Evaluation of chemical-specific IgG antibodies in male workers from a urethane foam factory

Background: Plastic resins are complex chemicals that contain toluene diisocyanate (TDI) and/or trimellitic anhydride (TMA), which cause occupational allergies (OA), including respiratory allergies. Serum IgGs against TDI and TMA have been suggested as potential markers of the exposure status and as exploring cause of OA. Although TDI-specific IgG has been examined for suspected OA, TMA-specific IgG is not commonly evaluated in a urethane foam factory. This study therefore investigated both TDI- and TMA-specific IgGs in suspected OA patients and to evaluate the usefulness of the measurement of multiple chemical-specific IgG measurement for practical monitoring. Methods: Blood samples were collected from two male workers who developed respiratory allergies supposedly caused by occupational exposure to TDI and/or TMA for the presence of TDI- and TMA-specific IgGs. In addition, blood samples from 75 male workers from a urethane foam factory, along with 87 male control subjects, were collected in 2014 and tested for the same IgGs in 2014. The presence and levels of TDI- and TMA-specific serum IgGs were measured using dot blot assays. Results: We found that controls had mean concentrations of TDI- and TMA-specific IgGs of 0.98 and 2.10 μg/mL, respectively. In the two workers with respiratory allergies, the TDI-specific IgG concentrations were 15.6 and 9.51 μg/ mL, and TMA-specific IgG concentrations were 4.56 and 14.4 μg/mL, which are clearly higher than those in controls. Mean concentrations of TDI- and TMA-specific IgGs in the factory workers were 1.89 and 2.41 μg/mL, respectively, and are significantly higher than those of the controls (P < 0.001 and P < 0.026 for TDI- and TMA-specific IgGs, respectively). Conclusion: The workers suspected of OA showed an evidently high level of TDI- and TMA-specific IgG, and these levels in workers at the urethane foam factory were also significantly higher than those in controls. In conclusion, the measurement of TDI- and TMA-specific IgG among workers using plastic resins is helpful to monitor their exposure status.This study was funded by an Industrial Disease Clinical Research Grant (grant number 14040101-02 to M.T.) and the JSPS KAKENHI (grant numbers 22790546 and 25860472 to M.T.)

Pleiotropic Effects of Linagliptin Monotherapy on Levels of Nitric Oxide, Nitric Oxide Synthase, and Superoxide Dismutase in Hemodialysis Patients with Diabetes

Linagliptin is an anti-diabetic drug and the only bile-excreted dipeptidyl peptidase-4 inhibitor. Malnutrition-inflammation-atherosclerosis syndrome is an important prognostic factor for hemodialysis patients, and we previously reported anti-inflammatory effects of linagliptin in hemodialysis patients with diabetes. Inflammation can accelerate oxidative stress, vasoconstriction, and platelet aggregation. However, few studies have investigated the pleiotropic effects of linagliptin treatment on inflammation in hemodialysis patients. In this study, we have extended our previous investigations of these effects in a longer and more thorough follow-up of hemodialysis patients with diabetes. We examined 20 hemodialysis patients with diabetes who were not receiving oral diabetes drugs or insulin therapy and who exhibited inadequate glycemic control (glycated albumin levels＞20%). Linagliptin (5mg) was administered daily, and we evaluated the patients’ superoxide dismutase, 8-hydroxydeoxyguanosine, nitric oxide, nitric oxide synthase, and asymmetric dimethylarginine levels in serum at baseline and after 1, 3, and 6 months of treatment. After 6 months of treatment, superoxide dismutase levels had significantly decreased from 8.8±0.5U/ml to 7.0±0.5U/ml. Nitric oxide synthase levels were significantly increased at 3 and 6 months (maximum, 94.2±13.2µg/ml; baseline, 31.6±5.5µg/ml). After 3 months of treatment, nitric oxide levels had significantly increased from 64.5±6.6µmol/l to 104±15.4µmol/l, and remained significantly elevated at 6 months. Asymmetric dimethylarginine and 8-hydroxydeoxyguanosine levels did not change during the 6-month treatment course, and no patients exhibited hypoglycemia or other significant adverse effects. Linagliptin treatment significantly changed various markers of inflammation relevant to the atherosclerosis in malnutrition-inflammation-atherosclerosis syndrome. Therefore, linagliptin monotherapy has pleiotropic effects on inflammation in hemodialysis patients with diabetes, and may improve their prognosis

Construction of an All-in-one Double-conditional shRNA Expression Vector

Gene silencing by RNA interference （RNAi） is widely used for assessing gene function. An important advance in the RNAi field was the discovery that plasmid-based RNAi can substitute for synthetic small interfering RNA in vitro and in vivo. However, constitutive and ubiquitous knockdown of gene expression by RNAi in mice can limit the scope of experiments because this process can lead to embryonic lethality, or result in compensatory overexpression of other genes such that no phenotypic abnormalities occur. Either way, analyses of the physiological roles of the gene of interest in adult mice are not possible. To overcome these limitations, we previously constructed a double-conditional short-hairpin RNA （shRNA） expression vector that can regulate shRNA expression in a spatio-temporal manner with a tetracycline-inducible floxed stuffer sequence selectively excised by application of Cre recombinase. In this study, we aimed to modify this vector to create an all-in-one vector that produces double-conditional transgenic mice through a single round of gene transfer to fertilized eggs. We added a coding region for nuclear localizing Cre （NCre） recombinase with a multi-cloning site for a cell-specific promoter into the double-conditional short-hairpin RNA （shRNA） expression vector that we previously constructed. Using Escherichia coli, we confirmed successful construction of the vector. First, we confirmed isopropyl-β-D-thiogalactopyranoside-induced expression of NCre recombinase through the lac operon as a specific promoter by western blotting. Second, we confirmed functional recombination of the floxed sequence of loxP-like TATA-lox by analysing restriction enzyme-digested fragments. This all-in-one double-conditional shRNA expression vector will be useful for reversible in vitro and in vivo knockdown of target gene expression, in target cells via promoter-specific expression of NCre, and at specific times by tetracycline application

Inter-rater reliability of the AFTD-pitting test among elderly patients in a long-term medical facility

BACKGROUND and AIM: The pitting test has been reported in various methods as a standard for evaluating chronic oedema, but a unified method has not been determined. This makes it difficult to accurately specify the prevalence of oedema. The present study aimed to evaluate inter-rater reliability of the AFTD-pitting test, which included 4 factors: Anatomical locations of oedema; Force required to pit; the amount of Time; and the Definition of oedema. The present study is the first stage of an international epidemiological study of chronic oedema.METHODS: This cross-sectional observational study was performed at a long-term care hospital in Ishikawa Prefecture, Japan. The inter-rater reliability of the pitting test for evaluating oedema using the AFTD-pitting test was tested for 34 locations on the body, with 10 seconds of pitting with a similar force to that of the reference rater and assessed using the modified Fukazawa method. One reference rater and four raters evaluated oedema in five patients. Then, the agreement rate and Cohen-s kappa coefficient were calculated.RESULTS: All protocols were completed by four raters for five bedridden patients. Agreement among the four raters was high, at >0.85, and the kappa coefficient showed almost perfect, moderate, and fair agreement for one (0.81), four (0.51-0.60) rater, respectively.CONCLUSION: The inter-rater reliability of four nurses who applied the AFTD-pitting test was high, and the kappa coefficient showed at least fair agreement. Therefore, the AFTD-pitting test is a useful method to assess whole-body chronic oedema

A novel ex vivo lung cancer model based on bioengineered rat lungs

Introduction: Two-dimensional cell cultures have contributed substantially to lung cancer research, but 3D cultures are gaining attention as a new, more efficient, and effective research model. A model reproducing the 3D characteristics and tumor microenvironment of the lungs in vivo, including the co-existence of healthy alveolar cells with lung cancer cells, is ideal. Here, we describe the creation of a successful ex vivo lung cancer model based on bioengineered lungs formed by decellularization and recellularization.Methods: Human cancer cells were directly implanted into a bioengineered rat lung, which was created with a decellularized rat lung scaffold reseeded with epithelial cells, endothelial cells and adipose-derived stem cells. Four human lung cancer cell lines (A549, PC-9, H1299, and PC-6) were applied to demonstrate forming cancer nodules on recellularized lungs and histopathological assessment were made among these models. MUC-1 expression analysis, RNA-seq analysis and drug response test were performed to demonstrate the superiority of this cancer model.Results: The morphology and MUC-1 expression of the model were like those of lung cancer in vivo. RNA sequencing revealed an elevated expression of genes related to epithelial-mesenchymal transition, hypoxia, and TNF-α signaling via NF-κB; but suppression of cell cycle-related genes including E2F. Drug response assays showed that gefitinib suppressed PC-9 cell proliferation equally well in the 3D lung cancer model as in 2D culture dishes, albeit over a smaller volume of cells, suggesting that fluctuations in gefitinib resistance genes such as JUN may affect drug sensitivity.Conclusions: A novel ex vivo lung cancer model was closely reproduced the 3D structure and microenvironment of the actual lungs, highlighting its possible use as a platform for lung cancer research and pathophysiological studies

Apoptosis-induced Proliferation in UV-Irradiated Human Conjunctival Epithelial Cells

A pterygium is a benign growth that develops on the conjunctiva and, in some cases, extends to the cornea and interferes with vision. Excessive exposure to ultraviolet (UV) light is one of the causes of pterygium development. We previously reported that UV-induced apoptosis is led by production of reactive oxygen species (ROS) that activate p38 mitogen-activated protein kinase (MAPK) in human conjunctival epithelial (HCE) cells. Also, ROS-dependent induction of interleukin-11 (IL-11) has been reported to upregulate MAPK pathways, which results in compensatory proliferation. In this study, we examined the effect of UV exposure on HCE cells, in terms of change in apoptosis, ROS generation, phosphorylation of c-Jun N-terminal kinase (JNK), levels of IL-11 (a key cytokine in tissue repair and compensatory proliferation), production of activator protein 1 (AP-1), and expression of c-myc, c-fos and c-jun (which provides evidence of healthy cell proliferation). Apoptosis in HCE cells was induced by UV light irradiation (312nm, 4.94mW/cm2). Apoptosis was measured using the Muse Annexin V and Dead Cell Assay Kit. ROS generation was measured by using 5-(and 6-) chloromethyl-2\u277\u27-dichlorodihydrofluorescein diacetate, acetyl ester. JNK phosphorylation, IL-11 levels and AP-1 production were measured by enzyme-linked immunosorbent assays (ELISAs). Imnunocytochemical staining was used to measure c-myc, c-fos and c-jun expression. UV irradiation increased ROS generation, phosphorylation of JNK, and apoptotic cell count. IL-11 levels and AP-1 production were significantly increased by UV irradiation. The irradiated cells had increased expression of c-myc, c-fos and c-jun, and treatment of the cells with IL-11 significantly increased expression of c-myc, c-fos and c-jun. These results suggest that the release of IL-11 from UV-induced apoptotic HCE cells and surrounding healthy cells could promote proliferation to maintain homeostasis

Increased In Vitro Intercellular Barrier Function of Lung Epithelial Cells Using Adipose-Derived Mesenchymal Stem/Stromal Cells

With the emergence of coronavirus disease-2019, researchers have gained interest in the therapeutic efficacy of mesenchymal stem/stromal cells (MSCs) in acute respiratory distress syndrome; however, the mechanisms of the therapeutic effects of MSCs are unclear. We have previously reported that adipose-derived MSCs (AD-MSCs) strengthen the barrier function of the pulmonary vessels in scaffold-based bioengineered rat lungs. In this study, we evaluated whether AD-MSCs could enhance the intercellular barrier function of lung epithelial cells in vitro using a transwell coculture system. Transepithelial electrical resistance (TEER) measurements revealed that the peak TEER value was significantly higher in the AD-MSC coculture group than in the AD-MSC non-coculture group. Similarly, the permeability coefficient was significantly decreased in the AD-MSC coculture group compared to that in the AD-MSC non-coculture group. Immunostaining of insert membranes showed that zonula occuldens-1 expression was significantly high at cell junctions in the AD-MSC coculture group. Moreover, cell junction-related gene profiling showed that the expression of some claudin genes, including claudin-4, was upregulated in the AD-MSC coculture group. Taken together, these results showed that AD-MSCs enhanced the barrier function between lung epithelial cells, suggesting that both direct adhesion and indirect paracrine effects strengthened the barrier function of lung alveolar epithelium in vitro

Study on Incident Causal Factors About The Oral Medicine By Nurses: Analysis From The Administration Prosses

The aim of this study was to elucidate the differences in the causes of incidents at each stage of drug administration between nurses with different numbers of years of experience and current nursing unit tenure. We evaluated 510 reports involving medications taken after meals where medication charts were utilized. These reports were selected from 1,173 incident reports involving drug administration by hospital ward nurses in advanced treatment hospitals with 640 bed capacities in the northeastern area of Japan between fiscal 2012 and 2014. Approximately 40% of these were related to the drug preparation stage. There was no difference in the frequency of incidents between nurses with less than 2 years and those with 11 or more years of experience. As per the results of our correspondence analysis of the relation between the nursing unit tenure and the causal factors for each stage, “insufficient checking” at all stages was the most common factor. At the drug preparation and distribution stages, 11 or more years of experience were related to physical conditions. At all stages, nursing unit tenure of less than 2 years was related to psychological conditions. The findings indicated that nurse training and organized approach are required to minimize human factors involved in medical incidents