Clinical Features and Treatment Outcomes of Bone and Joint Nontuberculous Mycobacterial Infections According to Immune Status: a 9-year Retrospective Observational Cohort.

OBJECTIVES: Nontuberculous mycobacteria (NTM) bone and joint infections (BJIs) are uncommon. We evaluated the characteristics of BJIs and identified differences according to immune status. METHODS: We performed a multicenter retrospective study in France involving patients with documented NTM BJI over a 9-year period. We collected the clinical and microbiological characteristics, management, and clinical outcomes of the patients. RESULTS: Ninety-five patients were included, of whom 50.5% (48/95) were immunosuppressed. Tenosynovitis was more frequent in the immunocompetent group, and native arthritis more common in the immunosuppressed group. M. marinum and M. abscessus complex were significantly more frequent in the immunocompetent group, and M. avium and M. xenopi were significantly more frequent in the immunosuppressed group. The combination of antibiotherapy with surgery tended to be more frequent in the immunocompetent than the immunosuppressed group (63.8% (30/47) vs 47.8% (22/46), respectively); of the latter, 45.7% (21/46) received antimicrobial therapy alone, a higher frequency than in the immunocompetent group (23.4%, 11/47). The median duration of antimicrobial treatment was similar in the two groups (11 months). Mortality was significantly higher in the immunosuppressed group. CONCLUSIONS: Although the clinical presentations and the NTM species involved in BJI differed according to immune status, most recovered completely after treatment.

Bile salt hydrolase acyltransferase activity expands bile acid diversity.

Bile acids (BAs) are steroid detergents in bile that contribute to the absorption of fats and fat-soluble vitamins while shaping the gut microbiome because of their antimicrobial properties1-4. Here we identify the enzyme responsible for a mechanism of BA metabolism by the gut microbiota involving amino acid conjugation to the acyl-site of BAs, thus producing a diverse suite of microbially conjugated bile acids (MCBAs). We show that this transformation is mediated by acyltransferase activity of bile salt hydrolase (bile salt hydrolase/transferase, BSH/T). Clostridium perfringens BSH/T rapidly performed acyl transfer when provided various amino acids and taurocholate, glycocholate or cholate, with an optimum at pH 5.3. Amino acid conjugation by C. perfringens BSH/T was diverse, including all proteinaceous amino acids except proline and aspartate. MCBA production was widespread among gut bacteria, with strain-specific amino acid use. Species with similar BSH/T amino acid sequences had similar conjugation profiles and several bsh/t alleles correlated with increased conjugation diversity. Tertiary structure mapping of BSH/T followed by mutagenesis experiments showed that active site structure affects amino acid selectivity. These MCBA products had antimicrobial properties, where greater amino acid hydrophobicity showed greater antimicrobial activity. Inhibitory concentrations of MCBAs reached those measured natively in the mammalian gut. MCBAs fed to mice entered enterohepatic circulation, in which liver and gallbladder concentrations varied depending on the conjugated amino acid. Quantifying MCBAs in human faecal samples showed that they reach concentrations equal to or greater than secondary and primary BAs and were reduced after bariatric surgery, thus supporting MCBAs as a significant component of the BA pool that can be altered by changes in gastrointestinal physiology. In conclusion, the inherent acyltransferase activity of BSH/T greatly diversifies BA chemistry, creating a set of previously underappreciated metabolites with the potential to affect the microbiome and human health.

Longitudinal microbial and molecular dynamics in the cystic fibrosis lung after Elexacaftor-Tezacaftor-Ivacaftor therapy.

BACKGROUND: Cystic fibrosis (CF) is a genetic disorder causing poor mucociliary clearance in the airways and subsequent respiratory infection. The recently approved triple therapy Elexacaftor-Tezacaftor-Ivacaftor (ETI) has significantly improved lung function and decreased airway infection in persons with CF (pwCF). This improvement has been shown to occur rapidly, within the first few weeks of treatment. The effects of longer term ETI therapy on lung infection dynamics, however, remain mostly unknown. RESULTS: Here, we applied 16S rRNA gene amplicon sequencing, untargeted metabolomics, and neutral models to high-resolution, longitudinally collected sputum samples from pwCF on ETI therapy (162 samples, 7 patients) and compared to similarly collected data set from pwCF not taking ETI (630 samples, 9 patients). Because ETI reduces sputum production, samples were collected in freezers provided in the subject's homes at least 3 months after first taking ETI, with those on ETI collecting a sample approximately weekly. The lung function (%ppFEV1) of those in our longitudinal cohort significantly improved after ETI (6.91, SD = 7.74), indicating our study cohort was responsive to ETI. The daily variation of alpha- and beta-diversity of both the microbiome and metabolome was higher for those on ETI, reflecting a more dynamic microbial community and chemical environment during treatment. Four of the seven subjects on ETI were persistently infected with Pseudomonas or Burkholderia in their sputum throughout the sampling period while the total bacterial load significantly decreased with time (R = - 0.42, p = 0.01) in only one subject. The microbiome and metabolome dynamics on ETI were personalized, where some subjects had a progressive change with time on therapy, whereas others had no association with time on treatment. To further classify the augmented variance of the CF microbiome under therapy, we fit the microbiome data to a Hubbell neutral dynamics model in a patient-stratified manner and found that the subjects on ETI had better fit to a neutral model. CONCLUSION: This study shows that the longitudinal microbiology and chemistry in airway secretions from subjects on ETI has become more dynamic and neutral and that after the initial improvement in lung function, many are still persistently infected with CF pathogens.

Longitudinal Microbial and Molecular Dynamics in the Cystic Fibrosis Lung after Elexacaftor-Tezacaftor-Ivacaftor therapy.

Background: Cystic fibrosis (CF) is a genetic disorder causing poor mucociliary clearance in the airways and subsequent respiratory infection. The recently approved triple therapy Elexacaftor-Tezacaftor-Ivacaftor (ETI) has significantly improved the lung function and decreased airway infection of persons with CF (pwCF). This improvement has been shown to occur rapidly, within the first few weeks of treatment. The effects of longer term ETI therapy on lung infection dynamics, however, remains mostly unknown. Results: Here, we applied 16S rRNA gene amplicon sequencing, untargeted metabolomics, and neutral models to high-resolution, longitudinally collected sputum samples from pwCF on ETI therapy (162 samples, 7 patients) and compared to similarly collected data set of CF subjects not taking ETI (630 samples, 9 patients). Because ETI reduces sputum production, samples were collected in freezers provided in the subject's homes at least 3 months after first taking ETI, with those on ETI collecting a sample approximately weekly. The lung function (%ppFEV1) of those in our longitudinal cohort significantly improved after ETI (6.91, SD = 7.74), indicating our study cohort was responsive to ETI. The daily variation of alpha- and beta-diversity of both the microbiome and metabolome was higher for those on ETI, reflecting a more dynamic microbial community and chemical environment during treatment. Four of the seven subjects on ETI were persistently infected with Pseudomonas or Burkholderia in their sputum throughout the sampling period. The microbiome and metabolome dynamics on ETI were personalized, where some subjects had a progressive change with time on therapy, whereas others had no association with time on treatment. To further classify the augmented variance of the CF microbiome under therapy, we fit the microbiome data to a Hubbell neutral dynamics model in a patient-stratified manner and found that the subjects on ETI had better fit to a neutral model. Conclusion: This study shows that the longitudinal microbiology and chemistry in airway secretions from subjects on ETI has become more dynamic and neutral, and that after the initial improvement in lung function, many are still persistently infected with CF pathogens.

Single-polyp metabolomics reveals biochemical structuring of the coral holobiont at multiple scales.

All biology happens in space, and spatial structuring plays an important role in mediating biological processes at all scales from cells to ecosystems. However, the metabolomic structuring of the coral holobiont has yet to be fully explored. Here, we present a method to detect high-quality metabolomic data from individual coral polyps and apply this method to study the patterning of biochemicals across multiple spatial (~1 mm - ~100 m) and organizational scales (polyp to population). The data show a strong signature for individual coral colonies, a weaker signature of branches within colonies, and variation at the polyp level related to the polyps' location along a branch. Mapping metabolites to either the coral or algal components of the holobiont reveals that polyp-level variation along the length of a branch was largely driven by molecules associated with the cnidarian host as opposed to the algal symbiont, predominantly putative sulfur-containing metabolites. This work yields insights on the spatial structuring of biochemicals in the coral holobiont, which is critical for design, analysis, and interpretation of studies on coral reef biochemistry.

Mechanical activation of lung epithelial cells through the ion channel Piezo1 activates the metalloproteinases ADAM10 and ADAM17 and promotes growth factor and adhesion molecule release.

In the lung, pulmonary epithelial cells undergo mechanical stretching during ventilation. The associated cellular mechanoresponse is still poorly understood at the molecular level. Here, we demonstrate that activation of the mechanosensitive cation channel Piezo1 in a human epithelial cell line (H441) and in primary human lung epithelial cells induces the proteolytic activity of the metalloproteinases ADAM10 and ADAM17 at the plasma membrane. These ADAMs are known to convert cell surface expressed proteins into soluble and thereby play major roles in proliferation, barrier regulation and inflammation. We observed that chemical activation of Piezo1 promotes cleavage of substrates that are specific for either ADAM10 or ADAM17. Activation of Piezo1 also induced the synthesis and ADAM10/17-dependent release of the growth factor amphiregulin (AREG). In addition, junctional adhesion molecule A (JAM-A) was shed in an ADAM10/17-dependent manner resulting in a reduction of cell contacts. Stretching experiments combined with Piezo1 knockdown further demonstrated that mechanical activation promotes shedding via Piezo1. Most importantly, high pressure ventilation of murine lungs increased AREG and JAM-A release into the alveolar space, which was reduced by a Piezo1 inhibitor. Our study provides a novel link between stretch-induced Piezo1 activation and the activation of ADAM10 and ADAM17 in lung epithelium. This may help to understand acute respiratory distress syndrome (ARDS) which is induced by ventilation stress and goes along with perturbed epithelial permeability and release of growth factors.

Welding Fume Instillation in Isolated Perfused Mouse Lungs-Effects of Zinc- and Copper-Containing Welding Fumes.

Zinc- and copper-containing welding fumes can cause systemic inflammation after exposure in humans. Recent ex vivo studies have shown that the observed inflammation originates from exposed immune cells. In vitro studies identified the soluble fraction of metal particles as the main effectors. Isolated perfused mouse lungs (IPLs) were perfused and ventilated for 270 min. Lungs were instilled with saline solution (control), welding fume particle suspension (WFs) or the soluble fraction of the welding fumes (SF-WFs). Bronchoalveolar lavage fluid (BALF) and perfusate samples were analyzed for cytokine levels and lung tissue mRNA expression levels were analyzed via RT-PCR. All lungs instilled with WFs did not complete the experiments due to a fatal reduction in tidal volume. Accordingly, IL-6 and MPO levels were significantly higher in BALF of WF lungs compared to the control. IL-6 and MPO mRNA expression levels were also increased for WFs. Lungs instilled with SF-WFs only showed mild reactions in tidal volume, with BALF and mRNA expression levels not significantly differing from the control. Zinc- and copper-containing welding fume particles adversely affect IPLs when instilled, as evidenced by the fatal loss in tidal volume and increased cytokine expression and secretion. The effects are mainly caused by the particles, not by the soluble fraction.

Platelet-derived growth factor (PDGF)-BB regulates the airway tone via activation of MAP2K, thromboxane, actin polymerisation and Ca2+-sensitisation.

BACKGROUND: PDGFR-inhibition by the tyrosine kinase inhibitor (TKI) nintedanib attenuates the progress of idiopathic pulmonary fibrosis (IPF). However, the effects of PDGF-BB on the airway tone are almost unknown. We studied this issue and the mechanisms beyond, using isolated perfused lungs (IPL) of guinea pigs (GPs) and precision-cut lung slices (PCLS) of GPs and humans. METHODS: IPL: PDGF-BB was perfused after or without pre-treatment with the TKI imatinib (perfused/nebulised) and its effects on the tidal volume (TV), the dynamic compliance (Cdyn) and the resistance were studied. PCLS (GP): The bronchoconstrictive effects of PDGF-BB and the mechanisms beyond were evaluated. PCLS (human): The bronchoconstrictive effects of PDGF-BB and the bronchorelaxant effects of imatinib were studied. All changes of the airway tone were measured by videomicroscopy and indicated as changes of the initial airway area. RESULTS: PCLS (GP/human): PDGF-BB lead to a contraction of airways. IPL: PDGF-BB decreased TV and Cdyn, whereas the resistance did not increase significantly. In both models, inhibition of PDGFR-(ß) (imatinib/SU6668) prevented the bronchoconstrictive effect of PDGF-BB. The mechanisms beyond PDGF-BB-induced bronchoconstriction include activation of MAP2K and TP-receptors, actin polymerisation and Ca2+-sensitisation, whereas the increase of Ca2+ itself and the activation of EP1-4-receptors were not of relevance. In addition, imatinib relaxed pre-constricted human airways. CONCLUSIONS: PDGFR regulates the airway tone. In PCLS from GPs, this regulatory mechanism depends on the ß-subunit. Hence, PDGFR-inhibition may not only represent a target to improve chronic airway disease such as IPF, but may also provide acute bronchodilation in asthma. Since asthma therapy uses topical application. This is even more relevant, as nebulisation of imatinib also appears to be effective.

Never Change a Flowing System? The Effects of Retrograde Flow on Isolated Perfused Lungs and Vessels.

Retrograde perfusion may occur during disease, surgery or extracorporeal circulation. While it is clear that endothelial cells sense and respond to changes in blood flow, the consequences of retrograde perfusion are only poorly defined. Similar to shear stress or disturbed flow, retrograde perfusion might result in vasomotor responses, edema formation or inflammation in and around vessels. In this study we investigated in rats the effects of retrograde perfusion in isolated systemic vessels (IPV) and in pulmonary vessels of isolated perfused lungs (IPL). Anterograde and retrograde perfusion was performed for 480 min in IPV and for 180 min in the IPL. Perfusion pressure, cytokine levels in perfusate and bronchoalveolar lavage fluid (BALF), edema formation and mRNA expression were studied. In IPV, an increased perfusion pressure and initially also increased cytokine levels were observed during retrograde perfusion. In the IPL, increased edema formation occurred, while cytokine levels were not increased, though dilution of cytokines in BALF due to pulmonary edema cannot be excluded. In conclusion, effects of flow reversal were visible immediately after initiation of retrograde perfusion. Pulmonary edema formation was the only effect of the 3 h retrograde perfusion. Therefore, further research should focus on identification of possible long-term complications of flow reversal.

Reevaluation of Lung Injury in TNF-Induced Shock: The Role of the Acid Sphingomyelinase.

Tumor necrosis factor (TNF) is a well-known mediator of sepsis. In many cases, sepsis results in multiple organ injury including the lung with acute respiratory distress syndrome (ARDS). More than 20-year-old studies have suggested that TNF may be directly responsible for organ injury during sepsis. However, these old studies are inconclusive, because they relied on human rather than conspecific TNF, which was contaminated with endotoxin in most studies. In this study, we characterized the direct effects of intravenous murine endotoxin-free TNF on cardiovascular functions and organ injury in mice with a particular focus on the lungs. Because of the relevance of the acid sphingomyelinase in sepsis, ARDS, and caspase-independent cell death, we also included acid sphingomyelinase-deficient (ASM-/-) mice. ASM-/- and wild-type (WT) mice received 50 µg endotoxin-free murine TNF intravenously alone or in combination with the pan-caspase inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone (zVAD) and were ventilated at low tidal volume while lung mechanics were followed. Blood pressure was stabilized by intra-arterial fluid support, and body temperature was kept at 37°C to delay lethal shock and to allow investigation of blood gases, lung histopathology, proinflammatory mediators, and microvascular permeability 6 hours after TNF application. Besides the lungs, also the kidneys and liver were examined. TNF elicited the release of inflammatory mediators and a high mortality rate, but failed to injure the lungs, kidneys, or liver of healthy mice significantly within 6 hours. Mortality in WT mice was most likely due to sepsis-like shock, as indicated by metabolic acidosis, high procalcitonin levels, and cardiovascular failure. ASM-/- mice were protected from TNF-induced hypotension and reflex tachycardia and also from mortality. In WT mice, intravenous exogenous TNF does not cause organ injury but induces a systemic inflammatory response with cardiovascular failure, in which the ASM plays a role.

Tolerability of facial electrostimulation in healthy adults and patients with facial synkinesis.

PURPOSE: To evaluate optimal stimulation parameters with regard to discomfort and tolerability for transcutaneous electrostimulation of facial muscles in healthy participants and patients with postparetic facial synkinesis. METHODS: Two prospective studies were performed. First, single pulse monophasic stimulation with rectangular pulses was compared to triangular pulses in 48 healthy controls. Second, 30 healthy controls were compared to 30 patients with postparetic facial synkinesis with rectangular pulse form. Motor twitch threshold, tolerability threshold, and discomfort were assessed using a numeric rating scale at both thresholds. RESULTS: Discomfort at motor threshold was significantly lower for rectangular than for triangular pulses. Average motor and tolerability thresholds were higher for patients than for healthy participants. Discomfort at motor threshold was significantly lower for healthy controls compared to patients. Major side effects were not seen. CONCLUSIONS: Surface electrostimulation for selective functional and tolerable facial muscle contractions in patients with postparetic facial synkinesis is feasible.

Acid sphingomyelinase regulates TH 2 cytokine release and bronchial asthma.

BACKGROUND: Allergic diseases and especially allergic asthma are widespread diseases with high prevalence in childhood, but also in adults. Acid sphingomyelinase (ASM) is a key regulator of the sphingolipid pathway. Previous studies defined the association of ASM with the pathogenesis of TH 1-directed lung diseases like cystic fibrosis and acute lung injury. Here, we define the role of ASM in TH 2-regulated allergic bronchial asthma. METHODS: To determine the role of Asm under baseline conditions, wild-type (WT) and Asm-/- mice were ventilated with a flexiVent setup and bronchial hyperresponsiveness was determined using acetylcholine. Flow cytometry and cytokine measurements in bronchoalveolar lavage fluid and lung tissue were followed by in vitro TH 2 differentiations with cells from WT and Asm-/- mice and blockade of Asm with amitriptyline. As proof of principle, we conducted an ovalbumin-induced model of asthma in WT- and Asm-/-  mice. RESULTS: At baseline, Asm-/- mice showed better lung mechanics, but unaltered bronchial hyperresponsiveness. Higher numbers of Asm-/- T cells in bronchoalveolar lavage fluid released lower levels of IL-4 and IL-5, and these results were paralleled by decreased production of typical TH 2 cytokines in Asm-/- T lymphocytes in vitro. This phenotype could be imitated by incubation of T cells with amitriptyline. In the ovalbumin asthma model, Asm-/- animals were protected from high disease activity and showed better lung functions and lower levels of eosinophils and TH 2 cytokines. CONCLUSION: Asm deficiency could induce higher numbers of TH 2 cells in the lung, but those cells release decreased TH 2 cytokine levels. Hereby, Asm-/- animals are protected from bronchial asthma, which possibly offers novel therapeutic strategies, for example, with ASM blockade.

Retrograde perfusion in isolated perfused mouse lungs-Feasibility and effects on cytokine levels and pulmonary oedema formation.

Retrograde lung vascular perfusion can appear in high-risk surgeries. The present report is the first to study long-term retrograde perfusion of isolated perfused mouse lungs (IPLs) and to use the tyrosine kinase ephB4 and its ligand ephrinB2 as potential markers for acute lung injury. Mouse lungs were subjected to anterograde or retrograde perfusion with normal-pressure ventilation (NV) or high-pressure ventilation (=overventilation, OV) for 4 hours. Outcome parameters were cytokine, ephrinB2 and ephB4 levels in perfusate samples and bronchoalveolar lavage (BAL), and the wet-to-dry ratio. Anterograde perfusion was feasible for 4 hours, while lungs receiving retrograde perfusion presented considerable collapse rates. Retrograde perfusion resulted in an increased wet-to-dry ratio when combined with high-pressure ventilation; other physiological parameters were not affected. Cytokine levels in BAL and perfusate, as well as levels of soluble ephB4 in BAL were increased in OV, while soluble ephrinB2 BAL levels were increased in retrograde perfusion. BAL levels of ephrinB2 and ephB4 were also determined in vivo, including mice ventilated for 7 hours with normal-volume ventilation (NVV) or high-volume ventilation (HVV) with increased levels of ephB4 in HVV BAL compared to NVV. Retrograde perfusion in IPL is limited as a routine method to investigate effects due to collapse for yet unclear reasons. If successful, retrograde perfusion has an influence on pulmonary oedema formation. In BAL, ephrinB2 seems to be up-regulated by flow reversal, while ephB4 is a marker for acute lung injury.

PDGF-BB regulates the pulmonary vascular tone: impact of prostaglandins, calcium, MAPK- and PI3K/AKT/mTOR signalling and actin polymerisation in pulmonary veins of guinea pigs.

BACKGROUND: Platelet-derived growth factor (PDGF)-BB and its receptor PDGFR are highly expressed in pulmonary hypertension (PH) and mediate proliferation. Recently, we showed that PDGF-BB contracts pulmonary veins (PVs) and that this contraction is prevented by inhibition of PDGFR-ß (imatinib/SU6668). Here, we studied PDGF-BB-induced contraction and downstream-signalling in isolated perfused lungs (IPL) and precision-cut lung slices (PCLS) of guinea pigs (GPs). METHODS: In IPLs, PDGF-BB was perfused after or without pre-treatment with imatinib (perfused/nebulised), the effects on the pulmonary arterial pressure (PPA), the left atrial pressure (PLA) and the capillary pressure (Pcap) were studied and the precapillary (Rpre) and postcapillary resistance (Rpost) were calculated. Perfusate samples were analysed (ELISA) to detect the PDGF-BB-induced release of prostaglandin metabolites (TXA2/PGI2). In PCLS, the contractile effect of PDGF-BB was evaluated in pulmonary arteries (PAs) and PVs. In PVs, PDGF-BB-induced contraction was studied after inhibition of PDGFR-α/ß, L-Type Ca2+-channels, ROCK/PKC, prostaglandin receptors, MAP2K, p38-MAPK, PI3K-α/γ, AKT/PKB, actin polymerisation, adenyl cyclase and NO. Changes of the vascular tone were measured by videomicroscopy. In PVs, intracellular cAMP was measured by ELISA. RESULTS: In IPLs, PDGF-BB increased PPA, Pcap and Rpost. In contrast, PDGF-BB had no effect if lungs were pre-treated with imatinib (perfused/nebulised). In PCLS, PDGF-BB significantly contracted PVs/PAs which was blocked by the PDGFR-ß antagonist SU6668. In PVs, inhibition of actin polymerisation and inhibition of L-Type Ca2+-channels reduced PDGF-BB-induced contraction, whereas inhibition of ROCK/PKC had no effect. Blocking of EP1/3- and TP-receptors or inhibition of MAP2K-, p38-MAPK-, PI3K-α/γ- and AKT/PKB-signalling prevented PDGF-BB-induced contraction, whereas inhibition of EP4 only slightly reduced it. Accordingly, PDGF-BB increased TXA2 in the perfusate, whereas PGI2 was increased in all groups after 120 min and inhibition of IP-receptors did not enhance PDGF-BB-induced contraction. Moreover, PDGF-BB increased cAMP in PVs and inhibition of adenyl cyclase enhanced PDGF-BB-induced contraction, whereas inhibition of NO-formation only slightly increased it. CONCLUSIONS: PDGF-BB/PDGFR regulates the pulmonary vascular tone by the generation of prostaglandins, the increase of calcium, the activation of MAPK- or PI3K/AKT/mTOR signalling and actin remodelling. More insights in PDGF-BB downstream-signalling may contribute to develop new therapeutics for PH.

Assessment of the Cytotoxic and Immunomodulatory Effects of Substances in Human Precision-cut Lung Slices.

Respiratory diseases in their broad diversity need appropriate model systems to understand the underlying mechanisms and enable development of new therapeutics. Additionally, registration of new substances requires appropriate risk assessment with adequate testing systems to avoid the risk of individuals being harmed, for example, in the working environment. Such risk assessments are usually conducted in animal studies. In view of the 3Rs principle and public skepticism against animal experiments, human alternative methods, such as precision-cut lung slices (PCLS), have been evolving. The present paper describes the ex vivo technique of human PCLS to study the immunomodulatory potential of low-molecular-weight substances, such as ammonium hexachloroplatinate (HClPt). Measured endpoints include viability and local respiratory inflammation, marked by altered secretion of cytokines and chemokines. Pro-inflammatory cytokines, tumor necrosis factor alpha (TNF-α), and interleukin 1 alpha (IL-1α) were significantly increased in human PCLS after exposure to a sub-toxic concentration of HClPt. Even though the technique of PCLS has been substantially optimized over the past decades, its applicability for the testing of immunomodulation is still in development. Therefore, the results presented here are preliminary, even though they show the potential of human PCLS as a valuable tool in respiratory research.

Emergence of Nontuberculous Mycobacterial Lymphadenitis in Children After the Discontinuation of Mandatory Bacillus Calmette and GuÉrin Immunization in France.

Eighty-five children were diagnosed with culture-confirmed nontuberculous mycobacterial cervical lymphadenitis within the MYCOMED surveillance network from 2004 to 2013. The mean incidence sharply increased from 0.57 to 3.7 per 100,000 children per year, after the discontinuation of mandatory bacillus Calmette and Guérin immunization in 2007. Cases were documented as Mycobacterium avium (62.3%), Mycobacterium intracellulare (15.3%) and Mycobacterium lentiflavum (12.9%). Outcome was favorable in all, with or without surgery or antimycobacterial treatment.

Imatinib relaxes the pulmonary venous bed of guinea pigs.

BACKGROUND: Recently, the IMPRES study revealed that systemic imatinib improves exercise capacity in patients with advanced pulmonary arterial hypertension. Imatinib blocks the tyrosine kinase activity of the platelet-derived growth factor (PDGF)-receptor (PDGFR), acts antiproliferative and relaxes pulmonary arteries. However so far, the relaxant effects of imatinib on pulmonary veins (PVs) and on the postcapillary resistance are unknown, although pulmonary hypertension (PH) due to left heart disease (LHD) is most common and primarily affects PVs. Next, it is unknown whether activation of PDGFR alters the pulmonary venous tone. Due to the reported adverse effects of systemic imatinib, we evaluated the effects of nebulized imatinib on the postcapillary resistance. METHODS: Precision-cut lung slices (PCLS) were prepared from guinea pigs. PVs were pre-constricted with Endothelin-1 (ET-1) and the imatinib-induced relaxation was studied by videomicroscopy; PDGF-BB-related vascular properties were evaluated as well. The effects of perfused/nebulized imatinib on the postcapillary resistance were studied in cavine isolated perfused lungs (IPL). Intracellular cAMP/cGMP was measured by ELISA in PVs. RESULTS: In PCLS, imatinib (100 µM) relaxed pre-constricted PVs (126%). In PVs, imatinib increased cAMP, but not cGMP and inhibition of adenyl cyclase or protein kinase A reduced the imatinib-induced relaxation. Further, inhibition of KATP-channels, [Formula: see text]-channels or Kv-channels diminished the imatinib-induced relaxation, whereas inhibition of NO-signaling was without effect. In the IPL, perfusion or nebulization of imatinib reduced the ET-1-induced increase of the postcapillary resistance. In PCLS, PDGF-BB contracted PVs, which was blocked by imatinib and by the PDGFR-ß kinase inhibitor SU6668, whereas inhibition of PDGFR-α (ponatinib) had no significant effect. Conversely, PDGFR-ß kinase inhibitors (SU6668/DMPQ) relaxed PVs pre-constricted with ET-1 comparable to imatinib, whereas the PDGFR-α kinase inhibitor ponatinib did not. CONCLUSIONS: Imatinib-induced relaxation depends on cAMP and on the activation of K+-channels. Perfused or nebulized imatinib significantly reduces the postcapillary resistance in the pre-constricted (ET-1) pulmonary venous bed. Hence, nebulization of imatinib is feasible and might reduce systemic side effects. Conversely, PDGF-BB contracts PVs by activation of PDGFR-ß suggesting that imatinib-induced relaxation depends on PDGFR-ß-antagonism. Imatinib combines short-term relaxant and long-term antiproliferative effects. Thus, imatinib might be a promising therapy for PH due to LHD.

Variability of word discrimination scores in clinical practice and consequences on their sensitivity to hearing loss.

Speech perception scores are widely used to assess patient's functional hearing, yet most linguistic material used in these audiometric tests dates to before the availability of large computerized linguistic databases. In an ENT clinic population of 120 patients with median hearing loss of 43-dB HL, we quantified the variability and the sensitivity of speech perception scores to hearing loss, measured using disyllabic word lists, as a function of both the number of ten-word lists and type of scoring used (word, syllables or phonemes). The mean word recognition scores varied significantly across lists from 54 to 68%. The median of the variability of the word recognition score ranged from 30% for one ten-word list down to 20% for three ten-word lists. Syllabic and phonemic scores showed much less variability with standard deviations decreasing by 1.15 with the use of syllabic scores and by 1.45 with phonemic scores. The sensitivity of each list to hearing loss and distortions varied significantly. There was an increase in the minimum effect size that could be seen for syllabic scores compared to word scores, with no significant further improvement with phonemic scores. The use of at least two ten-word lists, quoted in syllables rather than in whole words, contributed to a large decrease in variability and an increase in sensitivity to hearing loss. However, those results emphasize the need of using updated linguistic material for clinical speech score assessments.

System-Level Design of an Integrated Receiver Front End for a Wireless Ultrasound Probe.

In this paper, a system-level design is presented for an integrated receive circuit for a wireless ultrasound probe, which includes analog front ends and beamformation modules. This paper focuses on the investigation of the effects of architectural design choices on the image quality. The point spread function is simulated in Field II from 10 to 160 mm using a convex array transducer. A noise analysis is performed, and the minimum signal-to-noise ratio (SNR) requirements are derived for the low-noise amplifiers (LNAs) and A/D converters (ADCs) to fulfill the design specifications of a dynamic range of 60 dB and a penetration depth of 160 mm in the B-mode image. Six front-end implementations are compared using Nyquist-rate and Σ∆ modulator ADCs. The image quality is evaluated as a function of the depth in terms of lateral full-width at half-maximum (FWHM) and -12-dB cystic resolution (CR). The designs that minimally satisfy the specifications are based on an 8-b 30-MSPS Nyquist converter and a single-bit third-order 240-MSPS Σ∆ modulator, with an SNR for the LNA in both cases equal to 64 dB. The mean lateral FWHM and CR are 2.4% and 7.1% lower for the Σ∆ architecture compared with the Nyquist-rate one. However, the results generally show minimal differences between equivalent architectures. Advantages and drawbacks are finally discussed for the two families of converters.