Involvement of autophagy in the maintenance of rat intervertebral disc homeostasis: an in-vitro and in-vivo RNA interference study of Atg5.

OBJECTIVE: In the largest avascular low-nutrient intervertebral disc, resident cells would utilize autophagy, a stress-response survival mechanism by self-digestion and recycling wastes. Our goal was to elucidate the involvement of autophagy in disc homeostasis through RNA interference of autophagy-related gene 5 (Atg5). DESIGN: In vitro, small interfering RNAs (siRNAs) targeting autophagy-essential Atg5 were transfected into rat disc cells. Cell viability with levels of autophagy including Atg5 expression, apoptosis, and senescence was assessed under serum starvation and/or pro-inflammatory interleukin-1 beta (IL-1ß) stimulation. In vivo, time-course autophagic flux was monitored following Alexa Fluor® 555-labeled Atg5-siRNA injection into rat tail discs. Furthermore, 24-h temporary static compression-induced disruption of Atg5 siRNA-injected discs was observed by radiography, histomorphology, and immunofluorescence. RESULTS: In disc cells, three different Atg5 siRNAs consistently suppressed autophagy with Atg5 protein knockdown (mean 44.4% [95% confidence interval: -51.7, -37.1], 51.5% [-80.5, -22.5], 62.3% [-96.6, -28.2]). Then, Atg5 knockdown reduced cell viability through apoptosis and senescence not in serum-supplemented medium (93.6% [-0.8, 21.4]) but in serum-deprived medium (66.4% [-29.8, -8.6]) further with IL-1ß (44.5% [-36.9, -23.5]). In disc tissues, immunofluorescence detected intradiscal signals for the labeled siRNA even at 56-d post-injection. Immunoblotting found 56-d autophagy suppression with prolonged Atg5 knockdown (33.2% [-52.8, -5.3]). With compression, Atg5 siRNA-injected discs presented radiographic height loss ([-43.9, -0.8]), histological damage ([-5.5, -0.2]), and immunofluorescent apoptosis ([2.2, 22.2]) and senescence ([4.1, 19.9]) induction compared to control siRNA-injected discs at 56 d. CONCLUSIONS: This loss-of-function study suggests Atg5-dependent autophagy-mediated anti-apoptosis and anti-senescence. Autophagy could be a molecular therapeutic target for degenerative disc disease.

Measurements of Strong-Interaction Effects in Kaonic-Helium Isotopes at Sub-eV Precision with X-Ray Microcalorimeters.

We have measured the 3d→2p transition x rays of kaonic ^{3}He and ^{4}He atoms using superconducting transition-edge-sensor microcalorimeters with an energy resolution better than 6 eV (FWHM). We determined the energies to be 6224.5±0.4(stat)±0.2(syst) eV and 6463.7±0.3(stat)±0.1(syst) eV, and widths to be 2.5±1.0(stat)±0.4(syst) eV and 1.0±0.6(stat)±0.3(stat) eV, for kaonic ^{3}He and ^{4}He, respectively. These values are nearly 10 times more precise than in previous measurements. Our results exclude the large strong-interaction shifts and widths that are suggested by a coupled-channel approach and agree with calculations based on optical-potential models.

Search for η

We measured missing mass spectrum of the ^{12}C(γ,p) reaction for the first time in coincidence with potential decay products from η^{'} bound nuclei. We tagged an (η+p) pair associated with the η^{'}N→ηN process in a nucleus. After applying kinematical selections to reduce backgrounds, no signal events were observed in the bound-state region. An upper limit of the signal cross section in the opening angle cosθ_{lab}^{ηp}<-0.9 was obtained to be 2.2 nb/sr at the 90% confidence level. It is compared with theoretical cross sections, whose normalization ambiguity is suppressed by measuring a quasifree η^{'} production rate. Our results indicate a small branching fraction of the η^{'}N→ηN process and/or a shallow η^{'}-nucleus potential.

IL4Rα and ADAM33 as genetic markers in asthma exacerbations and type-2 inflammatory endotype.

BACKGROUND: Genetic markers of susceptibility to asthma exacerbations in adults remain unclear. OBJECTIVE: To identify genetic markers of asthma exacerbations, particularly in patients with type-2 inflammatory endotype. METHODS: In this observational study of patients enrolled in the Kinki Hokuriku Airway disease Conference multicenter study, frequency of exacerbations requiring systemic corticosteroids during 2 years after enrolment and associated risk factors was determined. For genetic marker analysis, interleukin-4 receptor α (IL4RA) rs8832 and a disintegrin and metalloprotease 33 (ADAM33) S_2 (rs528557), T_1 (rs2280091), T_2 (rs2280090), and V_4 (rs2787094) variants were included. Elevated serum periostin levels at enrolment (≥95 ng/mL, defined as type-2 inflammatory endotype) were considered in the analysis. RESULTS: Among 217 patients who were successfully followed up for 2 years after enrolment, 60 patients showed at least one asthma exacerbation during the 2 years. Airflow limitation (%FEV1 <80%) and recent exacerbations but not genetic variants were identified as risk markers of exacerbations. A total of 27 patients showed type-2 inflammatory endotype (serum periostin ≥95 ng/mL at enrolment) and subsequent exacerbations; risk factors in these patients were airflow limitation (odds ratio, 6.51; 95% confidence interval (CI): 2.37-18.6; P=.0003), GG genotype of IL4RA rs8832 (odds ratio, 4.01; 95% CI: 1.47-11.0; P=.007), and A allele of ADAM33 T_2 (odds ratio, 2.81; 95% CI: 1.05-7.67; P=.04) by multivariate analysis. In addition, GG genotype of IL4RA rs8832 was associated with type-2 endotype, whereas A allele of ADAM33 T_2 was associated with mixed type of eosinophilic/type-2 and neutrophilic inflammations. CONCLUSIONS AND CLINICAL RELEVANCE: IL4RA and ADAM33 variants may be risk markers of asthma exacerbations in type-2 inflammatory endotype. Precise endotyping may facilitate the identification of genetic risk markers of asthma exacerbations.

Effect of sodium-glucose co-transporter-2 inhibitors on impaired ventricular repolarization in people with Type 2 diabetes.

AIMS: To test the hypothesis that treatment with a sodium-glucose co-transporter-2 inhibitor would reverse ventricular repolarization heterogeneity, a predictor of cardiovascular mortality, in people with Type 2 diabetes. METHODS: We retrospectively analysed changes in indices of ventricular repolarization before and after treatment with a sodium-glucose co-transporter-2 inhibitor in 46 people with Type 2 diabetes. RESULTS: Sodium-glucose co-transporter-2 inhibitor treatment reduced HbA1c concentration [62±13 mmol/mol (7.7±1.2%) vs 59±16 mmol/mol (7.5±1.4%)], body weight (77.8±13.9 vs 74.7±12.5 kg) and systolic blood pressure (133±18 vs 126±12 mmHg) in the study participants. Heart rate and QTc interval were not changed by sodium-glucose co-transporter-2 inhibitor treatment, but QTc dispersion was significantly reduced (median, 48.8 vs 44.2 ms). Sodium-glucose co-transporter-2 inhibitor treatment reversed QTc dispersion more in participants who had larger QTc dispersion before the treatment. Changes in systolic blood pressure (Spearman's ρ= 0.319; P=0.031), but not in HbA1c concentration, were correlated with changes in QTc dispersion after sodium-glucose co-transporter-2 inhibitor treatment. CONCLUSIONS: The findings suggest that sodium-glucose co-transporter-2 inhibitor treatment reverses ventricular repolarization heterogeneity in people with Type 2 diabetes, independently of its effect on glycaemic control. The favourable effect on ventricular repolarization heterogeneity could be the mechanism by which empaglifozin reduced cardiovascular events in a recent study.

IQ is an independent predictor of glycated haemoglobin level in young and middle-aged adults with intellectual disability.

AIMS: Here we examined whether intellectual disability is independently associated with hyperglycaemia. METHODS: We recruited 233 consecutive young and middle-aged adults with intellectual disability. After exclusion of subjects on medication for metabolic diseases or with severe intellectual disability (IQ < 35), 121 subjects were divided by IQ into a group with moderate intellectual disability (35 ≤ IQ ≤ 50), a mild intellectual disability group (51 ≤ IQ ≤ 70) and a borderline group (IQ > 70). RESULTS: HbA1c level was higher in subjects with moderate intellectual disability (42 ± 9 mmol/mol; 6.0 ± 0.8%) than those in the borderline group (36 ± 4 mmol/mol; 5.5 ± 0.3%) and mild intellectual disability group (37 ± 5 mmol/mol; 5.5 ± 0.5%) groups. HbA1c level was correlated with age, BMI, blood pressure, serum triglycerides and IQ in simple linear regression analysis. Multiple regression analysis indicated that IQ, age, BMI and diastolic blood pressure were independent explanatory factors of HbA1c level. CONCLUSIONS: An unfavourable effect of intellectual disability on lifestyle and untoward effect of hyperglycaemia on cognitive function may underlie the association of low IQ with hyperglycaemia.

GLCCI1 variant accelerates pulmonary function decline in patients with asthma receiving inhaled corticosteroids.

BACKGROUND: In steroid-naive patients with asthma, several gene variants are associated with a short-term response to inhaled corticosteroid (ICS) treatment; this has mostly been observed in Caucasians. However, not many studies have been conducted for other ethnicities. Here, we aimed to determine the relationship between the annual decline in forced expiratory flow volume in one second (FEV1 ) and the variant of the glucocorticoid-induced transcript 1 gene (GLCCI1) in Japanese patients with asthma receiving long-term ICS treatment, taking into account the effect of high serum periostin levels, a known association factor of pulmonary function decline and a marker of refractory eosinophilic/Th2 inflammation. METHODS: In this study, 224 patients with asthma receiving ICS treatment for at least 4 years were enrolled. The effects of single-nucleotide polymorphisms (SNPs) in GLCCI1, stress-induced phosphoprotein 1 (STIP1), and T gene on the decline in FEV1 of 30 ml/year or greater were determined. RESULTS: Besides the known contributing factors, that is, the most intensive treatment step, ex-smoking, and high serum periostin levels (≥95 ng/ml), the GG genotype of GLCCI1 rs37973, and not other SNPs, was independently associated with a decline in FEV1 of 30 ml/year or greater. When patients were stratified according to their serum periostin levels, the GG genotype of rs37973 was significantly associated with blood eosinophilia (≥250/µl) in the high serum periostin group. CONCLUSIONS: A GLCCI1 variant is a risk factor of pulmonary function decline in Japanese patients with asthma receiving long-term ICS treatment. Thus, GLCCI1 may be associated with response to ICS across ethnicities.

IL-13-producing BLT1-positive CD8 cells are increased in asthma and are associated with airway obstruction.

BACKGROUND: The role of CD8 T lymphocytes in the pathogenesis of asthma is not well understood. We investigated whether a subset of IL-13-producing BLT1-positive CD8 T lymphocytes are present in asthmatic airways and are associated with impaired lung function. METHODS: Bronchoalveolar lavage (BAL) cells were obtained from asthmatic (n = 39) and healthy control (n = 28) subjects. Cells were stimulated with phorbol ester and ionomycin in the presence of brefeldin A and stained for CD8, BLT1, and intracellular IL-13. The frequency of IL-13-producing BLT1-positive CD8 T lymphocytes was compared between the two groups and related to lung function, serum IgE levels, and reticular basement membrane (RBM) thickness. RESULTS: A subset of CD8 T lymphocytes expressing BLT1 and producing IL-13 were detected in the airways of all asthmatic subjects. The frequency of this subset among recovered lymphocytes was significantly higher in the airways of asthmatic subjects compared with controls (mean ± SEM: 16.2 ± 1.4 vs 5.3 ± 0.5, respectively, P < 0.001) and correlated positively with serum IgE levels and RBM thickness. More importantly, the frequency of CD8 T lymphocytes co-expressing BLT1 and IL-13 was inversely related to FEV1 and FEF[25-75] percent predicted values (P < 0.001). CONCLUSIONS: A subset of CD8 T lymphocytes expressing BLT1 and producing IL-13 is present in the airways of asthmatics. The accumulation of these cells is associated with airway obstruction, suggesting that they may play a significant pathogenic role in bronchial asthma.

Impact of immunoreactive substances contained in apheresis platelet concentrate on postoperative respiratory function in surgical patients receiving platelet transfusion: a prospective cohort study.

OBJECTIVES: To construct an alternative policy for the donor selection of platelet concentrate (PC), a clinical study exploring the features of lung injury following PC administration is needed. BACKGROUND: Although a male-donor-only policy for plasma products appears to have efficiently reduced transfusion-related acute lung injury (TRALI), this policy may not be applied to PC because of supply shortages. METHODS AND MATERIALS: We prospectively examined pulmonary function after the transfusion of PC in informed surgical patients treated at a tertiary university hospital in Japan. The contributions of immunoreactive substances contained in the PC to respiratory function after PC transfusion was then statistically examined. RESULTS: Eighty-six patients (56 men, 30 women) were enrolled in the analysis. Fifty-four cases experienced respiratory failure (PaO2 /FiO2 <300 mmHg) after transfusion. Five cases were diagnosed as possible TRALI based on permeability pulmonary oedema, while 23 cases were diagnosed as transfusion-associated circulatory overload (TACO) based on chest radiograph findings. A multivariate logistic regression analysis identified the presence of anti-granulocyte antibody as a significant predictor of possible TRALI [P = 0.023; odds ratio (OR), 13.0; 95% confidence interval (CI), 1.4-118.3]. Meanwhile, anti-leukocyte antibody class II was identified as a significant independent predictor of TACO (P = 0.010; OR, 18.4; 95% CI, 2.0-170.1). CONCLUSIONS: Our data suggest that antibodies contained in PC may contribute to the deterioration of respiratory function after PC transfusion, although the diagnoses of TACO and TRALI may have overlapped among the patients with pulmonary distress in this cohort.

Linezolid-induced pure red cell aplasia in a patient with Staphylococcus epidermidis infection after allogeneic stem cell transplantation.

Linezolid (LZD) is the first oxazolidinone antibiotic that is effective against drug-resistant gram-positive organisms. Hematological toxicities such as thrombocytopenia, anemia, and leukocytopenia are common in LZD therapy. However, LZD-induced pure red cell aplasia (PRCA) is very rare. A 56-year-old man with myelodysplastic syndrome underwent allogeneic bone marrow transplantation from a human leukocyte antigen-matched and ABO blood type-matched unrelated male donor. He had bacteremia caused by Staphylococcus epidermidis after engraftment of neutrophils and red blood cells. We first administered vancomycin, but then changed to intravenous LZD because of kidney damage. Two weeks after LZD therapy, the patient's hemoglobin and reticulocyte levels were 6.8 g/dL and 0.3%, respectively. Bone marrow examination revealed red blood cell aplasia (myeloid/erythroid ratio was 402). The patient showed rapid recovery of normal erythropoiesis within 2 weeks of LZD cessation. It is important to be aware of the hematological effects associated with LZD in the setting of stem cell transplantation,particularly for those with pre-existing myelosuppression, renal insufficiency, and those receiving concomitant drugs that produce bone marrow suppression. We advocate that a reticulocyte count be performed periodically for detecting bone marrow suppression, including PRCA, during LZD therapy.

Outcomes of endoscopic and surgical resection for a second primary cancer in the residual cervical esophagus after thoracic esophagectomy.

Patients who have received subtotal esophagectomy for thoracic esophageal cancer must be closely monitored for second primary malignancies. The purpose of this study is to review and assess patients who developed a second primary esophageal cancer in the residual cervical esophagus. Between 1996 and 2010, 10 patients were diagnosed in our hospital with esophageal squamous cell cancer in the residual cervical esophagus after undergoing thoracic esophagectomy and were treated with endoscopic or surgical resection. Data from these patients were reviewed retrospectively. Seven of the 10 patients (70%) had multiple primary carcinoma lesions at the time of their esophagectomy. A second primary cancer in the residual cervical esophagus was detected in eight patients during follow-up endoscopic examinations while the patients were still asymptomatic. Seven of the patients underwent endoscopic resection for a superficial cancer. None of those patients experienced any complications, and all are currently alive and cancer-free. The remaining three patients underwent resection of the cervical esophagus with regional lymph node dissection. Two of those patients experienced severe complications; one subsequently died (hospital death) from pneumonia, 12 months after surgery, while the other died from recurrence of his cancer. The third patient is alive and cancer-free. Early detection of a second primary malignancy in the residual cervical esophagus followed by endoscopic resection is the best treatment strategy for patients who previously received subtotal esophagectomy for thoracic esophageal cancer. Surgical resection puts patients at high risk of mortality or morbidity.

Circulating KL-6/MUC1 as an independent predictor for disseminated intravascular coagulation in acute respiratory distress syndrome.

OBJECTIVES: Acute respiratory distress syndrome (ARDS) patients show high levels of circulating mucin including KL-6/MUC1 (soluble MUC1 mucin). Because cancer mucin can bind vascular endothelial cells and platelets via selectins, mucin-selectin interactions are reported to trigger platelet aggregation and intravascular coagulation. Therefore, we hypothesized that KL-6/MUC1 is involved in the pathogenesis of disseminated intravascular coagulation (DIC) in ARDS. The aim of the current study is to evaluate the association between circulating KL-6/MUC1 and DIC in ARDS patients. DESIGN: Observational study with structured follow-up. SETTING: Intensive care unit in Hiroshima University Hospital. SUBJECTS: Fifty-six newly diagnosed patients with ARDS. INTERVENTIONS: Circulating levels of KL-6/MUC1 were measured during diagnosis and serially measured during the clinical course along with indices of respiratory failure, inflammation, coagulation and fibrinolysis and multiple organ dysfunction. RESULTS: Acute respiratory distress syndrome patients complicated with DIC showed significantly higher levels of serum KL-6/MUC1 than patients without DIC during the clinical course. Amongst the parameters analysed at diagnosis of ARDS, KL-6/MUC1 was an independent predictor for DIC complication. The baseline level of circulating KL-6/MUC1 at diagnosis of ARDS was significantly correlated with an increased DIC score following ARDS diagnosis. Using an optimum cutoff level of KL-6/MUC1 obtained by a receiver operating characteristic curve, the sensitivity and specificity for predicting future DIC development in ARDS patients were 88.9% and 55.3%, respectively. CONCLUSIONS: These results suggest that KL-6/MUC1 is associated with DIC development in ARDS patients. Elevated levels of KL-6/MUC1 at diagnosis could be a predictor of DIC complication in ARDS.

Overexpression of Rab3D enhances regulated amylase secretion from pancreatic acini of transgenic mice.

Rab3D, a member of the ras-related GTP-binding protein Rab family, is localized to secretory granules of various exocrine tissues such as acinar cells of the pancreas, chief cells of the stomach, and parotid and lacrimal secretory cells. To elucidate the function of Rab3D in exocytosis, we have generated transgenic mice that over-express Rab3D specifically in pancreatic acinar cells. Hemagglutinin-tagged Rab3D was localized to zymogen granules by immunohistochemistry, and was shown to be present on zymogen granule membranes by Western blotting; both results are similar to previous studies of endogenous Rab3D. Secretion measurements in isolated acinar preparations showed that overexpression of Rab3D enhanced amylase release. Amylase secretion from intact acini of transgenic mice 5 min after 10 pM cholecystokinin octapeptide (CCK) stimulation was enhanced by 160% of control. In streptolysin-O-permeabilized acini of transgenic mice, amylase secretion induced by 100 microM GTP-gamma-S was enhanced by 150%, and 10 microM Ca2+-stimulated amylase secretion was augmented by 206% of that of the control. To further elucidate Rab3D involvement in stimulus-secretion coupling, we examined the effect of CCK on the rate of GTP binding to Rab3D. Stimulation of permeabilized acini with 10 pM CCK increased the incorporation of radiolabeled GTP into HA-tagged Rab3D. These results indicate that overexpression of Rab3D enhances secretagogue-stimulated amylase secretion through both calcium and GTP pathways. We conclude that Rab3D protein on zymogen granules plays a stimulatory role in regulated amylase secretion from pancreatic acini.

Betacellulin and activin A coordinately convert amylase-secreting pancreatic AR42J cells into insulin-secreting cells.

Rat pancreatic AR42J cells possess exocrine and neuroendocrine properties. Activin A induces morphological changes and converts them into neuron-like cells. In activin-treated cells, mRNA for pancreatic polypeptide (PP) but not that for either insulin or glucagon was detected by reverse transcription-PCR. About 25% of the cells were stained by anti-PP antibody. When AR42J cells were incubated with betacellulin, a small portion of the cells were stained positively with antiinsulin and anti-PP antibodies. The effect of betacellulin was dose dependent, being maximal at 2 nM. Approximately 4% of the cells became insulin positive at this concentration, and mRNAs for insulin and PP were detected. When AR42J cells were incubated with a combination of betacellulin and activin A, approximately 10% of the cells became insulin positive. Morphologically, the insulin-positive cells were composed of two types of cells: neuron-like and round-shaped cells. Immunoreactive PP was found in the latter type of cells. The mRNAs for insulin, PP, glucose transporter 2, and glucokinase, but not glucagon, were detected. Depolarizing concentration of potassium, tolbutamide, carbachol, and glucagon-like peptide-1 stimulated the release of immunoreactive insulin. These results indicate that betacellulin and activin A convert amylase-secreting AR42J cells into cells secreting insulin. AR42J cells provide a model system to study the formation of pancreatic endocrine cells.

Conversion of amylase-secreting rat pancreatic AR42J cells to neuronlike cells by activin A.

When AR42J cells, an amylase-secreting pancreatic exocrine cell line, were treated with activin A, cells extended neuritelike processes, and, concomitantly, amylase-containing vesicles disappeared. Immunofluorescence and immunoelectron microscopy revealed that these processes had neurite-specific cytoskeletal architectures: neurofilaments and microtubule bundles with cross-bridges of microtubule-associated protein 2. In addition to such morphological changes, activin-treated cells exhibited a marked increase in cytoplasmic free calcium concentration in response to depolarizing concentration of potassium. Moreover, activin-treated AR42J cells expressed mRNA for alpha 1 subunit of the neuroendocrine/beta cell-type voltage-dependent calcium channel. In naive AR42J cells, a sulfonylurea compound, tolbutamide, did not affect free calcium concentration, while it induced a marked elevation of free calcium in activin-treated cells. Single channel recording of the membrane patch revealed the existence of ATP-sensitive potassium channel in activin-treated cells. These results indicate that activin A converts amylase-secreting AR42J cells to neuronlike cells. Given that pancreatic endocrine cells possess neuronlike properties and express ATP-sensitive potassium channel as well as neuroendocrine/beta cell-type voltage-dependent calcium channel, activin treatment of AR42J cells may provide an in vitro model system to study the conversion of pancreatic exocrine cells to endocrine cells in islets.

Effects of the short chain sugar acid 2-buten-4-olide on the hypothalamo-pituitary-adrenal axis in normal and adjuvant-induced arthritic rats.

The effects of intraperitoneal (i.p.) administration of 2-buten-4-olide (2-B4O), an endogenous sugar acid, on the hypothalamo-adenohypophysial system were examined in Lewis rats that were normal and in adjuvant-induced arthritic (AA) rats. In comparison with vehicle-treated rats, the plasma corticosterone and c-fos mRNA levels in the paraventricular nucleus (PVN) of normal rats increased significantly after i.p. administration of 2-B4O. Dual immunostaining revealed that almost all corticotrophin-releasing factor (CRF)-immunopositive neurones in the parvocellular division of the PVN exhibited Fos-like immunoreactivity (LI) 120 min after i.p. administration of 2-B4O (100 mg/kg). In the AA rats, repeated i.p. administration of 2-B4O (100 mg/kg) after immunisation significantly suppressed the expression of clinical symptoms and significantly increased plasma concentrations of corticosterone. Further, repeated i.p. administration of 2-B4O significantly increased CRF mRNA levels in the PVN and pro-opiomelanocortin mRNA levels in the anterior pituitary; however, they did not change arginine vasopressin mRNA levels in the parvocellular division of the PVN. These results suggest that i.p. administration of 2-B4O activates the hypothalamo-pituitary-adrenal (HPA) axis via the activation of CRF neurones in the PVN, and the activation of the HPA axis by i.p. administration of 2-B4O may be associated with the inhibition of AA in rats.

Surfactant protein D inhibits activation of non-small cell lung cancer-associated mutant EGFR and affects clinical outcomes of patients.

Tyrosine kinase inhibitor (TKI)-sensitive and TKI-resistant mutations of epidermal growth factor receptor (EGFR) are associated with lung adenocarcinoma. EGFR mutants were previously shown to exhibit ligand-independent activation. We have previously demonstrated that pulmonary surfactant protein D (SP-D, SFTPD) suppressed wild-type EGFR signaling by blocking ligand binding to EGFR. We herein demonstrate that SFTPD downregulates ligand-independent signaling in cells harboring EGFR mutations such as TKI-sensitive exon 19 deletion (Ex19del) and L858R mutation as well as TKI-resistant T790M mutation, subsequently suppressing cellular growth and motility. Lectin blotting and ligand blotting in lung cancer cell lines suggested that EGFR mutants express oligomannose-type N-glycans and interact with SFTPD directly. Cross-linking assay indicated that SFTPD inhibits ligand-independent dimerization of EGFR mutants. We also demonstrated that SFTPD reduced dimerization-independent phosphorylation of Ex19del and T790M EGFR mutants using point mutations that disrupted the asymmetric dimer interface. It was confirmed that SFTPD augmented the viability-suppressing effects of EGFR-TKIs. Furthermore, retrospective analysis of 121 patients with lung adenocarcinoma to examine associations between serum SFTPD levels and clinical outcome indicated that in TKI-treated patients with lung cancer harboring EGFR mutations, including Ex19del or L858R, high serum SFTPD levels correlated with a lower number of distant metastases and prolonged overall survival and progression-free survival. These findings suggest that SFTPD downregulates both TKI-sensitive and -resistant EGFR mutant signaling, and SFTPD level is correlated with clinical outcome. These findings illustrate the use of serum SFTPD level as a potential marker to estimate the efficacy of EGFR-TKIs.

Prevalence of, and risk factors for, carriage of carbapenem-resistant Enterobacteriaceae among hospitalized patients in Japan.

BACKGROUND: The prevalence of carbapenem-resistant Enterobacteriaceae (CRE) has been reported to be lower in Japan than in many other countries. However, extensive surveillance for CRE carriage has not been performed in Japan. AIM: To investigate the prevalence of CRE carriage in Japan among convalescent patients considered to be at high risk of being CRE carriers using an improved selective culture medium. METHODS: A cross-sectional survey was conducted in 22 acute care hospitals (ACHs) and 21 long-term care hospitals (LTCHs) in northern Osaka from December 2015 to January 2016. Patients who used incontinence aids, an enteral feeding tube or a urinary catheter were enrolled. Faecal specimens were examined using the newly developed M-ECC for imipenemase (IMP)-producing CRE, which is the most prevalent form of CRE in Japan. The positive isolates were analysed by polymerase chain reaction and sequencing. Risk factors associated with carriage were analysed by logistic regression. FINDINGS: Among 1507 patients, 184 (12.2%) carried CRE. The percentage of positive patients was significantly higher in LTCHs (14.9%) than in ACHs (3.6%) (P<0.001). Risk factors for CRE carriage were longer hospital stay [odds ratio (OR) 2.59; 95% confidence interval (CI) 1.87-3.60], enteral feeding (OR 3.03, 95% CI 2.08-4.42) and antibiotic exposure (OR 2.00, 95% CI 1.40-2.87). Among the 233 CRE isolates identified, 223 were IMP producers; the remaining isolates did not produce carbapenemase. CONCLUSIONS: This is the first Japanese report to demonstrate the significant spread of CRE in both ACHs and LTCHs using an improved selective medium. A coordinated regional approach may help to prevent further spread.