rottlerin protected the cells against TNF-induced necrosis, whereas it sensitized the cells to apoptosis induced by co-treatment with Hsp90 inhibitor geldanamycin and TNF, in a manner independent of its ability to inhibit PKC-delta. TNF treatment induced rapid accumulation of mitochondrial superoxide (O2") through the Nox1 NADPH oxidase when cells undergo necrosis. Moreover, pretreatment with rottlerin failed to induce the GTP-bound form of small GTPase Rac1 by TNF treatment, and subsequently suppressed mitochondrial O2(-) production and poly(ADP-ribose) polymerase activation, thus inhibiting necrotic cell death. Therefore, our study suggests that Nox1 NADPH oxidase is a new molecular target for anti-necrotic activity of rottlerin upon death-receptor ligation.
Acetophenones/*pharmacology ; Animals ; Benzopyrans/*pharmacology ; Cell Death/*drug effects ; Cell Line, Tumor ; Mice ; Protein Kinase Inhibitors/*pharmacology ; Superoxides/metabolism ; Tumor Necrosis Factor-alpha/*antagonists & inhibitors/pharmacology

BACKGROUND: Minimal (subclinical) hepatic encephalopathy (mHE) currently diagnosed by psychometric tests or neurophysiological test adversely affects daily functioning. In view of its sociomedical relevance, simple and reproducible tests for routine diagnosis are required. The aims of this study are to evaluate cognitive function of patients with chronic liver disease by computerized neuropsychological test (STIM), and the difference of cognitive function according to Child classification. METHODS: Between June, 2002 and February, 2003 We enrolled 61 randomized consecutive patients diagnosed with chronic liver disease by biochemical tests, ultrasonographic finding or histology. This study used finger tapping, visual CPT, spatial memory test, Wisconsin card sorting test chosen from Neuscan and STIM system (Neurosoft company, U.S.A) and global-local processing test. RESULTS: In the present study, significant correlation was found between neurologic abnormalities and the degree of liver disease. The result of neuropsychological test showed that cognitive function was decreased according to the severity of chronic liver disease, especially liver cirrhosis. Cirrhotic patients, especially Child C group, exhibited selective deficits in complex attentional and fine motor skills, visuospatial perception, with preservation of memory. CONCLUSION: The STIM in this study is simple, objective and reproducible method because it can subdivide evaluation of cognitive function and computerize the measurement of response. We assume that STIM may be used early detection method of mHE if the study will be in a large scale. Because psychomotor deficits found in mHE could have a disadvanting influence on daily functioning of patients, e.g., driving abilty of a car or performance at work, we concluded early detection of mHE and aggressive treatment of mHE in clinically asymptomatic cirrhotic patients is necessary for improvement of their quality of life.
Child ; Classification ; Diagnosis ; Fingers ; Hepatic Encephalopathy ; Humans ; Liver Cirrhosis ; Liver Diseases* ; Liver* ; Memory ; Motor Skills ; Neuropsychological Tests* ; Psychometrics ; Quality of Life ; Wisconsin