Immature mandarin orange extract increases the amount of hyaluronic acid in human skin fibroblast and keratinocytes.

Immature mandarin orange is thinned in order to maturation of orange. To use immature mandarin orange as a cosmetic functional material, we investigated the seasonal fluctuation changes in hesperidin and narirutin levels of immature mandarin oranges, and the effects on human skin cells. The contents of hesperidin from Aoshima, Otsu, and Shonan gold, is higher at about a month after flowering. Shonan gold has higher content of narirutin to compere that of Aoshima and Otsu. We found the addition of immature mandarin orange extracts to the human skin fibroblasts and keratinocytes, gene expression level of hyaluronic acid synthase and the hyaluronic acid contents in the medium are higher than that of the control. It was suggested that hesperidin in immature mandarin orange enhances the ability of skin cells to produce hyaluronic acid. Our findings indicate that the immature mandarin orange is a characteristic material on cosmetics and functional foods.

Preoperative sonographic evaluation of the defect size and the diaphragm rim in congenital diaphragmatic hernia - preliminary experience.

BACKGROUND: Sonographic assessment before congenital diaphragmatic hernia repair has rarely been studied. OBJECTIVE: To evaluate the accuracy of preoperative ultrasound in measuring the defect size and in anticipating the presence of a rim and thereby to determine ultrasound's usefulness in informing the surgical approach for definitive repair of congenital diaphragmatic hernia. MATERIALS AND METHODS: We performed a retrospective review of the medical records of seven children with left congenital diaphragmatic hernia who had undergone ultrasound and definitive repair between 2014 and 2017 at our institution. RESULTS: The estimated defect size by ultrasound to the actual defect size measured intraoperatively for each case were as follows: 23 × 25 mm to 20 × 26 mm (case 1); 23 × 30 mm to 20 × 30 mm (case 2); 43 × 25 mm to 30 × 30 mm (case 3); 21 × 23 mm to 20 × 25 mm (case 4); 19 × 24 mm to 10 × 30 mm (case 5); 32 × 33 mm to 30 × 50 mm (case 6); and almost total absence to 40 × 50 mm (case 7). Presence or absence of each part of the diaphragm rim evaluated by ultrasound was almost identical with the actual intraoperative findings. According to the ultrasound findings, we performed a successful thoracoscopic repair in cases 1-5 with relatively small defects and presence of all parts of the rim or absence of only posterolateral rim. CONCLUSION: There was good concordance between ultrasound findings and operative findings regarding the size of the defect and presence or absence of the diaphragm rim.

Expansion of functional human mucosal-associated invariant T cells via reprogramming to pluripotency and redifferentiation.

Mucosal-associated invariant T (MAIT) cells play an important physiological role in host pathogen defense and may also be involved in inflammatory disorders and multiple sclerosis. The rarity and inefficient expansion of these cells have hampered detailed analysis and application. Here, we report an induced pluripotent stem cell (iPSC)-based reprogramming approach for the expansion of functional MAIT cells. We found that human MAIT cells can be reprogrammed into iPSCs using a Sendai virus harboring standard reprogramming factors. Under T cell-permissive conditions, these iPSCs efficiently redifferentiate into MAIT-like lymphocytes expressing the T cell receptor Vα7.2, CD161, and interleukin-18 receptor chain α. Upon incubation with bacteria-fed monocytes, the derived MAIT cells show enhanced production of a broad range of cytokines. Following adoptive transfer into immunocompromised mice, these cells migrate to the bone marrow, liver, spleen, and intestine and protect against Mycobacterium abscessus. Our findings pave the way for further functional analysis of MAIT cells and determination of their therapeutic potential.

Paradoxical surge of corticotropin after glucocorticoid replacement in central adrenal insufficiency.

A 78-yr-old man was admitted in emergency with fatigue, anorexia, vomiting, hypothermia (35.1 °C on a hot August day), hypotension (89/56 mmHg) and hyponatraemia (126 mEq/l). Plasma corticotropin and cortisol were severely depressed: 0.84 pmol/L and 33.1 nmol/L respectively (reference range, 1.5-13.9 pmol/L and 110-505 nmol/L, respectively). Thyroid stimulating hormone was low-normal and free-triiodothyronine and free-thyroxine were subnormal. Magnetic resonance imaging revealed swelling of the pituitary gland and the stalk. The patient recovered after glucocorticoid replacement (200 mg/day intravenous hydrocortisone on Day 1 followed by tapering). Central diabetes insipidus which had become apparent had been treated with 1-desamino-8-D-arginine vasopressin. A surge of corticotropin and cortisol, 19.4 pmol/L and 712.1 nmol/L respectively, was found on Day 5 when luteinizing hormone, follicle stimulating hormone, and testosterone were subnormal and prolactin was slightly elevated. Subsequently, corticotropin and cortisol levels normalized together with normalization of luteinizing hormone, follicle stimulating hormone, anti-diuretic hormone, thyroid stimulating hormone, prolactin, testosterone and thyroid hormone levels. Shrinkage of the pituitary gland occurred after one month. Serum immunoglobulin G4 was elevated (3.21 and 6.02 g/l at 1- and 3-month follow-ups respectively). In conclusion, a paradoxical surge of corticotropin after glucocorticoid replacement was observed in a patient with central adrenal insufficiency due to immunoglobulin G4-related hypophysitis. Surge of ACTH in central adrenal insufficiency after glucocorticoid replacement has rarely been reported, and this is the second such case report.

Pancreatic nonfunctioning neuroendocrine tumor with the main pancreatic duct obstruction presenting as excessive hyperglycemia: a case report and review of the literature.

We present the case of a 65-year-old man with a pancreatic nonfunctioning neuroendocrine tumor causing main pancreatic duct obstruction that presented as excessive hyperglycemia. We considered the tumor elicited worsening of diabetes in this case, and we performed review of the relevant literature.

Development of monoclonal antibodies (MAbs) to feline interferon (fIFN)-γ as tools to evaluate cellular immune responses to feline infectious peritonitis virus (FIPV).

Feline infectious peritonitis virus (FIPV) can cause a lethal disease in cats, feline infectious peritonitis (FIP). The antibody-dependent enhancement (ADE) of FIPV infection has been recognised in experimentally infected cats, and cellular immunity is considered to play an important role in preventing the onset of FIP. To evaluate the importance of cellular immunity for FIPV infection, monoclonal antibodies (MAbs) against feline interferon (fIFN)-γ were first created to establish fIFN-γ detection systems using the MAbs. Six anti-fIFN-γ MAbs were created. Then, the difference in epitope which those MAbs recognise was demonstrated by competitive enzyme-linked immunosorbent assay (ELISA) and IFN-γ neutralisation tests. Detection systems for fIFN-γ (sandwich ELISA, ELISpot assay, and two-colour flow cytometry) were established using anti-fIFN-γ MAbs that recognise different epitopes. In all tests, fIFN-γ production from peripheral blood mononuclear cells (PBMCs) obtained from cats experimentally infected with an FIPV isolate that did not develop the disease was significantly increased by heat-inactivated FIPV stimulation in comparison with medium alone. Especially, CD8(+)fIFN-γ(+) cells, but not CD4(+)fIFN-γ(+) cells, were increased. In contrast, fIFN-γ production from PBMCs isolated from cats that had developed FIP and specific pathogen-free (SPF) cats was not increased by heat-inactivated FIPV stimulation. These results suggest that cellular immunity plays an important role in preventing the development of FIP. Measurement of fIFN-γ production with the anti-fIFN-γ MAbs created in this study appeared to be useful in evaluating cellular immunity in cats.

Screening and identification of T helper 1 and linear immunodominant antibody-binding epitopes in the spike 2 domain and the nucleocapsid protein of feline infectious peritonitis virus.

The antibody-dependent enhancement (ADE) of feline infectious peritonitis virus (FIPV) infection has been recognized in experimentally infected cats, and cellular immunity is considered to play an important role in preventing the onset of feline infectious peritonitis (FIP). In the present study, we synthesized eighty-one kinds of peptides derived from the spike (S)2 domain of type I FIPV KU-2 strain, the S2 domain of type II FIPV 79-1146 strain, and the nucleocapcid (N) protein of FIPV KU-2 strain. To detect the T helper (Th)1 epitope, peripheral blood mononuclear cells (PBMCs) obtained from FIPV-infected cats were cultured with each peptide, and Th1-type immune responses were measured using feline interferon (fIFN)-γ production as an index. To detect the linear immunodominant antibody-binding epitope, we investigated the reactivity of plasma collected from FIPV-infected cats against each peptide by ELISA. Four and 2 peptides containing Th1 epitopes were identified in the heptad repeat (HR)1 and inter-helical (IH) regions of the S2 domain of type I FIPV, respectively, and these were located on the N-terminal side of the regions. In the S2 domain of type II FIPV, 2, 3, and 2 peptides containing Th1 epitopes were identified in the HR1, IH, and HR2 regions, respectively, and these were mainly located on the C-terminal side of the regions. In the S2 domain of type I FIPV, 3 and 7 peptides containing linear immunodominant antibody-binding epitopes were identified in the IH and HR2 regions, respectively. In the S2 domain of type II FIPV, 4 peptides containing linear immunodominant antibody-binding epitopes were identified in the HR2 region. The Th1 epitopes in the S2 domain of type I and II FIPV were located in different regions, but the linear immunodominant antibody-binding epitopes were mostly located in the HR2 region. Eight peptides containing Th1 epitopes were identified in N protein, and 3 peptides derived from residues 81 to 100 and 137 to 164 showed strong inductivity of fIFN-γ production in PBMCs isolated from type I FIPV- and type II FIPV-infected non-FIP cats. In N protein, 4 peptides containing linear immunodominant antibody-binding epitopes were identified, and 2 peptides derived from residues 345 to 372 showed strong reactivity with plasma of type I FIPV- and type II FIPV-infected cats. The Th1 and linear immunodominant antibody-binding epitopes were located at different positions in both the S2 domain and N protein. Our results may provide important information for the development of peptide-based vaccine against FIPV infection.

Class III beta-tubulin expression in tumor cells is correlated with resistance to docetaxel in patients with completely resected non-small-cell lung cancer.

OBJECTIVE: To assess the relationship between in vitro chemosensitivity evaluated by the histoculture drug response assay (HDRA) and the expression of beta-tubulin isotypes in tumors of patients with completely resected NSCLC in order to determine the predictive value of beta-tubulin in chemotherapy for NSCLC. METHODS: Expression of beta-tubulin isotypes was immunohistochemically analyzed in a series of 58 tumor samples from patients with completely resected NSCLC. The sensitivity of individual tumors to anticancer agents was evaluated by HDRA. RESULTS: Class III beta-tubulin expression by tumor cells was significantly correlated with resistance to docetaxel (p=0.0250), but not related with resistance to gemcitabine. Patient characteristics (age, gender, histology, and stage) were not associated with class III beta-tubulin expression. CONCLUSION: An abundance of class III beta-tubulin in tumor cells could be a biomarker for resistance to docetaxel in patients with completely resected NSCLC.

In vitro evaluation of dose-response curve for paclitaxel in breast cancer.

BACKGROUND: The dose-response curve for anticancer agents cannot be evaluated by studying patients directly. To investigate individual differences in the dose-response curve for paclitaxel in breast cancer, we utilized the histoculture drug response assay (HDRA) technique. MATERIALS AND METHODS: Twenty specimens obtained from breast cancer patients who underwent surgical resection were used in this study. The inhibition rates of paclitaxel at several concentrations were measured and fitted to a sigmoid dose-response curve, using non-linear least squares analysis with the fitting equation y=A(1-1/(1+exp(B(x-log(C))))), where A denotes maximal response; B, slope factor; and C, ED50. RESULTS: A dose-response curve was obtained in all tumors. The mean value (+/-SD) of maximum response, slope factor, and ED50 were 90.2+/-5.5%, 9.4+/-4.3, and 36.8+/-17.2 microg/ml, respectively. The slope factor was higher in nuclear grade 3 tumors compared with nuclear grade 1 and 2 tumors. CONCLUSION: An individual dose-response curve for paclitaxel in breast cancer can be obtained using the HDRA technique. Nuclear grade 3 tumors appeared to have more uniform chemosensitivity to paclitaxel compared with nuclear grade 1 and 2 tumors.

Data acquisition for the histoculture drug response assay in lung cancer.

OBJECTIVE: Application of the histoculture drug response assay for lung cancer was investigated by using data acquired from lung cancer specimens. METHODS: From May 1994 through February 2005, histoculture drug response assay data were obtained from 359 lung cancer specimens held in our institute. We examined chemosensitivities of the tissues to cisplatin, doxorubicin, mitomycin C, 5-fluorouracil, docetaxel, paclitaxel, etoposide, irinotecan, and gemcitabine. Cutoff inhibition rates were determined with each drug for non-small cell lung cancer and were used to calculate predictabilities for chemotherapy responses. RESULTS: The evaluability of the histoculture drug response assay was high at 97.4%. Good predictability, including true-positive and true-negative rates of 73.2% and 100%, respectively, with an accuracy of 83.0%, was observed. CONCLUSION: The histoculture drug response assay appears to be applicable to non-small cell lung cancer for the prediction of responses to chemotherapy.

Evaluation of optimal drug concentration in histoculture drug response assay in association with clinical efficacy for head and neck cancer.

Induction chemotherapy or concomitant chemoradiotherapy has been used increasingly to improve survival, organ preservation and function in patients with head and neck cancer (HNC). However, this regimen encounters significant side effects with potential adverse reactions causing many disadvantages for patients. Therefore, reliable chemosensitivity assays are needed to accurately predict the response to chemotherapy and guide the selection and treatment of patients with HNC. The main purpose of this study was to examine the optimal drug concentrations for evaluating in vitro chemosensitivity using the histoculture drug response assay (HDRA). The tested tumor specimens included 7 from oral cavities (14.3%), 12 from oropharynx (24.5%), 10 hypopharynx (20.4%), 3 larynx (6.1%), 5 sinonasal (10.2%), 2 salivary glands (4.1%), and 10 from metastatic lymph nodes (20.4%), respectively. Histopathologic types of all 49 specimens were squamous cell carcinoma. We investigated the optimal drug concentrations in HDRA searching at doses of 4-100 microg/ml for cisplatin and 60-1500 microg/ml for 5-FU. We considered the concentration of 20 microg/ml to be appropriate for evaluating cisplatin sensitivity in HNC among the tested dosages. As for cisplatin sensitivity in vitro, the 50% cut-off inhibition index (I.I.) was found to have a significant association with the clinical response to chemotherapy, with an accurate prediction rate of 77.8%. The HDRA shows a predictive value for chemosensitivity in HNC patients using the optimal drug concentration cut-off with this site specificity.

Relationship between the contamination of the nurse's caps and their period of use in terms of microorganism numbers.

Nosocomial infections are a great problem in the health care facilities. The white uniforms of nurses are often washed to keep them clean, but the nurse's caps are not washed as frequently in comparison. It could be that the importance of these caps is being overlooked. If these caps are providing a residence for microorganisms causing nosocomial infection in the health care facility, then they should be washed as frequently as the uniforms. So far, the relationship between the contamination of the nurse's caps and nosocomial infection has not yet been studied. Therefore, this study was conducted to confirm if relationships exist among factors regarding the number of microorganisms on the nurse's caps, the period in which caps were used without being washed, and the individual characteristics of nurse wearing the caps. Results showed that the degree of contamination of the nurse's caps depended on individual characteristics and the period of use. Finally, results led to the conclusion that the nurse's caps should not be worn if their only purpose is to symbolize female workers in the health care facilities because, in actually, they provide a resistance for microorganisms causing nosocomial infections.

[Cut-off level of docetaxel, paclitaxel and gemcitabine in histoculture drug response assay for non-small cell lung cancer].

Cut-off levels of docetaxel (DOC), paclitaxel (PAC), and gemcitabine (GEM) in histoculture drug response assay are determined by data acquisition of non-small cell lung cancer. Inhibition rates were 47.5 +/- 22.2% in DOC (n=181), 66.6 +/- 25.1% in PAC (n=57), and 25.4 +/- 18.4% in GEM (n=63), respectively. Cut-off levels were determined as 50% in DOC, 60% in PAC, and 30% in GEM. The positive rates such as 47.5% in DOC, 68.4% in PAC, and 33.3% in GEM were obtained.

[Dose response curve of paclitaxel measured by histoculture drug response assay].

Dose response curves of paclitaxel were measured by histoculture drug response assay (HDRA) in 11 lung cancer patients. Inhibition rates of paclitaxel at several concentrations were measured and fitted to the sigmoid dose response curve, using non-linear least square analysis, with fitting equation y=A (1-1/(1+exp (b (x-log (ED50)). Parameters A, b, and ED50 were 88.3+/-6.0 (80.0-100.0) %, 9.57+/-4.32 (2.25-15.0), and 26.8+/-8.1 (15.0-41.0) microg/ml, respectively. The parameter b was lower in well-differentiated tumors compared with moderately and poorly-differentiated tumors. Dose response curves of paclitaxel could be measured by HDRA in lung cancer. This method provides us more information for drug sensitivity than the usual HDRA method. This may lead to the improved accuracy of HDRA.

Sequence analysis of amplified DNA fragments containing the region encoding the putative lipase substrate-binding domain and genotyping of Aeromonas hydrophila.

DNA fragments were amplified by PCR from all tested strains of Aeromonas hydrophila, A. caviae, and A. sobria with primers designed based on sequence alignment of all lipase, phospholipase C, and phospholipase A1 genes and the cytotonic enterotoxin gene, all of which have been reported to have the consensus region of the putative lipase substrate-binding domain. All strains showed lipase activity, and all amplified DNA fragments contained a nucleotide sequence corresponding to the substrate-binding domain. Thirty-five distinct nucleotide sequence patterns and 15 distinct deduced amino acid sequence patterns were found in the amplified DNA fragments from 59 A. hydrophila strains. The deduced amino acid sequences of the amplified DNA fragments from A. caviae and A. sobria strains had distinctive amino acids, suggesting a species-specific sequence in each organism. Furthermore, the amino acid sequence patterns appear to differ between clinical and environmental isolates among A. hydrophila strains. Some strains whose nucleotide sequences were identical to one another in the amplified region showed an identical DNA fingerprinting pattern by repetitive extragenic palindromic sequence-PCR genotyping. These results suggest that A. hydrophila, and also A. caviae and A. sobria strains, have a gene encoding a protein with lipase activity. Homologs of the gene appear to be widely distributed in Aeromonas strains, probably associating with the evolutionary genetic difference between clinical and environmental isolates of A. hydrophila. Additionally, the distinctive nucleotide sequences of the genes could be attributed to the genotype of each strain, suggesting that their analysis may be helpful in elucidating the genetic heterogeneity of Aeromonas.