Discordance between hyposalivation and xerostomia

Individuals with an objective decrease in salivary flow (objective dry mouth) may not be aware of subjective dry mouth (xerostomia). However, no clear evidence exists to explain the discordance between subjective and objective dry mouth. Therefore, this cross-sectional study aimed to assess the prevalence of xerostomia and decreased salivary flow among community-dwelling elderly adults. In addition, this study assessed several potential demographic and health status determinants of the discrepancy between xerostomia and reduced salivary flow. The 215 participants in this study were community-dwelling older people aged 70 years and above who underwent dental health examinations between January-February 2019. Symptoms of xerostomia were collected in the form of a questionnaire. The unstimulated salivary flow rate (USFR) was measured by a dentist using visual inspection. The stimulated salivary flow rate (SSFR) was measured using the Saxon test. We identified 19.1% of participants as having mild-severe USFR decline with xerostomia and 19.1% as having mild-severe USFR decline without xerostomia. Additionally, 26.0% of participants had low SSFR and xerostomia, and 40.0% had low SSFR without xerostomia. Except for the age trend, no factors could be associated with the discordance between USFR measurement and xerostomia. Furthermore, no significant factors were associated with the discordance between the SSFR and xerostomia. However, females were significantly associated (OR = 2.608, 95% CI = 1.174–5.791) with low SSFR and xerostomia, as compared to males. Age was a factor that was also significantly associated (OR = 1.105, 95% CI = 1.010–1.209) with low SSFR and xerostomia. Our findings indicate that approximately 20% of the participants had low USFR without xerostomia, and 40% had low SSFR without xerostomia. This study showed that age, sex, and the number of medications may not be factors in the discrepancy between the subjective feeling of dry mouth and reduced salivary flow

Muscle Fiber Type-Predominant Promoter Activity in Lentiviral-Mediated Transgenic Mouse

Variations in gene promoter/enhancer activity in different muscle fiber types after gene transduction was noticed previously, but poorly analyzed. The murine stem cell virus (MSCV) promoter drives strong, stable gene expression in hematopoietic stem cells and several other cells, including cerebellar Purkinje cells, but it has not been studied in muscle. We injected a lentiviral vector carrying an MSCV-EGFP cassette (LvMSCV-EGFP) into tibialis anterior muscles and observed strong EGFP expression in muscle fibers, primary cultured myoblasts, and myotubes isolated from injected muscles. We also generated lentiviral-mediated transgenic mice carrying the MSCV-EGFP cassette and detected transgene expression in striated muscles. LvMSCV-EGFP transgenic mice showed fiber type-dependent variations in expression: highest in types I and IIA, intermediate in type IID/X, and lowest in type IIB fibers. The soleus and diaphragm muscles, consisting mainly of types I and IIA, are most severely affected in the mdx mouse model of muscular dystrophy. Further analysis of this promoter may have the potential to achieve certain gene expression in severely affected muscles of mdx mice. The Lv-mediated transgenic mouse may prove a useful tool for assessing the enhancer/promoter activities of a variety of different regulatory cassettes

Th17 cells differentiated with mycelial membranes of Candida albicans prevent oral candidiasis

Candida albicans is a human commensal that causes opportunistic infections. Th17 cells provide resistance against mucosal infection with C. albicans; however, the T cell antigens remain little known. Our final goal is to find effective T cell antigens of C. albicans that are responsible for immunotherapy against candidiasis. Here, we prepared fractions including cytosol, membrane and cell wall from yeast and mycelial cells. Proteins derived from a membrane fraction of mycelial cells effectively induced differentiation of CD4+ T cells into IL-17A-producing Th17 cells. To confirm the immunological response in vivo of proteins from mycelial membrane, we performed adoptive transfer experiments using ex vivo stimulated CD4+ T cells from IL-17A-GFP reporter mice. Mycelial membrane-differentiated CD4+ Th17 cells adoptively transferred intravenously prevented oral candidiasis by oral infection of C. albicans, compared with control anti-CD3-stimulated CD4+ T cells. This was confirmed by the clinical score and the number of neutrophils on the infected tissues. These data suggest that effective T cell antigens against candidiasis could be present in the membrane protein fraction of mycelial cells. The design of novel vaccination strategies against candidiasis will be our next step.福岡歯科大学2017年

Identification of a Novel Alternatively Spliced Form of Inflammatory Regulator SWAP-70-Like Adapter of T Cells

福岡歯科大学2016年

The Effect of Medical Cooperation in the CKD Patients: 10-Year Multicenter Cohort Study

Introduction: While chronic kidney disease (CKD) is one of the most important contributors to mortality from non-communicable diseases, the number of nephrologists is limited worldwide. Medical cooperation is a system of cooperation between primary care physicians and nephrological institutions, consisting of nephrologists and multidisciplinary care teams. Although it has been reported that multidisciplinary care teams contribute to the prevention of worsening renal functions and cardiovascular events, there are few studies on the effect of a medical cooperation system. Methods: We aimed to evaluate the effect of medical cooperation on all-cause mortality and renal prognosis in patients with CKD. One hundred and sixty-eight patients who visited the one hundred and sixty-three clinics and seven general hospitals of Okayama city were recruited between December 2009 and September 2016, and one hundred twenty-three patients were classified into a medical cooperation group. The outcome was defined as the incidence of all-cause mortality, or renal composite outcome (end-stage renal disease or 50% eGFR decline). We evaluated the effects on renal composite outcome and pre-ESRD mortality while incorporating the competing risk for the alternate outcome into a Fine-Gray subdistribution hazard model. Results: The medical cooperation group had more patients with glomerulonephritis (35.0% vs. 2.2%) and less nephrosclerosis (35.0% vs. 64.5%) than the primary care group. Throughout the follow-up period of 5.59 +/- 2.78 years, 23 participants (13.7%) died, 41 participants (24.4%) reached 50% decline in eGFR, and 37 participants (22.0%) developed end-stage renal disease (ESRD). All-cause mortality was significantly reduced by medical cooperation (sHR 0.297, 95% CI 0.105-0.835, p = 0.021). However, there was a significant association between medical cooperation and CKD progression (sHR 3.069, 95% CI 1.225-7.687, p = 0.017). Conclusion: We evaluated mortality and ESRD using a CKD cohort with a long-term observation period and concluded that medical cooperation might be expected to influence the quality of medical care in the patients with CKD

Nik-related kinase regulates trophoblast proliferation and placental development by modulating AKT phosphorylation

<div><p>Nik-related kinase (Nrk) is a Ser/Thr kinase and was initially discovered as a molecule that was predominantly detected in skeletal muscles during development. A recent study using <i>Nrk</i>-null mice suggested the importance of <i>Nrk</i> in proper placental development; however, the molecular mechanism remains unknown. In this study, we demonstrated that differentiated trophoblasts from murine embryonic stem cells (ESCs) endogenously expressed <i>Nrk</i> and that <i>Nrk</i> disruption led to the enhanced proliferation of differentiated trophoblasts. This phenomenon may reflect the overproliferation of trophoblasts that has been reported in enlarged placentas of <i>Nrk</i>-null mice. Furthermore, we demonstrated that AKT phosphorylation at Ser473 was upregulated in <i>Nrk</i>-null trophoblasts and that inhibition of AKT phosphorylation cancelled the enhanced proliferation observed in differentiated <i>Nrk</i>-null trophoblasts. These results indicated that the upregulation of AKT phosphorylation was the possible cause of enhanced proliferation observed in <i>Nrk</i>-null trophoblasts. The upregulation of AKT phosphorylation was also confirmed in enlarged <i>Nrk</i>-null placentas <i>in vivo</i>, suggesting that proper regulation of AKT by <i>Nrk</i> was important for normal placental development. In addition, our detailed analysis on phosphorylation status of AKT isoforms in newly established trophoblast stem cells (TSCs) revealed that different levels of upregulation of AKT phosphorylation were occurred in <i>Nrk</i>-null TSCs depending on AKT isoforms. These results further support the importance of <i>Nrk</i> in proper development of trophoblast lineage cells and indicate the possible application of TSCs for the analysis of differently regulated activation mechanisms of AKT isoforms.</p></div

Enhanced proliferation observed in differentiated <i>Nrk</i>-null trophoblasts was associated with the upregulation of AKT Ser473 phosphorylation.

<p>(A) Western blot analysis of whole-cell extracts from ESCs and trophoblasts. Expressions of NRK, phosphorylated AKT at Ser473, total AKT, phosphorylated ERK1/2 at Thr202/Tyr204 and total ERK1/2 were compared between wild-type and <i>Nrk</i>-null cells. As an internal control, β-Actin was also detected. The relative density of phosphorylated vs total AKT was analyzed and was normalized to the value of wild-type. (B) The schematic representation of the experimental procedure to examine the effect of AKT inhibitor MK-2206. (C) Dose dependent effect of MK-2206 on the proliferation and AKT Ser473 phosphorylation in the differentiated <i>Nrk</i>-null trophoblasts. The proliferation rate of the trophoblasts on day 4 was calculated relative to the untreated day 1 control (n = 4) (top). Data are presented as mean plus SD. **<i>P</i><0.01compared to wild-type. n.s. = no significant difference. Expression of phosphorylated AKT at Ser473 and total AKT in trophoblasts on day 4 was analyzed by western blotting (bottom). As an internal control, β-Actin was also detected. The relative density of phosphorylated vs total AKT was analyzed and was normalized to the value of untreated wild-type control. Shown are cropped blots. The blots with indicated cropping lines are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0171503#pone.0171503.s003" target="_blank">S3 Fig</a>. WB: western blotting, ESC: embryonic stem cell, TB: trophoblast.</p

Upregulation of AKT phosphorylation in enlarged placentas derived from <i>Nrk</i>-null E18.5 mouse embryos.

<p>(A) The wild-type <i>Nrk</i> allele and the targeted <i>Nrk</i> allele are shown (left). The arrows indicate PCR primers for detection of each allele. PCR genotyping of mice (right). (B) Wild-type and <i>Nrk</i>-null mouse placentas and foetuses at E18.5 are shown (left). The placental weight was measured at E18.5 in wild-type (n = 24) and <i>Nrk</i>-null (n = 25) embryos (right). Data are presented as mean plus SD. **<i>P</i><0.01 compared to wild-type. (C) PAS staining of E18.5 placental sections. Scale bars indicate 1 mm. (D) The lysates were prepared from wild-type (n = 2) and <i>Nrk</i>-null (n = 2) placentas at E18.5 and were subjected to western blot analysis. Expressions of NRK, phosphorylated AKT at Ser473, phosphorylated AKT at Thr308 and total AKT were compared between wild-type and <i>Nrk</i>-null placentas (left). Expressions of NRK, phosphorylated ERK1/2 and total ERK1/2 were compared between wild-type and <i>Nrk</i>-null placentas (right). As an internal control, β-Actin was also detected. (E) Immunostaining of E18.5 placental sections with anti-AKT pSer473 antibodies. Scale bars indicate 1 mm. Shown are cropped gels/blots. The gels/blots with indicated cropping lines are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0171503#pone.0171503.s004" target="_blank">S4 Fig</a>.</p

Disruption of <i>Nrk</i> resulted in enhanced proliferation of trophoblasts without obvious influence on their differentiation.

<p>(A) <i>Nrk</i> expression in wild-type cells was examined using RT-PCR analysis. Total RNAs were isolated from undifferentiated ESCs (day 0) and differentiated trophoblasts (day 4 and day 8) and were subjected to RT-PCR amplification. As an internal control, <i>Gapdh</i> was also amplified. RT-PCR amplification of <i>Gapdh</i> was also performed in the absence of reverse transcriptase (RT-). (B) Construct of the gene-targeting vector for <i>Nrk</i> disruption. The endogenous <i>Nrk</i> expression was disturbed by a gene-trapping cassette inserted between exon 7 and 8 of <i>Nrk</i> gene. The arrowhead indicates AsiSI recognition site for linearization of the vector. The arrows indicate PCR primers for detection of the homologous recombination events. (C) Deficiency of <i>Nrk</i> expression in <i>Nrk</i>-null cells was confirmed using RT-PCR analysis. Total RNAs were isolated from <i>Nrk</i>-null cells and were subsequently subjected to RT-PCR amplification as described in (A). (D) RT-PCR analysis of trophoblast marker gene expressions in wild-type and <i>Nrk</i>-null cells. Total RNAs were isolated from undifferentiated ESCs (day 0) and differentiated trophoblasts (day 4 and day 8) and were subsequently subjected to RT-PCR amplification as described in (A). (E) Real-time PCR analysis of transcription factor expressions in wild-type and <i>Nrk</i>-null cells. Undifferentiated ESCs and differentiated trophoblasts (day 8) were analyzed (n = 3). The examined gene expressions were normalized to the amount of <i>Gapdh</i> control. Each gene expression in undifferentiated wild-type ESCs is indicated as 1. Data are presented as mean plus SD. n.s. = no significant difference. (F) Phase-contrast photomicrographs of undifferentiated ESCs (left) and differentiated trophoblasts at day 8 (right) with or without <i>Nrk</i>. Scale bars indicate 200 μm. (G) Proliferation of undifferentiated ESCs and differentiated ESCs into trophoblasts. Undifferentiated ESCs were seeded on a type I collagen-coated 24-well plate at 10⁶ cells/well in a LIF-supplemented ESF7 medium (left) in which undifferentiated state of ESCs was maintained. Undifferentiated ESCs were seeded on a laminin-coated 24-well plate at 10⁶ cells/well in an rhBMP4-supplemented ESF5 medium (right) in which ESCs were induced to differentiate into trophoblasts. The cells were counted every 24 h (n = 6). The value of day1 is indicated as 1. Data are presented as mean plus SD. **<i>P</i><0.01 compared to wild-type. n.s. = no significant difference. Shown are cropped gels. The gels with indicated cropping lines are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0171503#pone.0171503.s001" target="_blank">S1 Fig</a>. ESC: embryonic stem cell, TB: trophoblast.</p