Lymphocytes In Bronchoalveolar Lavage Fluid Research Articles

Bronchoalveolar lavage fluid analysis in patients with checkpoint inhibitor pneumonitis

BackgroundCheckpoint inhibitor pneumonitis (CIP) is a relatively uncommon but potentially life-threatening immune-related adverse event (irAE). Lung biopsies have not been commonly performed for CIP patients. Bronchoalveolar lavage fluid (BALF) analysis is a useful diagnostic approach for interstitial lung disease. However, BALF features were inconsistent across different studies.MethodsWe retrospectively reviewed the medical records of 154 patients with pathologically confirmed malignancies and suffering from CIPs between July 2018 and December 2022. Patients who had bronchoalveolar lavage (BAL) data available were enrolled in our study. Patient clinical, laboratory, radiological and follow-up data were reviewed and analyzed.ResultsThe BALF differential cell count and lymphocyte subset analysis were performed for 42 CIP patients. There were 32 males (76.2%). The mean age at diagnosis of CIP was 62.0 ± 10.4 (range: 31–78) years. The median time to onset of CIP was 98.5 days after the start of immunotherapy. There were 18 patients (42.9%) with low-grade CIPs and 24 patients (57.1%) with high-grade CIPs. The mean lymphocyte percentage was 36.7 ± 22.5%. There were 34 (81%) CIP patients with a lymphocytic cellular pattern. The median ratio of CD3+CD4+/CD3+CD8+ lymphocytes was 0.5 (0.3, 1.0). The ratio was less than 1.0 for 31 CIP patients (73.8%). However, there was no significant difference in the BALF features between patients with low-grade CIPs and those with high-grade CIPs.ConclusionsThe CD3+CD8+ lymphocytosis pattern was the main inflammatory profile in the BALF of CIP patients in this cohort. Targeting CD3+CD8+ lymphocytes might be a treatment option for CIPs.

The application of bronchoscopy in the assessment of immune checkpoint inhibitor-related pneumonitis severity and recurrence

To explore the role of bronchoscopy for the assessment of checkpoint inhibitor pneumonitis (CIP), a retrospective single-center study was conducted to assess patients diagnosed with CIP at grade 2 or above and also underwent bronchoscopy between January 2020 and December 2022. Clinical data and bronchoscopic findings were recorded. The treatment data and prognosis information were collected. Twenty-one patients who underwent bronchoscopy and were diagnosed with CIP were enrolled in this study. All patients underwent bronchoalveolar lavage fluid (BALF) analysis. Of them, T lymphocyte subsets of BALF were tested in 15 cases. Transbronchial cryobiopsy (TBCB) was performed in 8 patients, and transbronchial lung biopsy was performed in 5 patients. 3 patients developed pneumothorax after TBCB and all recovered without serious compilations.14 patients experienced grade 2 CIP, while 7 patients ≥ grade 3 CIP. Symptoms were improved in 19 (90.5%) patients after standard treatment adhering to CIP guidelines. However, 5 patients relapsed during steroid tapering. Factors related to the severity and recurrence of CIP were analyzed. Patients with previous interstitial lung disease (ILD) were more likely to develop high grade CIP than those without [83.3% (5/6) versus 15.4% (3/15), P = 0.011].The odds ratio (OR) was 32.5 (95% CI 2.284–443.145, P = 0.009). Increased BALF lymphocyte percentage was associated with high grade CIP, OR 1.095 (95% CI 1.001–1.197, P = 0.047), and higher possibility of CIP relapse, OR 1.123 (95% CI, 1.005–1.225, P = 0.040). Lymphocyte subsets were tested in 15 patients. CD4/CD8 > 1 was found in 80% (4/5) of relapsed patients and 20% (2/10) of patients without relapse (P = 0.047). The OR was 16.00 (95% CI 1.093–234.24, P = 0.043). In this retrospective study, patients with previous ILD was more likely to develop high grade CIP. Higher lymphocyte percentage in BALF was associated with high grade CIP and susceptibility to relapse during treatment of CIP. A CD4/CD8 ratio greater than 1 in lymphocyte subsets of BALF was associated with higher possibility of CIP relapse. We found that TBCB is a safe procedure in CIP patients.

Bronchoalveolar lavage fluid analysis in patients with checkpoint inhibitor pneumonitis.

e14683 Background: Checkpoint inhibitor pneumonitis (CIP) is a relatively uncommon but potentially life-threatening immune-related adverse event (irAE). Lung biopsies have not been commonly performed for CIP patients. Bronchoalveolar lavage fluid (BALF) analysis is a useful diagnostic approach for interstitial lung disease. However, BALF features were inconsistent across different studies. Methods: We retrospectively reviewed the medical records of 154 patients with pathologically confirmed malignancies and suffering from CIPs between July 2018 and December 2022. Patients who had bronchoalveolar lavage (BAL) data available were enrolled in our study. Patient clinical, laboratory, radiological and follow-up data were reviewed and analyzed. In our study, patients with grade 1 or grade 2 CIPs were defined as low-grade CIPs, and patients with grade 3, grade 4 or grade 5 CIPs were defined as high-grade CIPs. Results: The BALF differential cell count and lymphocyte subset analysis were performed for 42 CIP patients. There were 32 males (76.2%). The mean age at diagnosis of CIP was 62.0 ± 10.4 (range: 31-78) years. The median time to onset of CIP was 98.5 days after the start of immunotherapy. Most of the patients were diagnosed with lung cancer (36 patients, 85.7%). There were 3 patients with esophageal squamous cell cancer, 1 patient with hepatic cell cancer, 1 patient with gastric signet ring cell cancer and 1 patient with pericardial malignant mesothelioma. There were 18 patients (42.9%) with low-grade CIPs and 24 patients (57.1%) with high-grade CIPs. The mean lymphocyte percentage was 36.7 ± 22.5%. There were 34 (81%) CIP patients with a lymphocytic cellular pattern. The median ratio of CD3+CD4+/CD3+CD8+ lymphocytes was 0.5 (0.3, 1.0). The ratio was less than 1.0 for 31 CIP patients (73.8%). However, there was no significant difference in the BALF features between patients with low-grade CIPs and those with high-grade CIPs. Conclusions: The CD3+CD8+ lymphocytosis pattern was the main inflammatory profile in the BALF of CIP patients in this cohort. Targeting CD3+CD8+ lymphocytes might be a treatment option for CIPs.

Clinical findings and outcome predictors for multinodular pulmonary fibrosis in horses: 46 cases (2009-2019).

Prognostic indicators for equine multinodular pulmonary fibrosis (EMPF), an interstitial fibrosing lung disease, are poorly described. Describe diagnostic findings and outcome predictors for EMPF. Forty-six adult horses with EMPF. Retrospective multicenter case series from 2009 to 2019. Radiographic (n = 27) and ultrasonographic studies (n = 19) from EMPF horses and bronchoalveolar lavage fluid (BALF) cytology from 6 EMPF and 13 asthma cases were independently reviewed and blinded to diagnosis and outcome. Associations between predictor variables and survival were assessed by predictor screening followed by Fisher's exact and Wilcoxon rank sum tests. Primary clinical findings were weight loss (36/46, 78%), increased respiratory effort (33/46, 72%), tachypnea (32/46, 70%), and fever (18/46, 39%). Macrophage atypia was seen in more EMPF than asthmatic horse BALF (67% vs. 8%; P = .02). Equine herpesvirus 5 (EHV-5) was detected in 24 of 30 (80%) and hyperfibrinogenemia in 25 of 28 (89%) cases. Twenty-seven of 46 horses (59%) and 11 of 45 (24%) survived to discharge and to 3 months, respectively. Three-month survival was associated with lower median (range) respiratory rates (30 [24-36] vs. 41 [30-60] breaths per minute; P = .04), and higher BALF lymphocyte:neutrophil ratios (4.7 [1.4-22] vs. 0.47 [0.11-1.9]; P = .01) and blood lymphocyte counts (1.25 [0.93-2.55] vs. 0.90 [0.70-1.24] × 109/L; P = .03). Imaging findings, EHV-5 detection, and corticosteroid treatment were not associated with survival. Fever is not a sensitive clinical sign of EMPF. Diagnostic testing should be pursued for horses with increased respiratory rate and effort and weight loss. The prognosis for EMPF horses is poor. Corticosteroid treatment does not improve 3-month survival.

Changes in Cellular Morphology in Bronchoalveolar Lavage Fluid of Children with Mycoplasma Pneumoniae Pneumonia

Objective To study changes of cell morphology in BALF in children with Mycoplasma pneumoniae pneumonia (MPP). Methods From December 2021 to May 2022, a group of 32 children diagnosed with Acute MPP and admitted for treatment in the Pediatrics Department and PICU of the First Affiliated Hospital of Xinxiang Medical University were selected for our study. These patients underwent bronchoalveolar lavage as part of their clinical assessment. For comparison, we included a control group comprising 10 children who were not infected but had bronchial foreign bodies. We investigated the cellular composition in the bronchoalveolar lavage fluid (BALF) using Wright-Giemsa staining and microscopic evaluation, aiming to understand the relationship between shifts in cell proportions and extra-pulmonary symptoms associated with MPP. Results In this study, a total of 42 cases were enrolled, with 32 cases in the study group and 10 cases in the control group. There were no statistically significant differences in gender, age, height, weight, and BMI between the two groups (p > 0.05). The study group exhibited significantly higher levels of neutrophil percentage (GRA%), CRP, D-dimer, and LDH in blood routine tests compared to the control group (p < 0.05). Furthermore, the proportions of neutrophils (%) and macrophages (%) in BALF were significantly higher in the study group compared to the control group (p< 0.05), while the proportion of lymphocytes (%) in BALF showed no statistically significant difference between the two groups (p> 0.05). Conclusion In the acute phase of MPP in children, BALF is predominantly composed of neutrophils. A lower proportion of lymphocytes in BALF is associated with a higher incidence of extra-pulmonary manifestations and longer hospitalization duration.

CTRP3 regulates NF-κB and TGFβ1/Smad3 pathways to alleviate airway inflammation and remodeling in asthmatic mice induced by OVA.

Asthma is a common illness with chronic airway inflammation. C1q/tumor necrosis factor (TNF)-related protein 3 (CTRP3) plays a vital role ininflammatory response, but its effect on asthma is imprecise. Herein, we analyzed the functions of CTRP3 in asthma. The BALB/c mice were randomized into four groups: control, ovalbumin (OVA), OVA+vector, and OVA+CTRP3. The asthmatic mice model was established by OVA stimulation. Overexpression of CTRP3 was implemented by the transfection of corresponding adeno-associated virus 6 (AAV6). The contents of CTRP3, E-cadherin, N-cadherin, smooth muscle alpha-actin (α-SMA), phosphorylated (p)-p65/p65, transforming growth factor-beta 1 (TGFβ1), and p-Smad3/Smad3 were determined by Western blot analysis. The quantity of total cells, eosinophils, neutrophils, and lymphocytes in bronchoalveolar lavage fluid (BALF) was assessed by using a hemocytometer. The contents of tumor necrosis factor-α and interleukin-1β in BALF were examined by enzyme-linked immunesorbent serologic assay. The lung function indicators and airway resistance (AWR) were measured. The bronchial and alveolar structures were evaluated by hematoxylin and eosin staining and sirius red staining. The CTRP3 was downregulated in mice of OVA groups; however, AAV6-CTRP3 treatment markedly upregulated the expression of CTRP3. Upregulation of CTRP3 diminished asthmatic airway inflammation by decreasing the number of inflammatory cells and the contents of proinflammatory factors. CTRP3 markedly lessened AWR and improved lung function in OVA-stimulated mice. Histological analysis found that CTRP3 alleviated OVA-induced airway remodeling in mice. Moreover, CTRP3 modulated NF-κB and TGFβ1/Smad3 pathways in OVA-stimulated mice. CTRP3 alleviated airway inflammation and remodeling in OVA-induced asthmatic mice via regulating NF-κB and TGFβ1/Smad3 pathways.

A Combination of Cytological Biomarkers as a Guide in the Diagnosis of Acute Rejection in Lung Transplant Recipients

The usefulness of bronchoalveolar lavage fluid (BALF) to support the diagnosis of acute cellular (ACR) rejection in lung transplant (LTX) recipients remains controversial. ACR has been associated with blood eosinophil counts (EOS) in other solid organ recipients, but there are few studies in relation to lung transplants. Our aim was to assess the usefulness of the combined analysis of BALF cellularity and EOS for the diagnosis of ACR in lung transplant recipients. This is a retrospective study of findings observed simultaneously in 887 transbronchial biopsies (TBB), BALF, and blood samples obtained from 363 LTx patients transplanted between 2014 and 2020. The variables collected were: demographics, ACR degree, BALF cellularity, and simultaneous blood EOS counts. The lymphocyte count in BALF was significantly higher in patients with ACR than in those without (11.35% vs. 6.11%; p < 0.001). In parallel, EOS counts were also significantly higher in patients with ACR than in the non-ACR group (EOS 213 ± 206/mm3 vs. 83 ± 129/mm3; p < 0.001). Increases in both parameters were associated with an increased risk of ACR (lymphocytes OR 1.100; 95% CI 1.080–1.131; EOS OR 1.460; 95% CI 1.350–1.580). The diagnostic specificity of ACR for a lymphocyte count > 12% was 71.1%, which increased to 95.8% when taking into account a simultaneous blood EOS count > 200/mm3. Simultaneous assessment of BALF lymphocyte counts and blood eosinophil counts may be useful for diagnosing ACR in patients with risk factors for TBB or in the presence of inconclusive histological samples.

Peripheral blood lymphopenia in sarcoidosis associates with HLA-DRB1 alleles but not with lung immune cells and organ involvement.

Different human leukocyte antigen (HLA) alleles associate with disease phenotypes in sarcoidosis. Peripheral blood (PB) lymphopenia is reported as more common in sarcoidosis patients with worse prognosis. The mechanisms behind are unrecognized but a PB depletion due to lymphocytes migrating to lung and/or extra pulmonary organs has been suggested. Insights into associations between HLA alleles, lung immune cells, clinical phenotype including extra pulmonary manifestations (EPM), and PB lymphopenia may provide mechanistic clues and enable adequate intervention in this patient group. In this situdy,141 treatment naïve, newly diagnosed patients were retrospectively identified in a Swedish cohort of sarcoidosis patients. Data on HLA-DRB1 alleles, lung immune cells from bronchoalveolar lavage fluid (BALF), PB lymphocytes and clinical parameters including treatment and disease course (chronic vs. resolving) were collected. The patients were followed for 2 years. PB lymphopenia associated with male sex, development of non-resolving disease, a need for first- and second-line systemic immunosuppressant treatment and HLA- DRB1*07. No correlation between BALF and PB lymphocytes, and no difference in EPM was detected between patients with and without PB lymphopenia. In conclusion, PB lymphopenia is associated with a more severe disease phenotype and carriage of the HLA-DRB1*07 allele. The results do not lend support to the hypothesis about sarcoidosis PB lymphopenia being due to a migration of PB lymphocytes to other organs. Rather, they provide a basis for future studies on the connection between HLA-DRB1*07 and PB lymphopenia mechanisms.

Cellular analysis and metagenomic next-generation sequencing of bronchoalveolar lavage fluid in the distinction between pulmonary non-infectious and infectious disease.

The aim of the current study was to investigate the clinical value of cellular analysis and metagenomic next-generation sequencing (mNGS) of bronchoalveolar lavage fluid (BALF) in differentiating pulmonary non-infectious and infectious diseases in immunocompetent patients. The present retrospective study was conducted from December 2017 to March 2020, and included immunocompetent patients with suspected pulmonary infection. High-resolution computed tomography, total cell counts and classification of BALF, conventional microbiological tests (CMTs), laboratory tests and mNGS of BALF were performed. Patients were assigned to pulmonary non-infectious disease (PNID) and pulmonary infectious disease (PID) groups based on final diagnoses. PNID-predictive values were analyzed via areas under receiver operating characteristic curves (AUCs). Optimal cutoffs were determined by maximizing the sum of sensitivity and specificity. A total of 102 patients suspected of pulmonary infection were enrolled in the study, 23 (22.5%) with PNID and 79 (77.5%) with PID. The diagnostic efficiency of BALF mNGS for differentiating PID from PNID was better than that of CMTs. Neutrophil percentage (N%) and the ratio of neutrophils to lymphocytes (N/L) in BALF were significantly lower in the PNID group than in the PID group. The AUCs for distinguishing PNID and PID were 0.739 (95% confidence interval [CI] 0.636-0.825) for BALF N%, 0.727 (95% CI 0.624-0.815) for BALF N/L, and 0.799 (95% CI 0.702-0.876) for BALF mNGS, with respective cutoff values of 6.7%, 0.255, and negative. Joint models of BALF mNGS combined with BALF N/L or BALF N% increased the respective AUCs to 0.872 (95% CI 0.786-0.933) and 0.871 (95% CI 0.784-0.932), which were significantly higher than those for BALF mNGS, BALF N%, and BALF N/L alone. BALF N% ≤ 6.7% or BALF N/L ≤ 0.255 combined with a negative BALF mNGS result can effectively distinguish PNID from PID in immunocompetent patients with suspected pulmonary infection. BALF mNGS outperforms CMTs for identifying pathogens in immunocompetent patients, and the combination of mNGS and CMTs may be a better diagnostic strategy.

The anti-asthmatic potential of Rho-kinase inhibitor hydroxyfasudil in the model of experimentally induced allergic airway inflammation

ObjectiveThis experimental study evaluated the anti-asthmatic potential of the Rho-kinase inhibitor hydroxyfasudil in the settings of allergen-induced allergen-induced experimental asthma. MethodsChronic allergic airway inflammation was caused by 28 days-sensitisation of guinea pigs with ovalbumin (OVA). Hydroxyfasudil was administered intraperitoneally in two doses for the last two weeks (1 mg/kg b.w.; 10 mg/kg b.w.). The degree of allergic inflammation was determined based on concentrations of inflammatory Th2 cytokines (IL-4, IL-13), Th1 cytokines (TNF-α and IFN-γ) in the lung homogenate and leukocyte count in the bronchoalveolar lavage fluid (BALF). The markers of remodelling and fibrosis, the growth factors (TGF-β1, EGF), EGF receptor, collagen type III and V were estimated in lung homogenate. The changes in specific airway resistance (sRaw) were used as an in vivo bronchial hyperreactivity parameter. ResultsHydroxyfasudil administration at both doses significantly reduced sRaw after a week of therapy. We observed a decline of IL-13, TNF-α and IFN-γ in lung homogenate and a lower presence of lymphocytes in BALF after 14 days of hydroxyfasudil administration at both tested doses. Hydroxyfasudil 14 days-treatment at both doses effectively reduced the concentrations of TGF-β1, EGF receptors, collagen type III and V in BALF and modulated EGF levels. ConclusionsThese findings indicate that RhoA/Rho-kinase is involved in the pathophysiology of allergic airway inflammation and suggest that Rho-kinase inhibitor hydroxyfasudil has therapeutic potential for asthma management.

Lung CCR6−CXCR3− type 2 helper T cells as an indicator of progressive fibrosing interstitial lung diseases

Progressive fibrosing interstitial lung diseases (PF-ILDs) have a poor prognosis and may be resistant to corticosteroids and/or immunosuppressants, but antifibrotic therapies such as nintedanib and pirfenidone have been shown to slow the deterioration of lung function. The aim of this study was to identify the characteristic cellular profile of bronchoalveolar lavage fluid at diagnostic bronchoscopy for predicting PF-ILDs, defined as fibrotic diseases on chest high-resolution computed tomography with more than a 5% relative decline in the percent predicted value of forced vital capacity (FVC) over 6 months. The proportions of inflammatory cells, CCR6−CXCR3− T helper type 2 (Th2) cells among conventional CD4+ T cells in bronchoalveolar lavage fluid (BALF) and peripheral blood, were measured by flowcytometry. The proportion of lymphocytes in BALF was significantly higher in non-PF-ILD patients than in PF-ILD patients. The proportion of Th2 cells in BALF, but not in peripheral blood, was significantly higher in PF-ILD patients than in non-PF-ILD patients. Multivariate analysis showed that a greater population of Th2 cells in BALF was the only indicator for PF-ILDs. An increased proportion of Th2 cells in BALF is associated with greater deterioration of lung function in fibrotic interstitial lung diseases.

The role of calcium-sensitive receptor in ovalbumin-induced airway inflammation and hyperresponsiveness in juvenile mice with asthma.

The role of the calcium-sensitive receptor (CaSR) was assessed in a juvenile mouse model of asthma induced by ovalbumin (OVA). The experiment was divided into normal control, OVA, and OVA +2.5/5mg/kg NPS2143 (a CaSR antagonist) groups. OVA induction was performed in all groups except the normal control, followed by assessing airway hyperresponsiveness (AHR) and lung pathological changes. Serum OVA-specific IgE and IgG1 were detected with an enzyme-linked immunosorbent assay (ELISA), and inflammatory cells were counted in bronchoalveolar lavage fluid (BALF). Real-time quantitative polymerase chain reaction, ELISA, and western blotting were performed to detect gene and protein expression. NPS2143 improved the OVA-induced AHR in mice, and AHR was higher in the OVA +2.5mg/kg NPS2143 group than in the OVA +5mg/kg NPS2143 group. Furthermore, NPS2143 reduced the production of OVA-specific IgE and IgG1 in serum and the number of eosinophils and lymphocytes in BALF in OVA mice with reduced CaSR expression in lung tissues. Besides, OVA-induced mice exhibited peribronchial and perivascular inflammatory cell infiltration, which was accompanied by severe goblet cell hyperplasia/hyperplasia and airway mucus hypersecretion. Furthermore, these mice exhibited increased levels of Interleukin (IL)-5, IL-13, MCP-1, and eotaxin, which were alleviated by NPS2143. The 5mg/kg NPS2143 showed more effective than the 2.5mg/kg treatment. CaSR expression was elevated in the lung tissues of OVA-induced asthmatic juvenile mice, whereas the CaSR antagonist NPS2143 reduced AHR and attenuated the inflammatory response in OVA-induced juvenile mice, possibly exerting therapeutic effects on childhood asthma.

Immune cell composition of the bronchoalveolar lavage fluid in healthy and respiratory diseased dromedary camels

BackgroundRespiratory diseases are among the most common and expensive to treat diseases in camels with a great economic impact on camel health, welfare, and production. Bronchoalveolar lavage fluid (BALF) has been proven as a valuable sample for investigating the leukocyte populations in the respiratory tract of several species. In the present study, fluorescent antibody labeling and flow cytometry were used to study the immune cell composition of BALF in dromedary camels. Animals with clinical respiratory diseases (n = seven) were compared with apparently healthy animals (n = 10). In addition, blood leukocytes from the same animals were stained in parallel with the same antibodies and analyzed by flow cytometry.ResultsCamel BALF macrophages, granulocytes, monocytes, and lymphocytes were identified based on their forward and side scatter properties. The expression pattern of the cell markers CD172a, CD14, CD163, and MHCII molecules on BALF cells indicates a similar phenotype for camel, bovine, and porcine BALF myeloid cells. The comparison between camels with respiratory disease and healthy camels regarding cellular composition in their BALF revealed a higher total cell count, a higher fraction of granulocytes, and a lower fraction of macrophages in diseased than healthy camels. Within the lymphocyte population, the percentages of helper T cells and B cells were also higher in diseased than healthy camels. The elevated expression of the activation marker CD11a on helper T cells of diseased camels is an indication of the expansion of helper T cells population due to infection and exposure to respiratory pathogens. The higher abundance of MHCII molecules on BALF macrophages from diseased camels indicates a polarization toward an inflammatory macrophage phenotype (M1) in respiratory diseased camels. No significant differences were observed in the systemic leukogram between healthy and diseased animals.ConclusionsCollectively, the current study represents the first report on flow cytometric analysis of immune cell composition of bronchoalveolar lavage fluid (BALF) in dromedary camels.

IL-9 and IL-9 receptor expression in lymphocytes from bronchoalveolar lavage fluid of patients with interstitial lung disease

IntroductionIL-9, mainly produced by T helper 9 (Th9) cells, promotes allergic airway inflammation and remodeling through the interaction with its receptor (IL-9R). Th9 cells and IL-9 have also been implicated in tissue fibrosis and autoimmunity pathways. However, the role of IL-9/IL-9R in the pathogenesis of interstitial lung disease (ILD) is unknown. AimTo evaluate IL-9/IL-9R expression in bronchoalveolar lavage fluid (BALF) lymphocytes of patients with various ILDs. MethodsConsecutive patients with ILD, who underwent BAL for diagnostic purposes, were studied. As control group, consecutive patients without evidence of ILD were included. Immunocytochemical staining of BALF lymphocytes for IL-9 and IL-9R was performed and evaluated by two independent readers. Results45 patients, of them 8 had idiopathic pulmonary fibrosis (IPF), 12 nonspecific interstitial pneumonia (NSIP), 10 sarcoidosis, 9 hypersensitivity pneumonitis (HP), 6 cryptogenic organizing pneumonia (COP), and 24 controls were studied. In the ILD group, the highest BALF lymphocyte count was seen in HP followed by NSIP, COP, sarcoidosis, and IPF (p < 0.05 for HP vs IPF). The highest percentages of IL-9 and IL-9R positive lymphocytes were seen in COP. Conversely, NSIP showed the lowest rate of IL-9, and sarcoidosis the lowest rate of IL-9R positive lymphocytes. Only in NSIP, a direct correlation between IL and 9 and IL-9R positive lymphocytes was seen (r = 0.639, p = 0.025). ConclusionBALF lymphocytes IL-9 and IL-9R expression differs between various ILDs and could reflect different pathogenetic mechanisms.

Supplementary Role of Immunological Indicators in the Diagnosis and Prognosis of Pneumocystis Pneumonia in Non-HIV Immunocompromised Patients.

BackgroundPneumocystis pneumonia (PCP) has a high mortality in HIV-negative immunocompromised patients. The occurrence and development of PCP are believed to be correlated with the level of lymphocytes and their subsets. The aim of this study was to determine if the levels of lymphocyte subpopulations and immunoglobulin are associated with PCP.MethodsA total of 74 immunocompromised patients were enrolled in this single-center cohort study. Diagnosis of PCP was based on the relevant pulmonary symptoms and radiological imaging, and the detection of Pneumocystis jirovecii in BAL fluid or biopsy tissue by metagenomic next-generation sequencing (mNGS). All patients were divided into two groups (PCP group and non-PCP group) and the patients in PCP group were then divided into two groupsbased on the outcome of the disease during the hospitalization.ResultsWe observed a significant lower level of IgG (p=0.000) and B lymphocyte counts (p=0.017) in the PCP group comparing to that in the non-PCP group. CD4+ T cell counts, as well as the ratio of CD4+/CD8+ T cells in circulation and BAL fluid were also lower in the PCP group comparing to those in the non-PCP group. Lactate dehydrogenase (LDH) in the PCP group was significantly higher than that in the non-PCP group (p=0.029). In the PCP group, a lower level of total lymphocytes (p=0.004), T cells (p=0.001), CD4+ cells (p=0.001), and CD8+ cells (p=0.007), as well as the proportion of lymphocytes in BAL fluid (p=0.000) were found in deceased patients comparing to those in the survived group.ConclusionOur study revealed an important role of humoral immunity in the infection of Pneumocystis jirovecii. The level of B cells and IgG could be used as a supplement to predict the occurrence of PCP. The level of CD4+ and CD8+ lymphocytes was significantly correlated with the outcome of PCP.

Prognostic implication of bronchoalveolar lavage fluid analysis in patients with Pneumocystis jirovecii pneumonia without human immunodeficiency virus infection

BackgroundThe prognostic value of bronchoalveolar lavage (BAL) fluid analysis in non-human immunodeficiency virus (HIV)-infected patients with Pneumocystis jirovecii pneumonia (PJP) has not been well elucidated. We aimed to investigate the prognostic implication of BAL fluid analysis in non-HIV patients with PJP.MethodsThe data of 178 non-HIV patients diagnosed with PJP based on the results of the polymerase chain reaction assay of BAL fluid specimens between April 2018 and December 2020 were retrospectively reviewed. The clinical characteristics, laboratory findings, and BAL fluid analysis results of patients who died within 90 days after hospital admission were compared.ResultsTwenty patients (11.2%) died within 90 days from admission. The neutrophil count in BAL fluid was significantly higher (median 22.0%, interquartile range [IQR] 2.0–46.0% vs. median 6.0%, IQR 2.0–18.0%, P = 0.044), while the lymphocyte count was significantly lower (median 24.0%, IQR 7.0–37.0% vs. median 41.0%, IQR 22.5–60.5%, P = 0.001) in the non-survivor group compared with that in the survivor group. In the multivariate analysis, the C-reactive protein level (odds ratio [OR] 1.093, 95% confidence interval [CI] 1.020–1.170, P = 0.011) and a BAL fluid lymphocyte count of ≤ 30% (OR 3.353, 95% CI 1.101–10.216, P = 0.033) were independently associated with mortality after adjusting for albumin and lactate dehydrogenase levels.ConclusionA low lymphocyte count in BAL fluid may be a predictor of mortality in non-HIV patients with PJP.

G9a promotes inflammation in Streptococcus pneumoniae induced pneumonia mice by stimulating M1 macrophage polarization and H3K9me2 methylation in FOXP1 promoter region

BackgroundStreptococcus pneumoniae has become a leading cause of pneumonia in recent years. Here, we investigated the mechanism of histone methylase G9a in Streptococcus pneumoniae-induced pneumonia (Spn).MethodsG9a expression in Spn mouse tissue was measured. G9a lentivirus interference vector was injected into Spn mice to evaluate the wet and dry weight of the right upper lobe and the total lung water content (TLW) and wet/dry ratio (W/D). The number of neutrophils, macrophages, and lymphocytes in bronchoalveolar lavage fluid (BALF) was detected, and the levels of interleukin-1β (IL-1β), IL-6, tumor necrosis factor-α (TNF-α), and IL-10 in BALF were assessed. The expressions of M1 and M2 macrophage markers were also detected. The enrichment of histone 3 lysine 9 dimethylation (H3K9me2) in the Forkhead Box P1 (FOXP1) promoter was detected by chromatin immunoprecipitation (ChIP) assay, and the transcription level of FOXP1 was detected. Mouse macrophage RAW264.7 was induced by lipopolysaccharide (LPS) following G9a interference.ResultsG9a in the lung tissue of Spn mice was increased. After G9a knockdown, the mouse weight increased, the infiltration of inflammatory cells was decreased, levels of pro-inflammatory cytokines in BALF were decreased, CD86 and inducible nitric oxide synthase (iNOS) were decreased, and CD206 and arginase-1 (Arg-1) were elevated. In LPS-induced RAW264.7, G9a inhibited macrophage polarization to M1 and promoted macrophage polarization to M2. G9a promoted H3K9me2 methylation in the FOXP1 promoter region and inhibited its transcription, while FOXP1 downregulation reversed the inhibition of G9a knockdown on macrophage polarization to M1 and the inflammatory effect on Spn mice.ConclusionsG9a promotes M1 polarization of macrophages by promoting H3K9me2 methylation in the FOXP1 promoter region, promoting an inflammatory response in Spn mice.

Protective effects of dexamethasone combined with valsartan on chronic obstructive pulmonary disease in mice and its mechanism

Objective: To investigate the possible protective effects of combined dexamethasone and valsartan against cigarette induced chronic obstructive pulmonary disease (COPD) in mice. Methods: Forty C57BL/6 mice were randomly divided into control group, COPD group, dexamethasone treated group, valsartan treated group and dexamethasone + valsartan combined treatment group, with 8 mice in each group. Mice in COPD group were exposed to cigarette for 8 weeks. On the basis of cigarette exposure, mice in dexamethasone treated group were intraperitoneally injected with dexamethasone (2 mg / kg) before cigarette exposure for 5-8 weeks. Mice in valsartan treated group were intraperitoneally injected with valsartan (30 mg/kg) before cigarette exposure for 1-8 weeks. Dexamethasone (2 mg/kg) and valsartan (30 mg/kg) were injected intraperitoneally into mice in the dexamethasone + valsartan combined treatment group. After 8 weeks, the lung tissues and bronchoalveolar lavage fluid (BALF) of mice in each group were collected. The pathological score of lung tissue was evaluated. The activities of superoxide dismutase(SOD) and matrix metalloproteinase-9 (MMP-9), and the contents of malondialdehyde (MDA), intracellular adhesion molecule-1 (ICAM-1), C-reactive protein (CRP) and nitric oxide (NO) in BALF were determined. Results: Compared with the control group, COPD mice had emphysema and alveolar congestion, the levels of MDA, ICAM-1, MMP-9, CRP and lymphocytes in BALF were increased, while the levels of SOD, macrophages and NO were decreased (all P＜0.05). Compared with COPD group, there was no significant improvement in emphysema and alveolar congestion, the levels of SOD and NO in BALF were increased, and the levels of MDA, lymphocytes and macrophages were decreased in dexamethasone or valsartan group (all P＜0.05). Compared with dexamethasone or valsartan group, the dexamethasone + valsartan combined treatment was more effective in preventing pulmonary emphysema and alveolar congestion caused by cigarette smoke. The levels of MDA, ICAM-1, MMP-9, CRP and lymphocyte in BALF were decreased, while the levels of SOD, macrophage and NO were increased (all P＜0.05). Conclusion: Compared with dexamethasone or valsartan, dexamethasone combined with valsartan has a more effective protective effect in COPD mice by inhibiting oxidative stress and inflammation.

Diagnostic and Staging Value of Serum Angiotensin-Converting Enzyme in Sarcoidosis.

Objective To investigate the diagnostic and staging value of serum angiotensin-converting enzyme (sACE) in sarcoidosis. Methods Patients with suspected sarcoidosis treated in the Department of Pulmonary and Critical Care Medicine of the China-Japan Friendship Hospital from 2010 to 2020 were included. The data of sACE, erythrocyte sedimentation rate (ESR), complete blood count (CBC), lung function, bronchoalveolar lavage, and biopsy were collected. The differences between the sarcoidosis group and the nonsarcoidosis group and between different stages of sarcoidosis were compared. The receiver operating characteristic (ROC) curve analysis was used for the diagnostic test of sACE in sarcoidosis. Results A total of 84 cases with suspected sarcoidosis were included, among which 70 cases were confirmed to be sarcoidosis by biopsy. The mean value of sACE in sarcoidosis patients was 56.61 ± 30.80 U/L, which was significantly higher than that in nonsarcoidosis patients (28.07 ± 14.11 U/L, P = 0.001). The level of sACE in sarcoidosis patients with peripheral superficial lymph nodes and multiple system involvement was significantly higher than that in intrathoracic sarcoidosis patients (P = 0.009); the percentage of lymphocytes in bronchoalveolar lavage fluid (BALF) of sarcoidosis patients was 45.39 ± 22.87%, which was significantly higher than that of nonsarcoidosis patients (P < 0.001). There was no correlation between sACE and ESR (correlation coefficient = −0.167). According to ROC curve analysis, when sACE ≥ 44.0 U/L, the sensitivity of sarcoidosis diagnosis was 61.4%, the specificity was 92.9%, and the AUC was 0.819. Conclusion sACE has a good specificity in the diagnosis of sarcoidosis. sACE values in patients with sarcoidosis with systemic involvement were higher than those with simple intrathoracic sarcoidosis.

Interventional effect of IL-17A on chronic obstructive pulmonary disease and its mechanism

Objective: To investigate the intervention effects and mechanism of interleukin-17A (IL-17A) on chronic obstructive pulmonary disease (COPD). Methods: C57BL/6 mice were randomly divided into wild type blank control group, wild type COPD group and IL-7A knockout COPD group. Mice in wild type blank control group received no treatment, and mice in the other two groups were exposed to cigarette smoke to induce COPD (Cigarette: 1 cigarette / time, 4 times/day, 45 minutes/time; interval time: 1 hour; total intervention time: 90 days). Lung function of mice was assessed using animal pulmonary function machine. Bronchoalveolar lavage fluid (BALF) of mice was collected and BALF cell count and classification were determined. The lung tissue of mice was collected, the expression level of IL-17A in airway epithelium was determined by flow cytometry, and the levels of inflammatory factors in lung tissue were determined by enzyme-linked immunosorbent assay. The expression level of JNK/AP1 signaling pathway protein in mouse lung tissue was determined by Western blot. Results: Compared with the wild type blank control group mice, the wild type COPD group mice had significantly higher expression level of IL-17A, significantly lower peak inspiratory flow rate (PIF) and peak expiratory flow rate (PEF), significantly higher number of BALF neutrophils, eosinophils, lymphocytes and macrophage, significantly higher expression levels of CXC chemokine 1(CXCL1), CXC chemokine 2 (CXCL2), interleukin-1β (IL-1β) and interleukin-6 (IL-6), and significantly higher phosphorylation level of JNK, cJun and cFos and AP1 expression levels (P＜0.05). Compared with COPD mice, IL-17A expression level in airway epithelium of mice in IL-7A knockout COPD group was significantly lower, PIF and PEF were higher, the number of BALF neutrophils, eosinophils, lymphocytes and macrophage was significantly lower, the expression levels of CXCL1, CXCL2, IL-1β and IL-6 in lung tissue were lower, and the phosphorylation levels of JNK, cJun and cFos and AP1 expression levels were significantly lower (P＜0.05). Conclusion: Cigarette smoke can induce the production of IL-17A and reduce (or inhibit) the production (or expression or secretion) of IL-17A in mouse airway epithelium, thus inhibiting the JNK/AP1 signaling pathway to reduce the airway inflammation and improve the lung function of COPD mice.