Osimertinib, a third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, effectively treats EGFR-mutated non-small cell lung cancer. Drug-induced interstitial lung disease, a potentially fatal complication of osimertinib, involves an increase in lymphocytes in the bronchoalveolar lavage fluid (BALF). As the precise role of these lymphocytes is unknown, we investigated the pathogenesis of osimertinib-induced pneumonitis using BALF obtained from patients, and an osimertinib-induced pneumonitis mouse model. Mass cytometry revealed the presence of CCR7+ CD45RA+ naïve T cells in the BALF of patients with osimertinib-induced pneumonitis. Body weight measurements, BALF analysis, histopathological evaluation and RNA sequencing of mouse lung tissue were performed to investigate immune cell involvement and the mechanisms underlying osimertinib-induced pneumonitis. In the mouse model, administration of osimertinib after naphthalene-induced damage to the bronchiolar epithelium exacerbated lung inflammation and resulted in significant weight loss. Immunofluorescence staining revealed the infiltration of CCR7+ cells into the lungs of mice treated with naphthalene and osimertinib. Bulk RNA sequencing identified an upregulation of chemokine-related biological processes, with increased expression of C–C motif chemokine ligand 21 (Ccl21) and C–C motif chemokine ligand 8 (Ccl8) in the lung tissue. Additionally, immunofluorescence staining confirmed elevated expression of Ccl21 and Ccl8 in the distal bronchiolar epithelium. This study provides insights into the mechanisms underlying osimertinib-induced pneumonitis.

Mucin overproduction is a common feature of chronic airway diseases such as asthma and chronic obstructive pulmonary disease (COPD), and exacerbates their underlying respiratory condition. Surfactant protein D (SP-D) protects against airway diseases through modulation of immune reactions, but whether it also exerts direct effects on airway epithelial cells has remained unclear. Therefore, we sought to investigate the inhibitory role of SP-D on mucin production in airway epithelial cells. We prepared air–liquid interface (ALI) cultures of human primary bronchial epithelial cells (HBECs), which recapitulated a well-differentiated human airway epithelium. Benzo(a)pyrene (BaP), a key toxicant in cigarette smoke, induced mucin 5AC (MUC5AC) production in ALI-cultured HBECs, airway secretory cell lines, and airway epithelia of mice. Then, the protective effects of SP-D against the BaP-induced mucin overproduction were examined. BaP increased MUC5AC production in ALI cultures of HBECs, and this effect was attenuated by SP-D. SP-D also suppressed the BaP-induced phosphorylation of extracellular signal-regulated kinase (ERK) and MUC5AC expression in NCI-H292 goblet-like cells, but not in NCI-H441 club-like cells. Signal regulatory protein α (SIRPα) was found to be expressed in HBECs and NCI-H292 cells but absent in NCI-H441 cells. In NCI-H292 cells, SP-D activated SH2 domain-containing tyrosine phosphatase-1 (SHP-1), downstream of SIRPα, and knockdown of SIRPα abolished the suppressive effects of SP-D on BaP-induced ERK phosphorylation and MUC5AC production. Consistent with these in vitro findings, intratracheal instillation of SP-D prevented the BaP-induced phosphorylation of ERK and Muc5ac expression in airway epithelial cells in a mouse model. SP-D acts directly on airway epithelial cells to inhibit mucin secretion through ligation of SIRPα and SHP-1-mediated dephosphorylation of ERK. Targeting of SIRPα is therefore a potential new therapeutic approach to suppression of mucin hypersecretion in chronic airway diseases such as COPD and asthma.

Immune cells have been implicated in interstitial lung diseases (ILDs), although their phenotypes and effector mechanisms remain poorly understood. To better understand these cells, we conducted an exploratory mass cytometry analysis of immune cell subsets in bronchoalveolar lavage fluid (BALF) from patients with idiopathic pulmonary fibrosis (IPF), connective-tissue disease (CTD)-related ILD, and sarcoidosis, using two panels including 64 markers. Among myeloid cells, we observed the expansion of CD14+ CD36hi CD84hiCCR2- monocyte populations in IPF. These CD14+ CD36hi CD84hi CCR2- subsets were also increased in ILDs with a progressive phenotype, particularly in a case of acute exacerbation (AEx) of IPF. Analysis of B cells revealed the presence of cells at various stages of differentiation in BALF, with a higher percentage of IgG memory B cells in CTD-ILDs and a trend toward more FCRL5+ B cells. These FCRL5+ B cells were also present in the patient with AEx-IPF and sarcoidosis with advanced lung lesions. Among T cells, we found increased levels of IL-2R+ TIGIT+ LAG3+ CD4+ T cells in IPF, increased levels of CXCR3+ CD226+ CD4+ T cells in sarcoidosis, and increased levels of PD1+ TIGIT+ CD57+ CD8+ T cells in CTD-ILDs. Together, these findings underscore the diverse immunopathogenesis of ILDs.

Bronchiolitis obliterans syndrome (BOS) is a major complication of haematopoietic stem cell transplantation (HSCT), which affects the lungs and has limited treatment options [1]. BOS after HSCT is considered a manifestation of chronic graft-versus-host disease (GVHD). T-cells play a crucial role in its onset [2], while the contribution of myeloid cells has been largely unknown. This case study of a patient with BOS after HSCT found increased ST2+CD64+ macrophages in BALF, a potential therapeutic target for treatment-refractory BOS, and reduced CCR2+CD14+ monocytes compared to other lung disorders https://bit.ly/406Uyy9

Mass cytometry of BALF cells from a pulmonary alveolar proteinosis patient, positive for anti-GM-CSF antibodies, suggests potential impairment in human alveolar macrophage differentiation https://bit.ly/45JHUrz.

Background and Aims: Lymphangioleiomyomatosis (LAM) is a rare destructive lung disease characterized by multiple thin-walled pulmonary cysts.The currently proposed diagnostic algorithm emphasizes the characteristic cystic appearance on high-resolution computed tomography (HRCT) so atypical HRCT appearances present challenges to establishing the proper LAM diagnosis.The objective of this study is to accrue atypical chest HRCT appearances, determine frequencies in both tuberous sclerosis complex (TSC)-associated LAM (TSC-LAM) and sporadic LAM (S-LAM) patients.Methods: We retrospectively evaluated radiologic findings of chest HRCT (with a section thickness of 2 mm) from 311 females, including 272 patients with S-LAM (mean age 39.2 years) and 39 patients with TSC-LAM (mean age 38.3 years) who were seen at our hospital between April 2009 and December 2016.Results: We found 2 types of radiologic findings likely to make HRCT cyst appearance atypical: characteristics of the cyst itself and atypical findings in addition to cysts.We found that approximately 80% of LAM patients, whether TSC-associated or sporadic, showed typical HRCT appearance with mild to severe cystic destruction.The remaining 20% displayed unusual profiles in cyst appearance as well as additional findings aside from cyst: the former includes large cyst, thickened walls, and irregularly shaped whereas the latter includes ground glass attenuation and diffuse noncalcified nodules.

Sarcoidosis is a systemic, granulomatous disease caused by unknown immunological abnormalities. The organs most vulnerable to sarcoidosis are the lungs. Patients often resolve spontaneously, but the lungs can also be severely affected. Although details regarding prognostic factors in sarcoidosis patients with lung involvement remain unclear, several reports have suggested that immune checkpoint molecules are involved in the pathogenesis of sarcoidosis. In this study, we divided sarcoidosis patients into two groups based on chest computed tomography (CT) findings and compared immune checkpoint molecules expressed on T cells in bronchoalveolar lavage fluid (BALF) in the two groups, using flow cytometry. We found elevated programmed cell death 1 (PD-1) or T cell immunoglobulin- and mucin-domain-containing molecule-3 (TIM-3) expression on T cells in BALF in patients with spontaneous improvement in CT findings, compared with those in patients without improvement in CT findings. In conclusion, our study implies that PD-1 or TIM-3 expression on T cells in BALF may be a prognostic factor for pulmonary lesions in sarcoidosis.

The quantification of hepatitis C virus (HCV) is essential for the management of chronic hepatitis C therapy. We have developed a fully automated microfluidic RT-qPCR system for rapid quantitative detection of HCV RNA in human EDTA-plasma and serum, and the performance of the method was assessed. The platform for the assay, µTASWako g1 Fully Automated Genetic Analyzer, performs automated sample preparation and RNA extraction, followed by amplification and detection on an integrated RT-qPCR-CE (capillary electrophoresis (CE)) microfluidic chip. The total assay time from sample input to data output is less than 120 minutes. The HCV assay has a linear quantitative range of 15 to 107 IU/mL, with a limit of detection (LOD) of 10.65 IU/mL in EDTA-plasma and 12.43 IU/mL in serum. The assay has a reproducibility of SD ≤ 0.16 log10 IU/mL and an accuracy of ≤ 0.22 log10 IU/mL difference when compared to the assigned values. The main HCV genotypes 1 to 6 are detected with an accuracy of ± 0.3 log10 IU/mL. The assay is specific for HCV RNA and is free of interference from non-HCV pathogens, elevated levels of anti-viral and anti-bacterial drugs, and common endogenous interferents. In the linear quantitative range, the assay is highly correlated with the Roche cobas AmpliPrep/cobas TaqMan HCV Test, version 2.0 (r2 = 0.949). As the assay is highly sensitive, accurate and specific, and provides reliable quantification of HCV in plasma and serum, it can potentially be applicable for monitoring the therapy and management of HCV infection.

Pembrolizumab, an antiprogrammed cell death protein-1 monoclonal antibody, is predominantly administered in patients with advanced NSCLC. Recently, a few cases of immune checkpoint inhibitor (ICI)–induced Takotsubo cardiomyopathy (TC) were reported.1 However, it is difficult to clarify the correlation between ICI administration and TC because patients with cancer have several risk factors of TC. Here, we describe a case of a patient who was strongly suspected to have TC caused by an infusion reaction immediately after pembrolizumab administration.

spindled cells. Immunohistochemical stains were positive for SMA, HMB-45, ER, and PR. Both serum and ascitic fluid VEGF-D levels were high (928.6 pg/ml and 6980.9 pg/ml, respectively). She had no signs of tuberous sclerosis, therefore, she was diagnosed with sporadic LAM. Treatment was started with sirolimus 1mg/day, but her ascites did not improve. After two weeks, sirolimus was increased to 2mg/day, and leuprorelin was added. Thereafter, her ascites immediately disappeared and mass in the retroperitoneum and pelvis also shrank. After two months of this treatment, leuprorelin was discontinued and sirolimus was reduced by 1mg/day. Her ascites has not recurred for seven months. Discussion: To the best of our knowledge, the presentation of our patient is thought to be very rare and educational. Moreover, we think this is the first case report in which the treatment with sirolimus and leuprorelin was effective for refractory ascites in LAM. Conclusion: LAM should be considered in individuals presenting with chylous ascites, even with few lung lesions. Sirolimus is, of course, effective for refractory ascites in LAM patients, but additional treatment with leuprorelin may also be effective if its efficacy is poor.

THE EFFECTIVENESS AND SAFETY OF ANTIFIBROTICS FOR THE TREATMENT OF IDIOPATHIC PULMONARY FIBROSIS: A RETROSPECTIVE SINGLE CENTER ANALYSIS ATSUSHI SHIMAUCHI, KAZUYA TSUBOUCHI, RITSU IBUSUKI, TAKAFUMI KAYUKAWA, DAISUKE ETO, YUKI OKAMATSU, KATSUHIRO INOUE, TAISHI HARADA Department of Respiratory Medicine, Japan Community Health care Organization Kyushu Hospital, Kishinoura, Yahatanishi-ku, Fukuoka, Japan

Quantitative PCR (qPCR) has been widely used for the detection and monitoring of a variety of infectious diseases. PCR and CE were integrated into a microfluidic chip that was designed to achieve rapid real-time amplicon sampling, separation, and quantitation without requiring various probes. A novel chip design allows the overlapped execution of PCR and CE, minimizing the time required for CE analysis after each PCR cycle. The performance of the on-chip qPCR method was demonstrated using a 45-minutes model assay protocol for the phiX174 bacteriophage, and the multiplexing capability of the method was demonstrated by adding a second target, E. coli genomic DNA, to the model assay. The results indicate good sensitivity, reproducibility, and linearity over the tested assay range, 50 to 2 × 10(4) copies/25 μL reaction. Based on this performance, the on-chip qPCR method should be applicable to a wide variety of infectious disease detection and monitoring assays with the addition of suitable sample preparation protocols.