IL23R on myeloid cells is involved in murine pulmonary granuloma formation.

PURPOSE OF THE STUDY: The involvement of the IL-23/IL23R pathway is well known in the disease pathogenesis of sarcoidosis and other inflammatory diseases. To date, the pathogenic mechanism of IL-23 is most notably described on CD4+ Th17 lymphocytes. However, the function of the IL23R on myeloid cells in sarcoidosis is poorly understood. Thus, the aim of the study is to investigate the role of the IL23R on myeloid cell in pulmonary granuloma formation. Methods: We generated IL23RLysMCre mice lacking the IL23R gene in myeloid cells. The importance of IL23R in myeloid cells for the development of sarcoidosis was studied in a mouse model of inflammatory lung granuloma formation through embolization of PPD from Mycobacterium bovis-coated Sepharose beads into previously PPD-immunized mice. In addition the function of IL23R on myeloid cells was studied in LPS or IFNγ stimulated BMDMs and BMDCs. The mRNA and protein expression levels of relevant cytokines were analyzed by RT-PCR (TaqMan) and ELISA. The composition of immune cells in BALF was quantified by flow cytometry and alteration in granuloma sizes were observed by H&E stained lung sections. Results: Mycobacterium Ag-elicted pulmonary granulomas tend to be smaller in IL23RLysMCre mice and NF-κB dependent Th1 cytokines in the murine lungs are reduced compared to wildtype mice. In line, we observed that IL23R-deficient bone marrow-derived macrophages show a reduced production of Th1 cytokines after LPS stimulation. Conclusion: We here for the first time demonstrate a role for IL23R on myeloid cells in pulmonary inflammation and granuloma formation. Our findings provide essential insights in the pathogenesis of inflammatory lung diseases like sarcoidosis, which might be useful for the development of novel therapeutics targeting distinct immunological pathways like IL-23/IL23R.

Pseudomonas aeruginosa populations in the cystic fibrosis lung lose susceptibility to newly applied ß-lactams within 3 days.

BACKGROUND: Chronic pulmonary infections by Pseudomonas aeruginosa require frequent intravenous antibiotic treatment in cystic fibrosis (CF) patients. Emergence of antimicrobial resistance is common in these patients, which to date has been investigated at long-term intervals only. OBJECTIVES: To investigate under close to real-time conditions the dynamics of the response by P. aeruginosa to a single course of antibiotic therapy and the potentially associated rapid spread of antimicrobial resistance, as well as the impact on the airway microbiome. METHODS: We investigated a cohort of adult CF patients that were treated with a single course of antimicrobial combination therapy. Using daily sampling during treatment, we quantified the expression of resistance by P. aeruginosa (median of six isolates per daily sample, 347 isolates in total), measured bacterial load by P. aeruginosa-specific quantitative PCR and characterized the airway microbiome with a 16S rRNA-based approach. WGS was performed to reconstruct intrapatient strain phylogenies. RESULTS: In two patients, we found rapid and large increases in resistance to meropenem and ceftazidime. Phylogenetic reconstruction of strain relationships revealed that resistance shifts are probably due to de novo evolution and/or the selection of resistant subpopulations. We observed high interindividual variation in the reduction of bacterial load, microbiome composition and antibiotic resistance. CONCLUSIONS: We show that CF-associated P. aeruginosa populations can quickly respond to antibiotic therapy and that responses are patient specific. Thus, resistance evolution can be a direct consequence of treatment, and drug efficacy can be lost much faster than usually assumed. The consideration of these patient-specific rapid resistance shifts can help to improve treatment of CF-associated infections, for example by deeper sampling of bacteria for diagnostics, repeated monitoring of pathogen susceptibility and switching between drugs.

Occupation-Associated Fatal Limbic Encephalitis Caused by Variegated Squirrel Bornavirus 1, Germany, 2013.

Limbic encephalitis is commonly regarded as an autoimmune-mediated disease. However, after the recent detection of zoonotic variegated squirrel bornavirus 1 in a Prevost's squirrel (Callosciurus prevostii) in a zoo in northern Germany, we retrospectively investigated a fatal case in an autoantibody-seronegative animal caretaker who had worked at that zoo. The virus had been discovered in 2015 as the cause of a cluster of cases of fatal encephalitis among breeders of variegated squirrels (Sciurus variegatoides) in eastern Germany. Molecular assays and immunohistochemistry detected a limbic distribution of the virus in brain tissue of the animal caretaker. Phylogenetic analyses demonstrated a spillover infection from the Prevost's squirrel. Antibodies against bornaviruses were detected in the patient's cerebrospinal fluid by immunofluorescence and newly developed ELISAs and immunoblot. The putative antigenic epitope was identified on the viral nucleoprotein. Other zoo workers were not infected; however, avoidance of direct contact with exotic squirrels and screening of squirrels are recommended.

Oral glutamine supplementation improves intestinal permeability dysfunction in a murine acute graft-vs.-host disease model.

Although a profound barrier dysfunction has been reported, little is known about the pathophysiological mechanism evoking gastrointestinal graft-vs.-host disease (GI-GvHD) and apparent therapeutic options. The aim of this study was to evaluate the influence of oral glutamine on the course of GI-GvHD in an acute semiallogenic graft-vs.-host disease (GvHD) in irradiated B6D2F1 mice. An acute semiallogenic GvHD was induced by intraperitoneal injection of lymphocytes from C57BL/6 mice to irradiated B6D2F1 mice. Half of the GvHD animals received oral glutamine supplementation for 6 days started at the time of lymphocyte transfer. Six days after induction of the semiallogenic GvHD, jejunum specimens were prepared. The expression of the proinflammatory cytokine TNF-α and the tight junction protein occludin was investigated by PCR. Histological changes along with the apoptotic response were evaluated and intestinal permeability was assessed. Animals with GvHD showed a strong increase in paracellular permeability as a sign of the disturbed barrier function. TNF-α expression was significantly increased and the expression of the tight junction protein occludin decreased. GvHD led to mucosal atrophy, crypt hyperplasia, crypt apoptosis, and a disintegration of the tight junctions. Glutamine-treated mice showed reduced expression of TNF-α, increased occludin expression, fewer histological changes in the jejunum, smaller number of apoptotic cells in the crypt, and reduced gastrointestinal permeability. In conclusion, oral glutamine seems to have beneficial effects on the severity of inflammatory changes in the course of GvHD and might be a therapeutic option.

Acute Kidney Injury in Patients with Severe ARDS Requiring Extracorporeal Membrane Oxygenation: Incidence, Prognostic Impact and Risk Factors.

(1) Background: Acute kidney injury (AKI) is a common but under-investigated complication in patients receiving extracorporeal membrane oxygenation (ECMO). We aimed to define the incidence and clinical course, as well as the predictors of AKI in adults receiving ECMO support. (2) Materials and Methods: This is a retrospective analysis of all patients undergoing veno-venous ECMO treatment in a tertiary care center between December 2008 and December 2017. The primary endpoint was the new occurrence of an AKI of stage 2 or 3 according to the Kidney Disease: Improving Global Outcomes (KDIGO) classification after ECMO implantation. (3) Results: During the observation period, 103 patients underwent veno-venous ECMO implantation. In total, 59 patients (57.3%) met the primary endpoint with an AKI of stage 2 or 3 and 55 patients (53.4%) required renal replacement therapy. Patients with an AKI of 2 or 3 suffered from more bleeding and infectious complications. Whereas weaning failure from ECMO (30/59 (50.8%) vs. 15/44 (34.1%), p = 0.08) and 30-day mortality (35/59 (59.3%) vs. 17/44 (38.6%), p = 0.06) only tended to be higher in the group with an AKI of stage 2 or 3, long-term survival of up to five years was significantly lower in the group with an AKI of stage 2 or 3 (p = 0.015). High lactate, serum creatinine, and ECMO pump-speed levels, and low platelets, a low base excess, and a low hematocrit level before ECMO were independent predictors of moderate to severe AKI. Primary hypercapnic acidosis was more common in AKI non-survivors (12 (32.4%) vs. 0 (0.0%), p < 0.01). Accordingly, pCO2-levels prior to ECMO implantation tended to be higher in AKI non-survivors (76.12 ± 27.90 mmHg vs. 64.44 ± 44.31 mmHg, p = 0.08). In addition, the duration of mechanical ventilation prior to ECMO-implantation tended to be longer (91.14 ± 108.16 h vs. 75.90 ± 86.81 h, p = 0.078), while serum creatinine (180.92 ± 115.72 mmol/L vs. 124.95 ± 77.77 mmol/L, p = 0.03) and bicarbonate levels were significantly higher in non-survivors (28.22 ± 8.44 mmol/L vs. 23.36 ± 4.19 mmol/L, p = 0.04). (4) Conclusion: Two-thirds of adult patients receiving ECMO suffered from moderate to severe AKI, with a significantly increased morbidity and long-term mortality.

Increased intestinal permeability and tight junction disruption by altered expression and localization of occludin in a murine graft versus host disease model.

BACKGROUND: Hematopoietic stem cell transplantation is increasingly performed for hematologic diseases. As a major side effect, acute graft versus host disease (GvHD) with serious gastrointestinal symptoms including diarrhea, gastrointestinal bleeding and high mortality can be observed. Because surveillance and biopsies of human gastrointestinal GvHD are difficult to perform, rare information of the alterations of the gastrointestinal barrier exists resulting in a need for systematic animal models. METHODS: To investigate the effects of GvHD on the intestinal barrier of the small intestine we utilized an established acute semi allogenic GvHD in C57BL/6 and B6D2F1 mice. RESULTS: By assessing the differential uptake of lactulose and mannitol in the jejunum, we observed an increased paracellular permeability as a likely mechanism for disturbed intestinal barrier function. Electron microscopy, immunohistochemistry and PCR analysis indicated profound changes of the tight-junction complex, characterized by downregulation of the tight junction protein occludin without any changes in ZO-1. Furthermore TNF-α expression was significantly upregulated. CONCLUSIONS: This analysis in a murine model of GvHD of the small intestine demonstrates serious impairment of intestinal barrier function in the jejunum, with an increased permeability and morphological changes through downregulation and localization shift of the tight junction protein occludin.

Utilization and determinants of use of non-pharmacological interventions in COPD: Results of the COSYCONET cohort.

BACKGROUND: Guidelines for chronic obstructive pulmonary disease (COPD) recommend supplementing pharmacotherapy with non-pharmacological interventions. Little is known about the use of such interventions by patients. We analyzed the utilization of a number of non-pharmacological interventions and identified potential determinants of use. METHODS: Based on self-reports, use of interventions (smoking cessation, influenza vaccination, physiotherapy, sports program, patient education, pulmonary rehabilitation) and recommendation to use were assessed in 1410 patients with COPD. The utilization was analyzed according to sex and severity of disease. Potential determinants of utilization included demographic variables and disease characteristics and were analyzed using logistic regression models. RESULTS: Influenza vaccination in the previous autumn/winter was reported by 73% of patients. About 19% were currently participating in a reimbursed sports program, 10% received physiotherapy, 38% were ever enrolled in an educational program, and 34% had ever participated in an outpatient or inpatient pulmonary rehabilitation program. Out of 553 current or former smokers, 24% had participated in a smoking cessation program. While reports of having received a recommendation to use mainly did not differ according to sex, women showed significantly (p < 0.05) higher utilization rates than men for all interventions except influenza vaccination. Smoking was a predictor for not having received a recommendation for utilization and also significantly associated with a reduced odds of utilization. We found a correlation between recommendation to use and utilization. CONCLUSIONS: Utilization of non-pharmacological interventions was lower in men and smokers. A recommendation or offer to use by the physician could help to increase uptake.

Terminal part of thoracic duct: high-resolution US imaging.

PURPOSE: To assess ultrasonographic (US) examination results of the cervical part of the thoracic duct, to provide standard diameters, and to evaluate the diameter of the cervical thoracic duct in certain diseases suspected to involve an abnormal load of chyle (liver, heart, and inflammatory bowel diseases). MATERIALS AND METHODS: The study was approved by the institutional review committee, and written informed consent was obtained from all subjects. Diameter and variations of the cervical thoracic duct were assessed by using US in 265 healthy volunteers (age range, 21-82 years) from a population-based study, in 196 subjects with documented liver cirrhosis (age range, 19-87 years), in 68 subjects with chronic hepatitis (age range, 17-73 years), in 39 subjects with congestive heart failure (age range, 46-85 years), and in 17 subjects with inflammatory bowel disease (age range, 18-66 years). US examinations were performed with high-resolution linear probes (7-12 MHz). RESULTS: A standard imaging approach guided by anatomic structures was established. Dynamic imaging of the chyle flow and valve function was possible. The thoracic duct was visualized in 564 (96%) of 585 examinations. The average thoracic duct diameter in healthy volunteers was 2.5 mm, which was independent of the subjects' age. The diameter was significantly higher in subjects with congestive heart failure (6.3 mm, P < .0001) and liver cirrhosis (5.6 mm, P < .0001). Anatomic variations were present in 27% of subjects. CONCLUSION: High-resolution US with linear probes allows assessment of the cervical thoracic duct with high detection rates. Recognition of local anatomy, diameter, and chyle flow may aid functional assessment.

Combination of National Quality Assurance Data Collection With a Standard Operating Procedure in Community-Acquired Pneumonia: A Win-Win Strategy?

PURPOSE: The primary contact for German physicians with national quality assurance in community-acquired pneumonia (CAP) is frequently experienced as time-consuming obligatory documentation. Since the regular feedback loop stretches up to 18 months, the immediate impact on quality is perceived as rather low. Ultimately, a method leading to increase in the quality of data collection, clarification on expected clinical treatment standards, and improvement in the acceptance and feedback mechanism is needed. METHODS: We developed a form merging data collection for quality indicators with a standard operating procedure (SOP) in CAP and implemented it in the daily routine of a university's department for internal medicine. Fulfillment of quality indicators before and after the implementation of the new form was measured. RESULTS: Critical parameters such as the documentation of breathing rate and clinical parameters at discharge strongly improved after implementation of the intervention. Uncritical parameters showed slight improvement or stable results at a high level. CONCLUSION: The combination of collection of quality data with a clinical SOP and context information may improve the impact of quality measures by increasing acceptance, quality of data capture, short-loop feedback, and possibly quality of care.

CAT score single item analysis in patients with COPD: Results from COSYCONET.

The COPD Assessment Test (CAT) is in widespread use for the evaluation of patients with chronic obstructive pulmonary disease (COPD). We assessed whether the CAT items carry additional information beyond the sum score regarding COPD characteristics including emphysema. Patients of GOLD grades 1 to 4 from the COPD cohort COSYCONET (German COPD and Systemic Consequences - Comorbidities Network) with complete CAT data were included (n = 2270), of whom 493 had chest CT evaluated for the presence of emphysema. Comorbidities and lung function were assessed following standardised procedures. Cross-sectional data analysis was based on multiple regression analysis of the single CAT items against a panel of comorbidities, lung function, or CT characteristics (qualitative score, 15th percentile of mean lung density), with age, BMI and gender as covariates. This was supported by exploratory factor analysis. Regarding the relationship to comorbidities and emphysema, there were marked differences between CAT items, especially items 1 and 2 versus 3 to 8. This grouping was basically confirmed by factor analysis. Items 4 and 5, and to a lower degree 1, 2 and 6, appeared to be informative regarding the presence of emphysema, whereas the total score was not or less informative. Regarding comorbidities, similar findings as for the total CAT score were obtained for the modified Medical Research Council scale (mMRC) which was also informative regarding emphysema. Our findings suggest that the usefulness of the CAT can be increased if evaluated on the basis of single items which may be indicating the presence of comorbidities and emphysema.

Standardized quantification of pulmonary fibrosis in histological samples.

The Ashcroft scale for the evaluation of bleomycin-induced lung fibrosis is the analysis of stained histological samples by visual assessment. Based on the knowledge that this procedure is not standardized in animals and results are highly variable, we hypothesized that modification of this method may improve quantification of lung fibrosis in small animals. To prove our hypothesis, we evaluated pulmonary fibrosis in Lewis rats induced by a single intratracheal injection of 0.3 mg/kg body weight bleomycin (n = 13) compared with the same amount of saline in a control group (n = 4). We modified the Ashcroft scale by precisely defining the assignment of grades from 0 to 8 for the increasing extent of fibrosis in lung histological samples. Thirty-two observers were randomly assigned to evaluate 108 photographs of slides using either the Ashcroft scale or the modified scale. Consistent with our hypothesis, there was a significant reduction in the variability of standard deviations with the modified scale compared with the Ashcroft scale (mean of variability 0.25 versus 0.62, P < 0.0001). Applying the kappa index, the Ashcroft scale showed only a fair to moderate agreement (0.23-0.59) between the observers and a low intra-observer agreement (0.51-0.74) in contrast to the modified scale, which demonstrated a moderate to good agreement between the observers (0.65-0.93, P < 0.0001) and a high intra-observer agreement (0.87-0.91, P < 0.05). To test the modified scale in vivo, we compared both scales with the results of computed tomography (CT) of the lungs obtained from the same mice. In agreement, the modified scale demonstrated a better correlation to CT scans (R = 0.58) compared with the Ashcroft scale (R = 0.33). In summary, quantification of lung fibrosis in histological lung sections using the modified scale is reliable and reproducible.

NT-proBNP is not elevated in patients with obstructive sleep apnoea.

BACKGROUND: N-terminal pro-brain natriuretic peptide (NT-ProBNP) has emerged as an important marker of cardiac stress and may reflect the severity of underlying cardiac dysfunction, which is thought to be associated with obstructive sleep apnoea syndrome (OSAS). METHODS: This study evaluated the plasma concentration of NT-ProBNP in 60 consecutive patients (median age 55.7 years, median body mass index (BMI) 31.8) who were referred to a sleep laboratory with a suspicion of OSAS. Each subject underwent measurement of morning NT-ProBNP plasma levels, polysomnography and echocardiography. Patients were treated with nasal continuous or bilevel positive airway pressure ventilation (nCPAP/BIPAP) or without mechanical respiratory support, depending on clinical symptoms and results of polysomnography. Three months after treatment of OSAS 28 of the patients were reassessed for re-evaluation of NT-ProBNP and polysomnography. RESULTS: Low or high levels of NT-proBNP were not associated with AHI and other sleep related indices (p>0.3). There was no correlation between NT-proBNP and AHI or other sleep related indices. In multiple regression analysis, NT-proBNP was significantly correlated with left ventricular ejection fraction, creatinine clearance and the presence of systemic arterial hypertension but not with AHI. CONCLUSIONS: Our results show by a robust multiple regression analysis, that NT-pro BNP is not associated with OSAS and NT-pro BNP cannot be used as a sensitive marker for underlying cardiovascular abnormalities in patients with OSAS.

Parenting stress in mothers with cystic fibrosis.

AIM: To assess the parenting experience of mothers with cystic fibrosis (CF) and to compare with normative data. METHODS: Cross-sectional study with a validated generic parental stress questionnaire (PSQ). This PSQ differentiates four components of parental stress: main factor "parental stress", compounding factor "role restrictions", protective factors "support from spouse", and "social support". Cut-off scores categorise results as "normal", "borderline" or "concerning". SAMPLE: Seventy-three women were informed by their local CF centre. Of these, 36 enrolled and had a first-born child aged 1-12 years (consistent with reference values of the PSQ). Of these, 31 (86%) returned the PSQ. Mean age of mothers was 32.6 years ± 6.9 years, mean age of first-born child was 5.2 years ± 3.4 years. Most of the mothers had one biological child, five women had two children and one had three children. RESULTS: Parental stress scores were normally distributed, the same applies for contributing factors and for the two protective factors. Favourable scores were twice as frequent as concerning scores. Mothers of younger children scored slightly better than mothers of school-aged children. CONCLUSION: In line with the only comparable study, mothers with CF seem to be a remarkably resilient group who mostly cope well with parental stress even in the face of a progressive, chronic disease requiring time-consuming treatment. IMPLICATIONS FOR REHABILITATION: Today, motherhood is increasingly becoming an option in fertile women with cystic fibrosis. The additional burden of parenting seems to be rewarded by fulfilling essential personal goals. CF clinics should routinely address a possible wish for a child and to discuss it, openly.

Lung Transplantation in a Multidrug-Resistant Gram-Negative Acinetobacter Baumannii-Colonized Patient: A Case Report.

Colonization or infection with various pathogens is frequently found in patients listed for lung transplantation. We describe a case of a 50-year-old woman with α-1-antitrypsin deficiency, which was listed for double-lung transplantation, with multidrug-resistant gram-negative Acinetobacter baumannii (MRGN4-Ab) skin colonization. Transplantation was successfully performed. Colistin (Polymyxine E) was administered intravenously and through inhalation in the first month. MRGN4-Ab was still detectable in skin swabs without evidence of infection. After good recovery and clinical inapparence, the patient was discharged 2 months after transplantation.

Clinical and laboratory effects of recombinant human activated protein C in the treatment of a patient with sepsis-induced multiple organ failure.

OBJECTIVES: To evaluate clinical and laboratory effects of the administration of recombinant human activated protein C (rhAPC) in the treatment of a 25-year-old patient with septic shock and multiple organ failure secondary to perinephritic abscesses. INTERVENTIONS: Intravenous administration of rhAPC-or drotrecogin alfa (activated)-(24 mcg/kg/h) for a total of 80 h as an adjunct to antimicrobial therapy, mechanical ventilation, hemodynamic support, hemodiafiltration and surgical intervention. MEASUREMENTS AND MAIN RESULTS: The administration of rhAPC was associated with a rapid recovery of the patient's clinical condition reflected by decreasing Sepsis-related Organ Failure Assessment (SOFA) scores. Laboratory parameters monitoring inflammation and coagulopathy improved during the treatment. No drug-related adverse events were noted. CONCLUSIONS: RhAPC has been observed to have anticoagulant, anti-inflammatory and profibrinolytic properties in vitro and in vivo. This report describes the effects of rhAPC administration on standard laboratory parameters indicating that no single laboratory parameter exists that is capable of monitoring the effects of rhAPC on the coagulation cascade and the clinical course of sepsis. This description of a patient suffering from sepsis-induced multiple organ failure may illustrate a possible beneficial effect of rhAPC on the course of coagulopathy and systemic inflammatory response and provides evidence for rhAPC complementing standard intensive care therapy in severe sepsis.

Analysis of the GM-CSF and GM-CSF/IL-3/IL-5 receptor common beta chain in a patient with pulmonary alveolar proteinosis.

OBJECTIVE: To investigate the expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) and GM-CSF/IL-3/IL-5 receptor common beta chain (beta c receptor) in an adult patient with idiopathic pulmonary alveolar proteinosis (PAP), so as to demonstrate the possible association of the GM-CSF and beta c receptor with the pathogenesis of human PAP. METHODS: The GM-CSF levels were measured with a commercial ELISA kit (sensitivity 5 pg/ml) and the beta c receptor expression on the cell surface was detected by flow cytometry analysis. Reverse transcription polymerase chain reaction (RT-PCR) analysis was employed to detect the expression of the GM-CSF mRNA and the beta c receptor mRNA in peripheral blood mononuclear cells and alveolar macrophages. The entire coding regions of the GM-CSF cDNA and the beta c receptor cDNA were sequenced by the Sanger dideoxy-mediated chain termination method to detect possible mutations. RESULTS: The patient with PAP failed to release the GM-CSF protein either from circulating mononuclear cells or from alveolar macrophages. The expression of the GM-CSF mRNA was normal after the stimulation of lipopolysaccharide, whereas a point mutation at position 382 of the GM-CSF cDNA from "T" to "C" was revealed by cDNA sequencing, which caused a change in amino acid 117 of the protein from isoleucine to threonine. The beta c receptor expression on the cell surface was normal, and the beta c receptor mRNA expression and the sequence of the entire coding region of the beta c receptor were also normal. CONCLUSIONS: The decreased GM-CSF production is associated with the pathogenesis of human PAP. A point mutation of the GM-CSF cDNA may contribute to the decreased GM-CSF production in our adult PAP patient. The mutation of the beta c receptor in some of paediatric patients with PAP may not be a common problem in adult patients.