Mechanical ventilation and mortality among 223 critically ill patients with coronavirus disease 2019: A multicentric study in Germany.

BACKGROUND: There are large uncertainties with regard to the outcome of patients with coronavirus disease 2019 (COVID-19) and mechanical ventilation (MV). High mortality (50-97%) was proposed by some groups, leading to considerable uncertainties with regard to outcomes of critically ill patients with COVID-19. OBJECTIVES: The aim was to investigate the characteristics and outcomes of critically ill patients with COVID-19 requiring intensive care unit (ICU) admission and MV. METHODS: A multicentre retrospective observational cohort study at 15 hospitals in Hamburg, Germany, was performed. Critically ill adult patients with COVID-19 who completed their ICU stay between February and June 2020 were included. Patient demographics, severity of illness, and ICU course were retrospectively evaluated. RESULTS: A total of 223 critically ill patients with COVID-19 were included. The majority, 73% (n = 163), were men; the median age was 69 (interquartile range = 58-77.5) years, with 68% (n = 151) patients having at least one chronic medical condition. Their Sequential Organ Failure Assessment score was a median of 5 (3-9) points on admission. Overall, 167 (75%) patients needed MV. Noninvasive ventilation and high-flow nasal cannula were used in 31 (14%) and 26 (12%) patients, respectively. Subsequent MV, due to noninvasive ventilation/high-flow nasal cannula therapy failure, was necessary in 46 (81%) patients. Renal replacement therapy was initiated in 33% (n = 72) of patients, and owing to severe respiratory failure, extracorporeal membrane oxygenation was necessary in 9% (n = 20) of patients. Experimental antiviral therapy was used in 9% (n = 21) of patients. Complications during the ICU stay were as follows: septic shock (40%, n = 90), heart failure (8%, n = 17), and pulmonary embolism (6%, n = 14). The length of ICU stay was a median of 13 days (5-24), and the duration of MV was 15 days (8-25). The ICU mortality was 35% (n = 78) and 44% (n = 74) among mechanically ventilated patients. CONCLUSION: In this multicentre observational study of 223 critically ill patients with COVID-19, the survival to ICU discharge was 65%, and it was 56% among patients requiring MV. Patients showed high rate of septic complications during their ICU stay.

Dexamethasone and N-acetyl-cysteine attenuate Pseudomonas aeruginosa-induced mucus expression in human airways.

BACKGROUND: Infection with Pseudomonas aeruginosa (PA) induces mucus hypersecretion in airways. Therapeutic options to attenuate excessive mucus expression are sparse. OBJECTIVE: To investigate the effect of steroids and N-acetyl-cysteine (NAC) on PA-induced mucus expression. MATERIAL AND METHODS: Calu-3 cells and explanted human mucosa from the upper airways were stimulated with either PA, lipopolysaccharide from alginate producing PA (smooth, sPA-LPS) or non-alginate producing PA (rough, rPA-LPS). Dexamethasone (DEX) and NAC were added in different concentrations. Expression of mucin (MUC5AC) gene and mucin protein expression was quantified using PAS (periodic acids Schiff) staining and real time PCR. RESULTS: PA, sPA-LPS or rPA-LPS significantly induced mucin protein and MUC5AC gene expression in Calu-3 cells and explanted mucosal tissue (P < 0.05). Both DEX and NAC significantly decreased PA-, sPA-LPS- and rPA-LPS-induced mucin protein expression both in vitro and ex vivo (P < 0.05). A significant reduction was also observed for MUC5AC gene expression with the two agents (P < 0.05) except for sPA-LPS-induced mucin gene expression in vitro (P > 0.05). DISCUSSION AND CONCLUSION: Our data show that both an anti-inflammatory drug (DEX) and an anti-oxidative agent (NAC) can attenuate PA-induced mucus expression in human airways. These results support the use of steroids and NAC in clinical practice to treat PA-induced mucus hypersecretion.

Inhalation with fucose and galactose for treatment of Pseudomonas aeruginosa in cystic fibrosis patients.

BACKGROUND: Colonisation of cystic fibrosis (CF) lungs with Pseudomonas aeruginosa is facilitated by two lectins, which bind to the sugar coat of the surface lining epithelia and stop the cilia beating. OBJECTIVES: We hypothesized that P. aeruginosa lung infection should be cleared by inhalation of fucose and galactose, which compete for the sugar binding site of the two lectins and thus inhibit the binding of P. aeruginosa. METHODS: 11 adult CF patients with chronic infection with P. aeruginosa were treated twice daily with inhalation of a fucose/galactose solution for 21 days (4 patients only received inhalation, 7 patients received inhalation and intravenous antibiotics). Microbial counts of P. aeruginosa, lung function measurements, and inflammatory markers were determined before and after treatment. RESULTS: The sugar inhalation was well tolerated and no adverse side effects were observed. Inhalation alone as well as combined therapy (inhalation and antibiotics) significantly decreased P. aeruginosa in sputum (P < 0.05). Both therapies also significantly reduced TNFalpha expression in sputum and peripheral blood cells (P < 0.05). No change in lung function measurements was observed. CONCLUSIONS: Inhalation of simple sugars is a safe and effective measure to reduce the P. aeruginosa counts in CF patients. This may provide an alternative therapeutical approach to treat infection with P. aeruginosa.

Effect of dexamethasone and ACC on bacteria-induced mucin expression in human airway mucosa.

Gram-negative bacteria can stimulate mucin production, but excessive mucus supports bacterial infection and consequently leads to airway obstruction. Therefore, the effect of dexamethasone (DEX) and the antioxidant acetyl-cysteine (ACC) on bacteria-induced mucus expression was investigated. Explanted human airway mucosa and mucoepidermoid cells (Calu-3) were stimulated with lipopolysaccharide (LPS) or PAM3 (a synthetic lipoprotein). DEX or ACC were added to either LPS- or PAM3-stimulated airway mucosa or Calu-3 cells. Mucin mRNA expression (MUC5AC) and total mucus glycoconjugates (mucin protein) were quantified using real-time PCR and periodic acid Schiff staining. LPS and PAM3 significantly increased mucin expression in airway mucosa and Calu-3 cells (P < 0.05). DEX alone had no significant effect on mucin expression in airway mucosa or Calu-3 cells (P > 0.05). In contrast, DEX significantly reduced LPS- and PAM3-induced mucin expression in explanted mucosal tissue and mucin expression in Calu-3 cells (P < 0.05). In explanted human airway mucosa ACC alone significantly increased mucin expression (P < 0.05). In contrast, ACC significantly decreased LPS- and PAM3-induced mucin expression (P < 0.05). In Calu-3 cells ACC alone had no significant effect on mucin expression (P > 0.05). ACC decreased LPS- and PAM3-induced mucin expression, but this effect was not significant (P > 0.05). These data suggest that DEX can effectively reduce bacteria-induced mucin expression in the airways. ACC alone may increase mucin expression in noninfected mucosa, but it decreased bacteria-induced mucin expression. Further studies are warranted to evaluate whether the effect of DEX or ACC is clinically relevant.

LPS-induced mucin expression in human sinus mucosa can be attenuated by hCLCA inhibitors.

BACKGROUND: hCLCA1 is a member of the calcium-activated chloride channel family and is associated with disease-inducible mucus expression. Niflumic acid (NFA) and a closely related chemical structure are reported inhibitors of calcium-activated chloride channels and endotoxin-inducible mucus expression in the mouse. Therefore, we tested the hypothesis that hCLCA1 may be involved in lipopolysaccharide (LPS) induced mucin up-regulation in human airways. We also investigated the effect of NFA and MSI-2216 on LPS-induced mucin up-regulation. MATERIALS AND METHODS: Explanted human airways and the muco-epidermoid cell line Calu-3 were stimulated with LPS. Different concentrations of NFA or MSI-2216 were added to LPS-stimulated airway mucosa and Calu-3 cells. Expression of hCLCA1 and MUC5AC mRNA and protein was quantified in human airways using real-time PCR and PAS staining. In addition, immunohistochemistry was performed for quantification of inflammatory cells (lymphocytes, monocytes, eosinophils, and neutrophils) in the submucosa of the airways. Expression of hCLCA1 protein in Calu-3 cells was analysed by FACS. RESULTS: LPS significantly induced hCLCA1 and MUC5AC mRNA and protein expression in human airway mucosa (P < 0.05). NFA and MSI-2216 significantly decreased LPS-induced mucus expression in explanted airway mucosa in a dose-dependent manner (P < 0.05). In Calu-3 cells, LPS significantly increased hCLCA1 surface expression whereas intracellular expression was significantly decreased (P < 0.05). In Calu-3 cells, NFA and MSI-2216 also significantly reduced MUC5AC mRNA expression (P < 0.05). CONCLUSIONS: These data suggest that hCLCA1 may play a role in LPS-induced mucin expression in human airway mucosa. Calcium-activated chloride channel inhibitors significantly decreased LPS-induced mucus expression both ex vivo and in vitro . Therefore, blocking of hCLCA1 may offer a therapeutic approach to reduce bacterial-induced mucus hypersecretion.

Up-Regulation of Interleukin-9 and the Interleukin-9-Associated Calcium-Activated Chloride Channel hCLCA1 in Nasal Mucosa Following In Vivo Allergen Challenge.

: Interleukin (IL)-9 is a pleiotropic T helper 2-type cytokine that has been shown to be up-regulated in allergic airway disease, including asthma. IL-9 has been demonstrated to be a potent stimulus for the production and secretion of mucus from airway epithelial cells via induction of a calcium-activated chloride channel, hCLCA1. The objective of this study was to investigate the expression of IL-9 and hCLCA1 following allergen challenge in the nasal mucosa of allergic rhinitis patients. Nasal biopsies were obtained from allergic rhinitis patients out of allergen season both before (baseline) and after local antigen challenge with either ragweed or diluent (control). Immunohistochemistry and in situ hybridization were used to assess IL-9 protein and hCLCA1 messenger ribonucleic acid. Eosinophils and T cells were detected using immunohistochemistry. IL-9 and hCLCA1 were very low at baseline, and expression was significantly up-regulated following ragweed challenge. Whereas the number of eosinophils increased after allergen challenge, T-cell counts did not change significantly. The results of this study demonstrate the relationship between specific allergen challenge and expression of both IL-9 and hCLCA1, suggesting a possible mechanism for the increased production of mucus from airway epithelial cells in allergic rhinitis.

Mucin overproduction in chronic inflammatory lung disease.

Mucus overproduction and hypersecretion are commonly observed in chronic inflammatory lung disease. Mucins are gel-forming glycoproteins that can be stimulated by a variety of mediators. The present review addresses the mechanisms involved in the upregulation of secreted mucins. Mucin induction by neutrophil elastase, bacteria, cytokines, growth factors, smoke and cystic fibrosis transmembrane conductance regulator malfunction are also discussed.

Toll-like receptors 4 and 2 expression in the bronchial mucosa of patients with cystic fibrosis.

BACKGROUND: Cystic fibrosis (CF) is a lung disease characterized by chronic infection with Gram-negative bacteria Pseudomonas aeruginosa and Gram-positive bacteria Staphylococcus aureus. Recently, toll-like receptor (TLR) 4 has been shown to be responsible for the lipopolysaccharide (LPS)-mediated immune response. While TLR2 mediates responses driven by bacterial lipoproteins and peptidoglycans from Gram-positive bacteria, LPS derived from P aeruginosa may stimulate the immune response in the airways of patients with CF via activation of TLR4. OBJECTIVES: To investigate TLR4 and TLR2 expression in the bronchial mucosa of patients with CF and normal control subjects. PATIENTS AND METHODS: Endoscopic bronchial biopsies from seven patients with CF and six healthy control subjects were obtained. TLR4 and TLR2 expression was assessed using immmunocytochemistry. Real-time polymerase chain reaction was used to detect TLR4 messenger RNA in blood cells from patients with CF and to compare TLR4 expression in CF bronchial epithelial cells with non-CF bronchial epithelial cells. RESULTS: In patients with CF, the number of TLR4-positive cells was significantly increased in their submucosa (P<0.05) but significantly reduced in their epithelium compared with control subjects (P<0.05). The majority of TLR4-positive cells were neutrophils. Patients with CF (n=4) and control subjects (n=4) had a similar percentage of TLR4-expressing neutrophils and monocytes/lymphocytes in peripheral blood. CF cells (IB-3) had significantly decreased basal TLR4 messenger RNA expression compared with non-CF cells (Calu-3) (P<0.05). Although there was a trend toward reduced TLR2 expression in the airway epithelium of patients with CF (P=0.07), there was no significant difference in TLR2 expression in the submucosa of patients with CF compared with that of control subjects. CONCLUSIONS: Both TLR4 and TLR2 expression in the bronchial epithelium of patients with CF were significantly reduced compared with healthy control subjects. In contrast, the number of TLR4-positive neutrophils in the submucosa of patients with CF was higher than in control subjects. This may suggest that the loss of epithelial TLR expression may contribute to the impaired defense against LPS.

Increased expression of the calcium-activated chloride channel hCLCA1 in airways of patients with obstructive chronic bronchitis.

BACKGROUND: Interleukin (IL)-9 and its effect on enhancing the human calcium-activated chloride channel 1 (hCLCA1) expression have been shown to induce mucin production. Increased expression of hCLCA1 may, in turn, contribute to mucus overproduction in chronic obstructive pulmonary disease (COPD) with a chronic bronchitis (CB) phenotype. OBJECTIVE: To determine the expression of IL-9, IL-9 receptor (IL-9R), hCLCA1 and mucoglycoconjugates in COPD. METHODS: Bronchial biopsies were obtained from six patients with obstructive CB and six healthy control subjects. IL-9, IL-9R and hCLCA1 expression were detected using immunohistochemistry. Additionally, in situ hybridization was performed to determine the expression of hCLCA1 messenger RNA. Mucin production was assessed using periodic acid-Schiff staining. RESULTS: There was a significantly higher number of IL-9 immunoreactive cells in the submucosa of patients with COPD than that of healthy control subjects (P<0.05). Also, a significant increase in the expression of IL-9R, hCLCA1 (protein and messenger RNA) and mucin (periodic acid-Schiff-positive cells) was noted in the bronchial epithelium of patients with COPD compared the control subjects (P<0.05). CONCLUSION: Increased expression of IL-9, IL-9R and hCLCA1 in the bronchial mucosa of patients with obstructive CB suggests that mucus overproduction in this disease may be, at least in part, due to hCLCA1.

Decreased interleukin-18 expression in BAL cells and peripheral blood mononuclear cells in adult cystic fibrosis patients.

Lymphocytes from cystic fibrosis (CF) patients secrete less interferon-gamma (IFN-gamma) upon stimulation compared to controls. Expression of interleukin (IL)-18 as an IFN-gamma inducing factor and of IL-10 as an IL-18 inhibiting factor were determined in bronchoalveolar lavage (BAL) cells from CF patients (n=5) and from normal control subjects (n=9) as well as in peripheral blood mononuclear cells (PBMC) from patients (n=12) and from control subjects (n=9) with RT-PCR. IL-18 and IL-10 serum protein levels were measured using ELISA. BAL cells and PBMC of CF patients expressed significantly less IL-18 compared to controls (p<0.05). There was no significant difference for IL-10 in BAL cells. However, PBMC from patients expressed significantly more IL-10 mRNA (p<0.05). IL-18 serum protein levels were decreased in the patient group, whereas IL-10 serum concentrations were elevated. Stimulation with rhIL-10 reduced IL-18 expression in PBMC from CF patients. Decreased IL-18 expression in CF patients may contribute to decreased IFN-gamma production. IL-10 may contribute to inhibit IL-18 expression in PBMC in CF.

Increased expression of Interleukin-13 but not Interleukin-4 in cystic fibrosis patients.

BACKGROUND: Many patients with cystic fibrosis (CF) suffer from allergic disease, which can complicate treatment of CF lung disease. Interleukin (IL)-4 and IL-13 have been shown to be important mediators in allergic disease. OBJECTIVE: To investigate the role of IL-4 and IL-13 in allergic and non-allergic CF patients. METHODS: Expression of IL-4 and IL-13 mRNA was investigated in peripheral blood mononuclear cells (PBM) of seven CF patients with allergy, of six patients without allergy and of nine healthy subjects as well as in BAL cells of four patients and of all controls. PBM from six patients were incubated with recombinant human IL-13 or human antiIL-13 antibody without and with LPS stimulation and TNFalpha levels were measured by ELISA. RESULTS: IL-13 mRNA expression was increased in allergic and non-allergic patients compared to controls. No significant difference in IL-4 expression could be found between patients and controls. Addition of IL-13 decreased TNFalpha in PBM culture supernatants. CONCLUSION: Our data suggest that IL-13 rather than IL-4 might play an important role in both allergic and non-allergic CF patients. IL-13 might also compromise host defence by decreasing TNFalpha production.

Increased expression of interleukin-9, interleukin-9 receptor, and the calcium-activated chloride channel hCLCA1 in the upper airways of patients with cystic fibrosis.

OBJECTIVES/HYPOTHESIS: Mucus overproduction is commonly found in airway disease in patients with cystic fibrosis. Interleukin-9 (IL-9) has been shown to mediate airway hyper-responsiveness and mucus overproduction. Recently, the calcium-activated chloride channel hCLCA1 has been described to be upregulated by IL-9 and has been thought to regulate the expression of soluble gel-forming mucins. We sought to examine the expression of IL-9, interleukin-9 receptor (IL-9R), and hCLCA1 in the upper airway of patients with cystic fibrosis in comparison to healthy control subjects and to demonstrate the relationship of IL-9, IL-9R, and hCLCA1 expression with mucus production. STUDY DESIGN: Prospective design. METHODS: Biopsy samples from nasal polyps of four patients with cystic fibrosis, nasal mucosa of six patients with cystic fibrosis, sinus mucosa of eight patients with cystic fibrosis, and nasal mucosa of six healthy control subjects were stained with periodic acid-Schiff (PAS) to identify mucus glycoconjugates. IL-9, IL-9R, and hCLCA1 expression was determined by immunocytochemical study. RESULTS: We demonstrated significant increases in IL-9, IL-9R, and hCLCA1 immunoreactivity in the mucosa of patients with cystic fibrosis compared with that found in control subjects (P <.05). There were no significant differences between the different locations (nasal polyps, nasal mucosa, and sinus mucosa) in the patient group (P >.05). We also observed a significant increase in the number of mucus-producing cells in biopsy specimens from patients with cystic fibrosis in comparison to control subjects. A positive correlation was found between hCLCA1-positive cells and IL-9-positive cells (correlation coefficient [r] = 0.79, P <.05) or IL-9R-positive cells (r = 0.92, P <.05). Moreover, a positive correlation was also present between PAS-positive (mucus-producing) cells and hCLCA1-positive cells (r = 0.64, P <.05) or IL-9R-positive cells (r = 0.64, P <.05). CONCLUSIONS: Increased expression of IL-9 and IL-9R, as well as upregulation of hCLCA1, in mucus-overproducing epithelium of patients with cystic fibrosis supports the hypothesis that IL-9 contributes to mucus overproduction in cystic fibrosis. Expression of hCLCA1 may also be responsible, in part, for the overproduction of mucus. These preliminary findings suggest that hCLCA1 might be an interesting new therapeutic target to control mucus overproduction in airway disease in patients with cystic fibrosis.

Cytokine profile of chronic sinusitis in patients with cystic fibrosis.

BACKGROUND: The inflammatory-cell and cytokine profiles of chronic sinusitis (CS) are well documented in the literature. In contrast, little is known about the pathogenesis of this condition in patients with cystic fibrosis (CF). OBJECTIVE: To determine whether patients with CF have inflammatory-cell and cytokine profiles that differ from other patients with CS. METHODS: Patients with CF (n = 7) and adults with CS (n = 7) undergoing functional endoscopic sinus surgery were recruited for the study. Patients with no allergies or sinus disease (n = 6) were used as controls. Using immunohistochemical analysis, we assessed sinus mucosal specimens for the presence of T lymphocytes, eosinophils, macrophages, and neutrophils. Using in situ hybridization, we assessed the expression of interleukin (IL) 4, IL-5, IL-8, IL-10, and interferon gamma. RESULTS: There was a higher number of neutrophils, macrophages, and cells expressing messenger RNA for interferon gamma and IL-8 in patients with CF than in patients with CS or in controls (P<.01). The number of eosinophils and cells expressing messenger RNA for IL-4, IL-5, and IL-10 was higher in patients with CS than in those with CF and controls (P<.01). CONCLUSIONS: Sinus disease in patients with CF presents different inflammatory-cell and cytokine profiles than that seen in other patients with CS. These results may explain the difference in response to treatment in the CF group.

Pharyngeal lavage lymphocytosis in patients with obstructive sleep apnea: a preliminary observation.

BACKGROUND: Upper airway inflammation has been previously demonstrated in obstructive sleep apnea (OSA). However, investigation has been hampered by the necessity of invasive tissue biopsies. OBJECTIVE: To evaluate the pharyngeal lavage (PHAL) as a new tool to analyze mucosal inflammation in the pharynx of patients with sleep-related disordered breathing. PATIENTS AND METHODS: 36 patients with a diagnosis of OSA, 14 patients with heavy snorer syndrome (HS) or body position dependent OSA (bd-OSA), and 14 healthy volunteers underwent PHAL. Inflammatory cell counts were compared. RESULTS: Neutrophils were the predominant cells in PHAL in all groups (94.3% ± 0.7%, 98.5% ± 0.6%, 94.3% ± 0.7%, and 96.2% ± 1.4%). OSA patients had significantly increased numbers of lymphocytes (3.2% ± 0.4%) compared to bd-OSA/HS and controls group (0.5% ± 0.1% and 0.6% ± 0.2%, respectively; P<0.05). Patients with moderate to severe OSA had significantly higher numbers of lymphocytes compared to patients with mild OSA (P<0.05). CONCLUSIONS: Data from this study suggest that PHAL is a feasible tool to investigate upper airway inflammation in OSA. In addition, PHAL demonstrates lymphocytic inflammation of the pharynx in OSA patients. Future studies are warranted to evaluate whether PHAL can be used to monitor disease and whether lymphocytic inflammation is affected by OSA treatment.

HOPE-BAL: improved molecular diagnostics by application of a novel technique for fixation and paraffin embedding.

The bronchoalveolar lavage (BAL) and its cells have been widely used as a support for clinical diagnosis and as a versatile tool for research questions since many years. Because there are no sufficient possibilities of long-term storage, the authors explore in this study the utility of a new fixative for fixation and paraffin embedding of human lavage cells with the possibility of implementing standard molecular biology techniques. HOPE-fixed, paraffin-embedded BAL cells of patients with different lung diseases (asthma, chronic obstructive pulmonary diseases, tuberculosis, sarcoidosis, emphysema, and fibrosis) were subjected to immunohistochemistry, in situ hybridization, quantitative polymerase chain reaction, and transcription microarray analysis. Furthermore, two-dimensional gel electrophoresis was conducted to evaluate the range of possible applications for research, diagnostics, and further implementing in biobanks. The authors show, by targeting some exemplary molecules, the power of screening and validating HOPE-BAL for new biomarkers. The transforming growth factor ß signaling pathway may play a central role in immunomodulation upon infection as well as asthma. Furthermore, haptoglobin was overexpressed in asthma and sarcoidosis. Because of the excellent preservation of nucleic acids, protein, and morphologic structures, HOPE-BAL is a step forward into enhanced molecular diagnostics and biobanking in pulmonary medicine.

Current and future treatment options in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic condition of unknown aetiology with deteriorating respiratory function leading to respiratory failure. Sequential acute lung injury leads to progressive fixed tissue fibrosis, architectural distortion and loss of function. An excess of profibrotic cytokines and/or a deficiency in antifibrotic cytokines have been implicated in the pathological process as has excessive oxidation. At present no specific therapy is available. Corticosteroids alone or in combination with immunosuppressive drugs such as azathioprine, colchicine, and cyclophosphamide have been used with limited success. Interferon-gamma-1b showed a significant improvement in pulmonary function only in one study. Pirfenidone, cyclosporine and acetylcysteine may also prove to be of benefit but data from studies are limited. Novel drugs, mainly antifibrotic, anticytokine and immunoregulatory, are currently being investigated in various trial phases. Most recently, endothelin receptor antagonists (eg. bosentan) have been shown to have possible beneficial effects in early stages of IPF. After a short overview on the current hypothesis on pathophysiology in IPF this review will discuss the present and possible future therapeutic options in IPF.

The other T helper cells in asthma pathogenesis.

The complex phenotype of allergic bronchial asthma involves a variable degree of bronchoobstruction, increased mucus production, and airway remodeling. So far it is suggested that it arises from multiple interactions of infiltrating and structural cells in the context of chronic airway inflammation that is orchestrated by T helper 2 (TH2) cells. By secreting a plethora of typical mediators such as interleukin (IL) 4, IL-5, and IL-13, these cells hold a key position in asthma pathogenesis. However, therapeutic approaches targeting these TH2-type mediators failed to improve asthma symptoms and impressively showed that asthma pathogenesis cannot be reduced by TH2 cell functions. Recently, other T helper cells, that is, TH9 and TH17 cells, have been identified and these cells also contribute to asthma pathogenesis, the processes leading to formation or aggravation of asthma. Furthermore, TH25 cells, TH3 cells, and regulatory T cells have also been implicated in asthma pathogenesis. This paper aims at summarizing recent insights about these new T helper cells in asthma pathogenesis.

Effect of Th2 type cytokines on hCLCA1 and mucus expression in cystic fibrosis airways.

Correlations between expression of interleukin (IL)-9, the calcium-activated chloride channel hCLCA1 and mucus expression in cystic fibrosis (CF) airways have suggested a causal relationship. To verify this hypothesis mucosal tissue from upper airways of CF patients (N=5) was stimulated with the Th2 type cytokines IL-4, IL-9, or IL-13. Expression of hCLCA1 mRNA and protein as well as mucus and mucin (MUC5AC) gene expression was quantified using real time PCR, immunohistochemistry (hCLCA1) and PAS staining (mucus). Th2 type cytokines significantly increased hCLCA1 protein expression (P<0.05) whereas increase in hCLCA1 mRNA expression failed to reach statistical significance (P>0.05). Mucin protein and MUC5AC mRNA expression were not significantly changed (P>0.05). These data suggest that Th2 type cytokines may increase hCLCA1 expression in CF but may not have a significant effect on mucus expression. Therefore the role of hCLCA1 as a mediator of mucus overexpression in CF has to be questioned.

Monoclonal antibody S60-4-14 reveals diagnostic potential in the identification of Pseudomonas aeruginosa in lung tissues of cystic fibrosis patients.

The lipopolysaccharide (LPS) of Pseudomonas aeruginosa has been identified to contain an inner-core structure expressing a Pseudomonas-specific epitope. This target structure is characterized by a highly phosphorylated and 7-O-carbamoyl-l-glycero-alpha-d-manno-heptopyranose (CmHep) and was found to be present in all human-pathogenic Pseudomonas species of the Palleroni (RNA)-classification I scheme. We raised and selected the monoclonal antibody S60-4-14 (mAb S60-4-14, subtype IgG1) from mice immunized with heat-killed Pseudomonas bacteria. The epitope of this mAb was found to reside in the inner-core structure of P. aeruginosa and, hence, successfully evaluated for the immunohistochemical detection of P. aeruginosa in formalin- or HOPE-fixed (Hepes-glutamic acid buffer-mediated organic solvent protection effect) and paraffin-embedded human lung tissue slices. Lung specimens, mainly from explanted lungs of cystic fibrosis (CF) patients, as well as P. aeruginosa isolates from patients suffering from CF and patients with extrapulmonar Pseudomonas infections were investigated by PCR, immunohistochemistry, and Western blot analysis with mAb S60-4-14. The results revealed an unequivocal coincidence of PCR and immunohistochemistry. Together with the Western blot results mAb S60-4-14 displays a potential diagnostic tool for the specific identification of P. aeruginosa in infected lungs of CF.