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1.  Acute Liver Injury in a Patient with Alcohol Dependence: A Case Resembling Autoimmune Hepatitis or Drug-Induced Liver Injury 
Case Reports in Gastroenterology  2014;8(1):129-133.
Some patients with alcohol dependence may initially present with atypical laboratory and histological features resembling autoimmune hepatitis (AIH) or drug-induced liver injury (DILI). Even with liver biopsy, it may be difficult to diagnose certain patients with alcohol dependence. However, careful follow-up of our patient and consultations with the attending psychiatrist were successful in diagnosing alcohol dependence and its liver injury. The immune mechanisms of alcoholic liver diseases, AIH and DILI may be overlapping. Certain patients are suffering from AIH with flares on a background of alcohol abuse. Certain patients with alcohol abuse may have a past history of DILI. This might be consistent with the fact that alcohol dependence initially presents with atypical laboratory features of AIH or DILI. With careful observation, the clinician should remind himself that alcohol dependence is not always required for developing liver disease, since many patients with liver disease do not meet the criteria for alcohol dependence.
doi:10.1159/000362442
PMCID: PMC4025146  PMID: 24847195
Acute liver injury; Alcohol dependence; Autoimmune hepatitis; Drug-induced liver injury
2.  Epidermal Cyst of the Breast Treated by Vacuum-Assisted Biopsy 
International Surgery  2013;98(1):65-69.
A 38-year-old woman presented for evaluation of a firm mass (measuring 20 × 20 mm) in the upper outer quadrant of her left breast. On the basis of the clinical and radiologic findings, we diagnosed a benign breast tumor and scheduled removal by a hand-held vacuum-assisted biopsy device (VABD) under ultrasonographic guidance. Because the first specimen removed from the tumor was white, flaky, and waxy material, we strongly suspected that the lesion was an epidermal cyst. We continued VABD treatment until the tumor was invisible on real-time ultrasonography. Histologic examination demonstrated that the tumor was composed of mature stratified squamous epithelium and laminated layers of keratin, findings consistent with a diagnosis of epidermal cyst. These cysts rarely occur in the breast and are sometimes difficult to distinguish from breast cancer. To our knowledge, this is the first report of an epidermal cyst treated by VABD.
doi:10.9738/CC74.1
PMCID: PMC3723168  PMID: 23438279
Epidermal cyst; Breast; Vacuum-assisted biopsy device (VABD)
3.  A New Protocol to Detect Multiple Foodborne Pathogens with PCR Dipstick DNA Chromatography after a Six-Hour Enrichment Culture in a Broad-Range Food Pathogen Enrichment Broth 
BioMed Research International  2013;2013:295050.
A quick foodborne pathogen screening method after six-hour enrichment culture with a broad-range food pathogen enrichment broth is described. Pathogenic factors of Salmonella enterica, Shigella spp., enteroinvasive Escherichia coli, and enterohemorrhagic E. coli are amplified with a cocktail primer and rapid polymerase chain reaction (PCR), which finishes amplification in 30 min. The PCR amplicon was differentiated with a dipstick DNA chromatography assay in 5–10 min. Starting from a four- to six-hour enrichment culture, this assay was finished within 45 min. Detection sensitivity of this protocol was less than 2.5 CFU/25 g for S. enterica and 3.3 CFU/25 g for enterohemorrhagic E. coli in spiked ground meat experiments.
doi:10.1155/2013/295050
PMCID: PMC3865639  PMID: 24364031
4.  Versatile Transformation System That Is Applicable to both Multiple Transgene Expression and Gene Targeting for Thraustochytrids 
A versatile transformation system for thraustochytrids, a promising producer for polyunsaturated fatty acids and fatty acid-derived fuels, was established. G418, hygromycin B, blasticidin, and zeocin inhibited the growth of thraustochytrids, indicating that multiple selectable marker genes could be used in the transformation system. A neomycin resistance gene (neor), driven with an ubiquitin or an EF-1α promoter-terminator from Thraustochytrium aureum ATCC 34304, was introduced into representatives of two thraustochytrid genera, Aurantiochytrium and Thraustochytrium. The neor marker was integrated into the chromosomal DNA by random recombination and then functionally translated into neor mRNA. Additionally, we confirmed that another two genera, Parietichytrium and Schizochytrium, could be transformed by the same method. By this method, the enhanced green fluorescent protein was functionally expressed in thraustochytrids. Meanwhile, T. aureum ATCC 34304 could be transformed by two 18S ribosomal DNA-targeting vectors, designed to cause single- or double-crossover homologous recombination. Finally, the fatty acid Δ5 desaturase gene was disrupted by double-crossover homologous recombination in T. aureum ATCC 34304, resulting in an increase of dihomo-γ-linolenic acid (C20:3n-6) and eicosatetraenoic acid (C20:4n-3), substrates for Δ5 desaturase, and a decrease of arachidonic acid (C20:4n-6) and eicosapentaenoic acid (C20:5n-3), products for the enzyme. These results clearly indicate that a versatile transformation system which could be applicable to both multiple transgene expression and gene targeting was established for thraustochytrids.
doi:10.1128/AEM.07129-11
PMCID: PMC3346472  PMID: 22344656
5.  Increase of Eicosapentaenoic Acid in Thraustochytrids through Thraustochytrid Ubiquitin Promoter-Driven Expression of a Fatty Acid Δ5 Desaturase Gene▿† 
Applied and Environmental Microbiology  2011;77(11):3870-3876.
Thraustochytrids, marine protists known to accumulate polyunsaturated fatty acids (PUFAs) in lipid droplets, are considered an alternative to fish oils as a source of PUFAs. The major fatty acids produced in thraustochytrids are palmitic acid (C16:0), n − 6 docosapentaenoic acid (DPA) (C22:5n − 6), and docosahexaenoic acid (DHA) (C22:6n − 3), with eicosapentaenoic acid (EPA) (C20:5n − 3) and arachidonic acid (AA) (C20:4n − 6) as minor constituents. We attempted here to alter the fatty acid composition of thraustochytrids through the expression of a fatty acid Δ5 desaturase gene driven by the thraustochytrid ubiquitin promoter. The gene was functionally expressed in Aurantiochytrium limacinum mh0186, increasing the amount of EPA converted from eicosatetraenoic acid (ETA) (C20:4n − 3) by the Δ5 desaturase. The levels of EPA and AA were also increased by 4.6- and 13.2-fold in the transgenic thraustochytrids compared to levels in the mock transfectants when ETA and dihomo-γ-linolenic acid (DGLA) (C20:3n − 6) were added to the culture at 0.1 mM. Interestingly, the amount of EPA in the transgenic thraustochytrids increased in proportion to the amount of ETA added to the culture up to 0.4 mM. The rates of conversion and accumulation of EPA were much higher in the thraustochytrids than in baker's yeasts when the desaturase gene was expressed with the respective promoters. This report describes for the first time the finding that an increase of EPA could be accomplished by introducing the Δ5 desaturase gene into thraustochytrids and indicates that molecular breeding of thraustochytrids is a promising strategy for generating beneficial PUFAs.
doi:10.1128/AEM.02664-10
PMCID: PMC3127612  PMID: 21478316
6.  Impact of Free-Living Amoebae on Presence of Parachlamydia acanthamoebae in the Hospital Environment and Its Survival In Vitro without Requirement for Amoebae▿  
Journal of Clinical Microbiology  2010;48(9):3360-3365.
Parachlamydia acanthamoebae is an obligately intracellular bacterium that infects free-living amoebae and is a potential human pathogen in hospital-acquired pneumonia. We examined whether the presence of P. acanthamoebae is related to the presence of Acanthamoeba in an actual hospital environment and assessed the in vitro survival of P. acanthamoebae. Ninety smear samples were collected between November 2007 and March 2008 (trial 1, n = 52) and between October 2008 and February 2009 (trial 2, n = 38) from the floor (dry conditions, n = 56) and sink outlets (moist conditions, n = 34) of a hospital. The prevalences of P. acanthamoebae DNA in the first and second trials were 64.3% and 76%, respectively. The prevalences of Acanthamoeba DNA in the first and second trials were 48% and 63.1%, respectively. A statistical correlation between the prevalence of P. acanthamoebae and that of Acanthamoeba was found (trial 1, P = 0.011; trial 2, P = 0.022), and that correlation increased when samples from just the dry area (floor smear samples, P = 0.002) were analyzed but decreased when samples from a moist area were analyzed (P = 0.273). The in vitro experiment showed that, without Acanthamoeba, P. acanthamoebae could not survive in dry conditions for 3 days at 30°C or 15 days at 15°C. Thus, both organisms were coincidentally found in an actual hospital environment, with the presence of Acanthamoeba having a significant effect on the long-term survival of P. acanthamoebae, suggesting that this potential human pathogen could spread through a hospital environment via Acanthamoeba.
doi:10.1128/JCM.00366-10
PMCID: PMC2937724  PMID: 20631104
7.  Human Artificial Chromosome with a Conditional Centromere for Gene Delivery and Gene Expression 
Human artificial chromosomes (HACs), which carry a fully functional centromere and are maintained as a single-copy episome, are not associated with random mutagenesis and offer greater control over expression of ectopic genes on the HAC. Recently, we generated a HAC with a conditional centromere, which includes the tetracycline operator (tet-O) sequence embedded in the alphoid DNA array. This conditional centromere can be inactivated, loss of the alphoidtet-O (tet-O HAC) by expression of tet-repressor fusion proteins. In this report, we describe adaptation of the tet-O HAC vector for gene delivery and gene expression in human cells. A loxP cassette was inserted into the tet-O HAC by homologous recombination in chicken DT40 cells following a microcell-mediated chromosome transfer (MMCT). The tet-O HAC with the loxP cassette was then transferred into Chinese hamster ovary cells, and EGFP transgene was efficiently and accurately incorporated into the tet-O HAC vector. The EGFP transgene was stably expressed in human cells after transfer via MMCT. Because the transgenes inserted on the tet-O HAC can be eliminated from cells by HAC loss due to centromere inactivation, this HAC vector system provides important novel features and has potential applications for gene expression studies and gene therapy.
doi:10.1093/dnares/dsq020
PMCID: PMC2955713  PMID: 20798231
human artificial chromosome; conditional centromere; gene delivery
8.  Contrast-enhanced intraoperative ultrasonography equipped with late Kupffer-phase image obtained by sonazoid in patients with colorectal liver metastases 
AIM: To find occult metastases during hepatectomy in patients with colorectal cancer liver metastases (CRCLM), contrast-enhanced intraoperative ultrasonography (CE-IOUS) was performed using a new microbubble agent, sonazoid, which provides a parenchyma-specific contrast image based on its accumulation in the Kupffer cells.
METHODS: Eight patients with CRCLM underwent CE-IOUS using sonazoid before hepatectomy. The liver was investigated during a late Kupffer-phase imaging, which is a valuable characteristic of sonazoid.
RESULTS: CE-IOUS using sonazoid provided the early vascular- and sinusoidal-phase images for
10 min followed by the late Kupffer-phase image up to 30 min after the injection of sonazoid. IOUS did not provide new findings of metastatic lesion in the 8 patients. However, during the late Kupffer-phase image of sonazoid, a metastatic lesion was newly found in two of the 8 patients. These newly detected lesions were removed by an additional hepatectomy and histopathologically diagnosed as a metastasis.
CONCLUSION: CE-IOUS using sonazoid can allow surgeons to investigate the whole liver with enough time and to find new metastases intraoperatively.
doi:10.3748/wjg.14.3207
PMCID: PMC2712854  PMID: 18506927
Contrast-enhanced intraoperative ultrasonography; Late Kupffer-phase image; Sonazoid; Colorectal cancer liver metastases; Occult hepatic lesions

Results 1-8 (8)