GLI1+ Cells Contribute to Vascular Remodeling in Pulmonary Hypertension.

BACKGROUND: The precise origin of newly formed ACTA2+ (alpha smooth muscle actin-positive) cells appearing in nonmuscularized vessels in the context of pulmonary hypertension is still debatable although it is believed that they predominantly derive from preexisting vascular smooth muscle cells (VSMCs). METHODS: Gli1Cre-ERT2; tdTomatoflox mice were used to lineage trace GLI1+ (glioma-associated oncogene homolog 1-positive) cells in the context of pulmonary hypertension using 2 independent models of vascular remodeling and reverse remodeling: hypoxia and cigarette smoke exposure. Hemodynamic measurements, right ventricular hypertrophy assessment, flow cytometry, and histological analysis of thick lung sections followed by state-of-the-art 3-dimensional reconstruction and quantification using Imaris software were used to investigate the contribution of GLI1+ cells to neomuscularization of the pulmonary vasculature. RESULTS: The data show that GLI1+ cells are abundant around distal, nonmuscularized vessels during steady state, and this lineage contributes to around 50% of newly formed ACTA2+ cells around these normally nonmuscularized vessels. During reverse remodeling, cells derived from the GLI1+ lineage are largely cleared in parallel to the reversal of muscularization. Partial ablation of GLI1+ cells greatly prevented vascular remodeling in response to hypoxia and attenuated the increase in right ventricular systolic pressure and right heart hypertrophy. Single-cell RNA sequencing on sorted lineage-labeled GLI1+ cells revealed an Acta2high fraction of cells with pathways in cancer and MAPK (mitogen-activated protein kinase) signaling as potential players in reprogramming these cells during vascular remodeling. Analysis of human lung-derived material suggests that GLI1 signaling is overactivated in both group 1 and group 3 pulmonary hypertension and can promote proliferation and myogenic differentiation. CONCLUSIONS: Our data highlight GLI1+ cells as an alternative cellular source of VSMCs in pulmonary hypertension and suggest that these cells and the associated signaling pathways represent an important therapeutic target for further studies.

Vitamin D constrains inflammation by modulating the expression of key genes on Chr17q12-21.1.

Vitamin D possesses immunomodulatory functions and vitamin D deficiency has been associated with the rise in chronic inflammatory diseases, including asthma (Litonjua and Weiss, 2007). Vitamin D supplementation studies do not provide insight into the molecular genetic mechanisms of vitamin D-mediated immunoregulation. Here, we provide evidence for vitamin D regulation of two human chromosomal loci, Chr17q12-21.1 and Chr17q21.2, reliably associated with autoimmune and chronic inflammatory diseases. We demonstrate increased vitamin D receptor (Vdr) expression in mouse lung CD4+ Th2 cells, differential expression of Chr17q12-21.1 and Chr17q21.2 genes in Th2 cells based on vitamin D status and identify the IL-2/Stat5 pathway as a target of vitamin D signaling. Vitamin D deficiency caused severe lung inflammation after allergen challenge in mice that was prevented by long-term prenatal vitamin D supplementation. Mechanistically, vitamin D induced the expression of the Ikzf3-encoded protein Aiolos to suppress IL-2 signaling and ameliorate cytokine production in Th2 cells. These translational findings demonstrate mechanisms for the immune protective effect of vitamin D in allergic lung inflammation with a strong molecular genetic link to the regulation of both Chr17q12-21.1 and Chr17q21.2 genes and suggest further functional studies and interventional strategies for long-term prevention of asthma and other autoimmune disorders.

GLI1+ cells are a source of repair-supportive mesenchymal cells (RSMCs) during airway epithelial regeneration.

Repair-supportive mesenchymal cells (RSMCs) have been recently reported in the context of naphthalene (NA)-induced airway injury and regeneration. These cells transiently express smooth muscle actin (Acta2) and are enriched with platelet-derived growth factor receptor alpha (Pdgfra) and fibroblast growth factor 10 (Fgf10) expression. Genetic deletion of Ctnnb1 (gene coding for beta catenin) or Fgf10 in these cells using the Acta2-Cre-ERT2 driver line after injury (defined as NA-Tam condition; Tam refers to tamoxifen) led to impaired repair of the airway epithelium. In this study, we demonstrate that RSMCs are mostly captured using the Acta2-Cre-ERT2 driver when labeling occurs after (NA-Tam condition) rather than before injury (Tam-NA condition), and that their expansion occurs mostly between days 3 and 7 following NA treatment. Previous studies have shown that lineage-traced peribronchial GLI1+ cells are transiently amplified after NA injury. Here, we report that Gli1 expression is enriched in RSMCs. Using lineage tracing with Gli1Cre-ERT2 mice combined with genetic inactivation of Fgf10, we show that GLI1+ cells with Fgf10 deletion fail to amplify around the injured airways, thus resulting in impaired airway epithelial repair. Interestingly, Fgf10 expression is not upregulated in GLI1+ cells following NA treatment, suggesting that epithelial repair is mostly due to the increased number of Fgf10-expressing GLI1+ cells. Co-culture of SCGB1A1+ cells with GLI1+ cells isolated from non-injured or injured lungs showed that GLI1+ cells from these two conditions are similarly capable of supporting bronchiolar organoid (or bronchiolosphere) formation. Single-cell RNA sequencing on sorted lineage-labeled cells showed that the RSMC signature resembles that of alveolar fibroblasts. Altogether, our study provides strong evidence for the involvement of mesenchymal progenitors in airway epithelial regeneration and highlights the critical role played by Fgf10-expressing GLI1+ cells in this context.

Fgfr2b signaling is essential for the maintenance of the alveolar epithelial type 2 lineage during lung homeostasis in mice.

Fibroblast growth factor receptor 2b (Fgfr2b) signaling is essential throughout lung development to form the alveolar epithelial lineage. However, its role in alveolar epithelial type 2 cells (AT2s) homeostasis was recently considered dispensable. SftpcCreERT2; Fgfr2bflox/flox; tdTomatoflox/flox mice were used to delete Fgfr2b expression in cells belonging to the AT2 lineage, which contains mature AT2s and a novel SftpcLow lineage-traced population called "injury activated alveolar progenitors" or IAAPs. Upon continuous tamoxifen exposure for either 1 or 2 weeks to delete Fgfr2b, a shrinking of the AT2 population is observed. Mature AT2s exit the cell cycle, undergo apoptosis and fail to form alveolospheres in vitro. However, the lung morphometry appears normal, suggesting the involvement of compensatory mechanisms. In mutant lungs, IAAPs which escaped Fgfr2b deletion expand, display enhanced alveolosphere formation in vitro and increase drastically their AT2 signature, suggesting differentiation towards mature AT2s. Interestingly, a significant increase in AT2s and decrease in IAPPs occurs after a 1-week tamoxifen exposure followed by an 8-week chase period. Although mature AT2s partially recover their alveolosphere formation capabilities, the IAAPs no longer display this property. Single-cell RNA seq analysis confirms that AT2s and IAAPs represent stable and distinct cell populations and recapitulate some of their characteristics observed in vivo. Our results underscore the essential role played by Fgfr2b signaling in the maintenance of the AT2 lineage in the adult lung during homeostasis and suggest that the IAAPs could represent a new population of AT2 progenitors.

FGF10 Triggers De Novo Alveologenesis in a Bronchopulmonary Dysplasia Model: Impact on Resident Mesenchymal Niche Cells.

Bronchopulmonary dysplasia (BPD) is a neonatal lung disease developing in premature babies characterized by arrested alveologenesis and associated with decreased Fibroblast growth factor 10 (FGF10) expression. One-week hyperoxia (HYX) exposure of newborn mice leads to a permanent arrest in alveologenesis. To test the role of Fgf10 signaling to promote de novo alveologenesis following hyperoxia, we used transgenic mice allowing inducible expression of Fgf10 and recombinant FGF10 (rFGF10) protein delivered intraperitoneally. We carried out morphometry analysis, and IF on day 45. Alveolospheres assays were performed co-culturing AT2s from normoxia (NOX) with FACS-isolated Sca1Pos resident mesenchymal cells (rMC) from animals exposed to NOX, HYX-PBS, or HYX-FGF10. scRNAseq between rMC-Sca1Pos isolated from NOX and HYX-PBS was also carried out. Transgenic overexpression of Fgf10 and rFGF10 administration rescued the alveologenesis defects following HYX. Alveolosphere assays indicate that the activity of rMC-Sca1Pos is negatively impacted by HYX and partially rescued by rFGF10 treatment. Analysis by IF demonstrates a significant impact of rFGF10 on the activity of resident mesenchymal cells. scRNAseq results identified clusters expressing Fgf10, Fgf7, Pdgfra, and Axin2, which could represent the rMC niche cells for the AT2 stem cells. In conclusion, we demonstrate that rFGF10 administration is able to induce de novo alveologenesis in a BPD mouse model and identified subpopulations of rMC-Sca1Pos niche cells potentially representing its cellular target.

An essential function for autocrine hedgehog signaling in epithelial proliferation and differentiation in the trachea.

The tracheal epithelium is a primary target for pulmonary diseases as it provides a conduit for air flow between the environment and the lung lobes. The cellular and molecular mechanisms underlying airway epithelial cell proliferation and differentiation remain poorly understood. Hedgehog (HH) signaling orchestrates communication between epithelial and mesenchymal cells in the lung, where it modulates stromal cell proliferation, differentiation and signaling back to the epithelium. Here, we reveal a previously unreported autocrine function of HH signaling in airway epithelial cells. Epithelial cell depletion of the ligand sonic hedgehog (SHH) or its effector smoothened (SMO) causes defects in both epithelial cell proliferation and differentiation. In cultured primary human airway epithelial cells, HH signaling inhibition also hampers cell proliferation and differentiation. Epithelial HH function is mediated, at least in part, through transcriptional activation, as HH signaling inhibition leads to downregulation of cell type-specific transcription factor genes in both the mouse trachea and human airway epithelial cells. These results provide new insights into the role of HH signaling in epithelial cell proliferation and differentiation during airway development.

TMPRSS2 Is the Major Activating Protease of Influenza A Virus in Primary Human Airway Cells and Influenza B Virus in Human Type II Pneumocytes.

Cleavage of influenza virus hemagglutinin (HA) by host cell proteases is essential for virus infectivity and spread. We previously demonstrated in vitro that the transmembrane protease TMPRSS2 cleaves influenza A virus (IAV) and influenza B virus (IBV) HA possessing a monobasic cleavage site. Subsequent studies revealed that TMPRSS2 is crucial for the activation and pathogenesis of H1N1pdm and H7N9 IAV in mice. In contrast, activation of H3N2 IAV and IBV was found to be independent of TMPRSS2 expression and supported by an as-yet-undetermined protease(s). Here, we investigated the role of TMPRSS2 in proteolytic activation of IAV and IBV in three human airway cell culture systems: primary human bronchial epithelial cells (HBEC), primary type II alveolar epithelial cells (AECII), and Calu-3 cells. Knockdown of TMPRSS2 expression was performed using a previously described antisense peptide-conjugated phosphorodiamidate morpholino oligomer, T-ex5, that interferes with splicing of TMPRSS2 pre-mRNA, resulting in the expression of enzymatically inactive TMPRSS2. T-ex5 treatment produced efficient knockdown of active TMPRSS2 in all three airway cell culture models and prevented proteolytic activation and multiplication of H7N9 IAV in Calu-3 cells and H1N1pdm, H7N9, and H3N2 IAV in HBEC and AECII. T-ex5 treatment also inhibited the activation and spread of IBV in AECII but did not affect IBV activation in HBEC and Calu-3 cells. This study identifies TMPRSS2 as the major HA-activating protease of IAV in human airway cells and IBV in type II pneumocytes and as a potential target for the development of novel drugs to treat influenza infections.IMPORTANCE Influenza A viruses (IAV) and influenza B viruses (IBV) cause significant morbidity and mortality during seasonal outbreaks. Cleavage of the viral surface glycoprotein hemagglutinin (HA) by host proteases is a prerequisite for membrane fusion and essential for virus infectivity. Inhibition of relevant proteases provides a promising therapeutic approach that may avoid the development of drug resistance. HA of most influenza viruses is cleaved at a monobasic cleavage site, and a number of proteases have been shown to cleave HA in vitro This study demonstrates that the transmembrane protease TMPRSS2 is the major HA-activating protease of IAV in primary human bronchial cells and of both IAV and IBV in primary human type II pneumocytes. It further reveals that human and murine airway cells can differ in their HA-cleaving protease repertoires. Our data will help drive the development of potent and selective protease inhibitors as novel drugs for influenza treatment.

The key protein of endosomal mRNP transport Rrm4 binds translational landmark sites of cargo mRNAs.

RNA-binding proteins (RBPs) determine spatiotemporal gene expression by mediating active transport and local translation of cargo mRNAs. Here, we cast a transcriptome-wide view on the transported mRNAs and cognate RBP binding sites during endosomal messenger ribonucleoprotein (mRNP) transport in Ustilago maydis Using individual-nucleotide resolution UV crosslinking and immunoprecipitation (iCLIP), we compare the key transport RBP Rrm4 and the newly identified endosomal mRNP component Grp1 that is crucial to coordinate hyphal growth. Both RBPs bind predominantly in the 3' untranslated region of thousands of shared cargo mRNAs, often in close proximity. Intriguingly, Rrm4 precisely binds at stop codons, which constitute landmark sites of translation, suggesting an intimate connection of mRNA transport and translation. Towards uncovering the code of recognition, we identify UAUG as specific binding motif of Rrm4 that is bound by its third RRM domain. Altogether, we provide first insights into the positional organisation of co-localising RBPs on individual cargo mRNAs.

Benefits of High-Intensity Exercise Training to Patients with Chronic Obstructive Pulmonary Disease: A Controlled Study.

BACKGROUND: Various exercise training programs are used for patients with chronic obstructive pulmonary disease (COPD) of different severity. OBJECTIVES: To investigate the impact of individualized high-intensity training on exercise capacity with COPD. METHODS: A total of 49 patients agreed to participate. Of these, 31 were assigned to the training group and 18 served as controls. The training group exercised twice a week for 90 min with consecutively increasing loads. At the time of enrollment (T0), as well as after 3 (T1) and 6 (T2) months, a 6-min walk test (6-MWT) was performed and data on health-related quality of life, femoral muscle thickness, and various serum markers were obtained. RESULTS: The training group improved in their 6-MWT results (T0 = 407 ± 152 m vs. T1 = 459 ± 127 m, p = 0.002, vs. T2 = 483.2 ± 130.1 m, p = 0.004), in their cross-sectional area of the musculus rectus femoris (T0 = 6.2 ± 1.2 cm2 vs. T1 = 6.9 ± 1.2 cm2, p = 0.003, vs. 7.5 ± 1.6 cm2, p = 0.002), and in their St. George's Respiratory Questionnaire (SGRQ) score (T0 = 43.3 ± 18.0 vs. T1 = 36.0 ± 18.4, p = 0.001, vs. T2 = 34.7 ± 18. 0, p = 0.004). Serum levels of myostatin, irisin, resistin, and α-Klotho did not change significantly within the training period. Of note, the exercise group showed an inverse relationship between serum levels of resistin and those of α-Klotho after 6 months (r = -0.608, p = 0.021). CONCLUSIONS: COPD patients undergoing an individualized, structured, high-intensity training program improved their exercise capacity, gained muscle mass, and improved their quality of life.

Reduction of glutamate-induced excitotoxicity in murine primary neurons involving calpain inhibition.

Excessive glutamate secretion leads to excitotoxicity, which has been shown to underlie neurodegenerative disorders. Excitotoxicity is in part exerted by overactivation of calpains, which promote neuronal cell death via induction of limited proteolysis of the cellular proteins p35, regulatory subunit of cyclin-dependent kinase 5, and αII-spectrin. We used primary murine neuronal cells in a model of glutamate toxicity. The protease inhibitor α1-antitrypsin was able to prevent glutamate toxicity as determined by MTT assay and immunofluorescence. Calpain and caspase 3 activity were reduced following α1-antitrypsin treatment, as assessed by calpain and caspase 3 activity assays. In addition we could observe a modulation of cleavage of the calpain/caspase substrates αII-spectrin and p35 in Western blots. In summary, α1-antitrypsin shows inhibitory effects on excitotoxicity of primary neurons involving the inhibition of calpain activity. The advantage of using α1-antitrypsin is that the substance is already in clinical use for the treatment of patients with hereditary α1-antitrypsin deficiency. Further experiments are required in animal models of neurodegenerative disorders to assess the suitability of this substance in patients suffering from Alzheimer's disease or Parkinson's disease.

How Can We Improve the Detection of Alpha1-Antitrypsin Deficiency?

The Z deficiency in α1-antitrypsin (A1ATD) is an under-recognized condition. Alpha1-antitrypsin (A1AT) is the main protein in the α1-globulin fraction of serum protein electrophoresis (SPE); however, evaluation of the α1-globulin protein fraction has received very little attention. Serum Z-type A1AT manifests in polymeric forms, but their interference with quantitative immunoassays has not been reported. Here, 214 894 samples were evaluated by SPE at the G. Fracastoro Hospital of Verona, Italy. Patients with an A1AT level ≤ 0.92 g/L were recalled to complete A1ATD diagnosis. In parallel, to qualitatively and quantitatively characterize A1AT, sera samples from 10 PiZZ and 10 PiMM subjects obtained at the National Institute of Tuberculosis and Lung Diseases in Warsaw, Poland, were subjected to non-denaturing 7.5% PAGE and 7.5% SDS-PAGE followed by Western blot. Moreover, purified A1AT was heated at 60°C and analyzed by a non-denaturing PAGE and 4-15% gradient SDS-PAGE followed by Western blot as well as by isolelectrofocusing and nephelometry. A total of 966 samples manifested percentages ≤ 2.8 or a double band in the alpha1-zone. According to the nephelometry data, 23 samples were classified as severe (A1AT ≤ 0.49 g/L) and 462 as intermediate (A1AT >0.49≤ 1.0 g/L) A1ATD. Twenty subjects agreed to complete the diagnosis and an additional 21 subjects agreed to family screening. We detected 9 cases with severe and 26 with intermediate A1ATD. Parallel experiments revealed that polymerization of M-type A1AT, when measured by nephelometry or isolelectrofocusing, yields inaccurate results, leading to the erroneous impression that it was Z type and not M-type A1AT. We illustrate the need for confirmation of Z A1AT values by "state of the art" method. Clinicians should consider a more in-depth investigation of A1ATD in patients when they exhibit serum polymers and low α1-globulin protein levels by SPE.

Therapy with plasma purified alpha1-antitrypsin (Prolastin®) induces time-dependent changes in plasma levels of MMP-9 and MPO.

The common Z mutation (Glu342Lys) of α1-antitrypsin (A1AT) results in the polymerization and intracellular retention of A1AT protein. The concomitant deficiency of functional A1AT predisposes PiZZ subjects to early onset emphysema. Clinical studies have implied that, among the biomarkers associated with emphysema, matrix metalloproteinase 9 (MMP-9) is of particular importance. Increased plasma MMP-9 levels are proposed to predict the decline of lung function as well as greater COPD exacerbations in A1AT deficiency-associated emphysema. The aim of the present study was to investigate the effect of A1AT therapy (Prolastin) on plasma MMP-9 and myeloperoxidase (MPO) levels. In total 34 PiZZ emphysema patients were recruited: 12 patients without and 22 with weekly intravenous (60 mg/kg body weight) A1AT therapy. The quantitative analysis of A1AT, MMP-9 and MPO was performed in serum and in supernatants of blood neutrophils isolated from patients before and after therapy. Patients with Prolastin therapy showed significantly lower serum MMP-9 and MPO levels than those without therapy. However, parallel analysis revealed that a rapid infusion of Prolastin is accompanied by a transient elevation of plasma MMP-9 and MPO levels. Experiments with freshly isolated blood neutrophils confirmed that therapy with Prolastin causes transient MMP-9 and MPO release. Prolastin induced the rapid release of MMP-9 and MPO when added directly to neutrophil cultures and this reaction was associated with the presence of IgA in A1AT preparation. Our data support the conclusion that changes in plasma levels of MMP-9 and MPO mirror the effect of Prolastin on blood neutrophils.

α1-antitrypsin modulates microglial-mediated neuroinflammation and protects microglial cells from amyloid-ß-induced toxicity.

BACKGROUND: One hallmark of Alzheimer disease is microglial activation. Therapeutic approaches for this neurodegenerative disease include the modulation of microglial cells. α1-antitrypsin (A1AT) has been shown to exert anti-inflammatory effects on macrophages and lung epithelial cells and an inhibition of calpain activity in neutrophil granulocytes. Nothing is known about the effect of A1AT on microglial-mediated neuroinflammation. Our aim was to investigate the effect of A1AT on amyloid-ß (Aß)- and LPS-treated microglial cells in vitro with respect to cytokine production, stress pathways, cell viability, phagocytotic abilities and the underlying mechanisms. METHODS: Primary microglial cells were isolated from Swiss Webster mouse embryos on embryonic day 13.5. Cytokines in the supernatants of treated primary microglial cells were analyzed with ELISAs, and accumulated nitrite was detected with Griess reagents. Intracellular stress pathways were investigated in cell lysates using western blotting. Intracellular calcium levels were detected in BV-2 microglial cells loaded with the Ca2+-sensitive (fluorescent) dye Fluo-4. Calpain activity in primary microglial cells was assessed by using a calpain activity assay. Cell viability of Aß-treated microglial cells was analyzed using MTT assay. Phagocytosis of Aß was evaluated with western blot analysis. RESULTS: Upon co-administration, A1AT reduced pro-inflammatory mediators induced by LPS or Aß. Interestingly, we detected a reduction in calpain activity and in the concentration of intracellular calcium that might mediate the anti-inflammatory effects of A1AT. Inhibition of the classic activation pathways, such as phosphorylation of mitogen-activated protein kinases or activation of protein kinase A were excluded as a mechanism of A1AT-mediated effects. In addition, A1AT increased the viability of Aß-treated microglial cells and reduced Aß phagocytosis. CONCLUSIONS: We provide evidence on the mechanism of action of A1AT on microglial-mediated neuroinflammation in vitro. Our in vitro data indicate that A1AT treatment modulates microglial cells in inflammatory conditions and that this modulation is due to an inhibition of calpain activity and intracellular calcium levels. The underlying mechanisms of the effects observed here are promising for future therapeutic strategies and should thus be further pursued in transgenic mouse models of Alzheimer disease.

Benefits of whole body vibration training in patients hospitalised for COPD exacerbations - a randomized clinical trial.

BACKGROUND: Patients with stable COPD show improvements in exercise capacity and muscular function after the application of whole body vibration. We aimed to evaluate whether this modality added to conventional physiotherapy in exacerbated hospitalised COPD patients would be safe and would improve exercise capacity and quality of life. METHODS: 49 hospitalised exacerbated COPD patients were randomized (1:1) to undergo physiotherapy alone or physiotherapy with the addition of whole body vibration. The primary endpoint was the between-group difference of the 6-minute walking test (day of discharge - day of admission). Secondary assessments included chair rising test, quality of life, and serum marker analysis. RESULTS: Whole body vibration did not cause procedure-related adverse events. Compared to physiotherapy alone, it led to significantly stronger improvements in 6-minute walking test (95.55 ± 76.29 m vs. 6.13 ± 81.65 m; p = 0.007) and St. Georges Respiratory Questionnaire (-6.43 ± 14.25 vs. 5.59 ± 19.15, p = 0.049). Whole body vibration increased the expression of the transcription factor peroxisome proliferator receptor gamma coactivator-1-α and serum levels of irisin, while it decreased serum interleukin-8. CONCLUSION: Whole body vibration during hospitalised exacerbations did not cause procedure-related adverse events and induced clinically significant benefits regarding exercise capacity and health-related quality of life that were associated with increased serum levels of irisin, a marker of muscle activity. TRIAL REGISTRATION: German Clinical Trials Register DRKS00005979. Registered 17 March 2014.

A randomized clinical trial to assess the influence of a three months training program (gym-based individualized vs. calisthenics-based non-invidualized) in COPD-patients.

INTRODUCTION: Pulmonary rehabilitation has been demonstrated to improve exercise capacity, dyspnoea, quality of life and to reduce the adverse effects of acute exacerbations. Current guidelines recommend exercise training in patients with mild to very severe disease. However, there is insufficient data comparing the efficacy of different training approaches and intensities. METHODS: Between January 2009 and December 2012, 105 COPD patients were screened to participate in the study. 61 patients were randomly assigned into an individualized training group or into a non-individualized training group. Both groups exercised once a week for 60 minutes over a time period of three months. At the beginning and after three months, the following measurements were performed: 6-minute walking test (6-MWT), health-related quality of life (St. Georges Respiratory Questionnaire; SGRQ and COPD-Assessment-Test; CAT), M. rectus femoris cross-sectional area, and inflammatory markers in peripheral blood. RESULTS: Only in the individualized training group we observed a significant change of the 6-MWT (increase of 32.47 m; p = 0.012) and the cross-sectional area of the M. rectus fermoris (increase of 0.57 cm2; p = 0.049), while no significant changes occurred in the non-individualized training group. Peroxisome-proliferator-activated receptor-γ coactivator 1α increased in the individualized training only after the three months training period (increase of 0.43 relative copies; p = 0.017), all other myokines and inflammatory markers were not influenced by either of the programs. The total drop-out-rate was 44.3%. CONCLUSION: A low frequency outpatient training program may induce modest improvements in exercise capacity and muscle mass only if it is performed on an individualized basis.

Endosomal transport of septin mRNA and protein indicates local translation on endosomes and is required for correct septin filamentation.

Endosomes transport lipids and proteins over long distances by shuttling along microtubules. They also carry mRNAs on their surface, but the precise molecular function of this trafficking process is unknown. By live cell imaging of polarized fungal hyphae, we show microtubule-dependent transport of septin mRNA and encoded septin protein on the same shuttling endosomes. Consistent with the hypothesis that septin mRNA is translated on endosomes, the accumulation of septin protein on endosomes requires the recruitment of septin mRNA. Furthermore, ribosomal proteins co-localise with shuttling endosomes, but only if mRNA is present. Importantly, endosomal trafficking is essential for an efficient delivery of septin protein to filaments at growth poles, a process necessary to establish unipolar growth. Thus, we propose that local mRNA translation loads endosomes with septins for assembly and efficient delivery to septin filaments.

Detection of obstructive sleep apnoea by an electronic nose.

Diagnosis of obstructive sleep apnoea syndrome (OSAS) is technically demanding, cost-intensive and time-consuming. The measurement of volatile organic compounds by an electronic nose is an innovative method that determines distinct molecular patterns of exhaled breath in different patient groups. We addressed the following questions: What is the diagnostic accuracy of an electronic nose in the detection of OSAS and the ability to detect effects of standard therapy in patients with OSAS? Are these results related to changes in distinct markers of airway inflammation and extracellular remodelling? We included 40 OSAS patients and 20 healthy controls. Exhaled breath of all participants was analysed using the Cyranose 320 electronic nose. Pharyngeal washings were performed to sample the upper airway compartment. For statistical analysis linear discriminant analysis was employed. We identified a linear discriminant function separating OSAS from control (p<0.0001). The corresponding area under the receiver-operating curve was 0.85 (95% CI 0.75-0.96; sensitivity 0.93 and specificity 0.7). In pharyngeal washing fluids of OSAS patients, we observed higher levels of α1-antitrypsin and markers of extracellular remodelling compared to controls. The electronic nose can distinguish between OSAS patients and controls with high accuracy.