Cigarette smoke-induced dysbiosis: comparative analysis of lung and intestinal microbiomes in COPD mice and patients.

BACKGROUND: The impact of cigarette smoke (CS) on lung diseases and the role of microbiome dysbiosis in chronic obstructive pulmonary disease (COPD) have been previously reported; however, the relationships remain unclear. METHODS: Our research examined the effects of 20-week cigarette smoke (CS) exposure on the lung and intestinal microbiomes in C57BL/6JNarl mice, alongside a comparison with COPD patients' intestinal microbiome data from a public dataset. RESULTS: The study found that CS exposure significantly decreased forced vital capacity (FVC), thickened airway walls, and induced emphysema. Increased lung damage was observed along with higher lung keratinocyte chemoattractant (KC) levels by CS exposure. Lung microbiome analysis revealed a rise in Actinobacteriota, while intestinal microbiome showed significant diversity changes, indicating dysbiosis. Principal coordinate analysis highlighted distinct intestinal microbiome compositions between control and CS-exposed groups. In the intestinal microbiome, notable decreases in Patescibacteria, Campilobacterota, Defferibacterota, Actinobacteriota, and Desulfobacterota were observed. We also identified correlations between lung function and dysbiosis in both lung and intestinal microbiomes. Lung interleukins, interferon-É£, KC, and 8-isoprostane levels were linked to lung microbiome dysbiosis. Notably, dysbiosis patterns in CS-exposed mice were similar to those in COPD patients, particularly of Global Initiative for Chronic Obstructive Lung Disease (GOLD) stage 4 patients. This suggests a systemic impact of CS exposure. CONCLUSION: In summary, CS exposure induces significant dysbiosis in lung and intestinal microbiomes, correlating with lung function decline and injury. These results align with changes in COPD patients, underscoring the important role of microbiome in smoke-related lung diseases.

Role of DCLK1/Hippo pathway in type II alveolar epithelial cells differentiation in acute respiratory distress syndrome.

BACKGROUND: Delay in type II alveolar epithelial cell (AECII) regeneration has been linked to higher mortality in patients with acute respiratory distress syndrome (ARDS). However, the interaction between Doublecortin-like kinase 1 (DCLK1) and the Hippo signaling pathway in ARDS-associated AECII differentiation remains unclear. Therefore, the objective of this study was to understand the role of the DCLK1/Hippo pathway in mediating AECII differentiation in ARDS. MATERIALS AND METHODS: AECII MLE-12 cells were exposed to 0, 0.1, or 1 µg/mL of lipopolysaccharide (LPS) for 6 and 12 h. In the mouse model, C57BL/6JNarl mice were intratracheally (i.t.) injected with 0 (control) or 5 mg/kg LPS and were euthanized for lung collection on days 3 and 7. RESULTS: We found that LPS induced AECII markers of differentiation by reducing surfactant protein C (SPC) and p53 while increasing T1α (podoplanin) and E-cadherin at 12 h. Concurrently, nuclear YAP dynamic regulation and increased TAZ levels were observed in LPS-exposed AECII within 12 h. Inhibition of YAP consistently decreased cell levels of SPC, claudin 4 (CLDN-4), galectin 3 (LGALS-3), and p53 while increasing transepithelial electrical resistance (TEER) at 6 h. Furthermore, DCLK1 expression was reduced in isolated human AECII of ARDS, consistent with the results in LPS-exposed AECII at 6 h and mouse SPC-positive (SPC+) cells after 3-day LPS exposure. We observed that downregulated DCLK1 increased p-YAP/YAP, while DCLK1 overexpression slightly reduced p-YAP/YAP, indicating an association between DCLK1 and Hippo-YAP pathway. CONCLUSIONS: We conclude that DCLK1-mediated Hippo signaling components of YAP/TAZ regulated markers of AECII-to-AECI differentiation in an LPS-induced ARDS model.

Assessing traffic-related air pollution-induced fiber-specific white matter degradation associated with motor performance declines in aged rats.

Fine particulate matter (PM2.5) is thought to exacerbate Parkinson's disease (PD) in the elderly, and early detection of PD progression may prevent further irreversible damage. Therefore, we used diffusion tensor imaging (DTI) for probing microstructural changes after late-life chronic traffic-related PM2.5 exposure. Herein, 1.5-year-old Fischer 344 rats were exposed to clean air (control), high-efficiency particulate air (HEPA)-filtered ambient air (HEPA group), and ambient traffic-related PM2.5 (PM2.5 group, 9.933 ± 1.021 µg/m3) for 3 months. Rotarod test, DTI tractographic analysis, and immunohistochemistry were performed in the end of study period. Aged rats exposed to PM2.5 exhibited motor impairment with decreased fractional anisotropy and tyrosine hydroxylase expression in olfactory and nigrostriatal circuits, indicating disrupted white matter integrity and dopaminergic (DA) neuronal loss. Additionally, increased radial diffusivity and lower expression of myelin basic protein in PM2.5 group suggested ageing progression of demyelination exacerbated by PM2.5 exposure. Significant production of tumor necrosis factor-α was also observed after PM2.5 exposure, revealing potential inflammation of injury to multiple fiber tracts of DA pathways. Microstructural changes demonstrated potential links between PM2.5-induced inflammatory white matter demyelination and behavioral performance, with indication of pre-manifestation of DTI-based biomarkers for early detection of PD progression in the elderly.

Data on lung and intestinal microbiome after air pollution exposure in ageing rats.

Air pollution has been linked to respiratory diseases, and urban air pollution can be attributed to a number of emission sources. The emitted particles and gases are the primary components of air pollution that enter the lungs during respiration. Particulate matter with an aerodynamic diameter of ≤ 2.5 µm (PM2.5) can deposit deep into the respiratory tract via inhalation and has been proposed as a causative agent for adverse respiratory health. In addition, the lung contains a diverse microbial community (microbiome) that maintains normal homeostasis and is significantly altered in a variety of pulmonary disorders. Air pollution, specifically PM2.5, has previously been shown to significantly alter the composition of the lower airway microbiome, which has been linked to decreased lung function in chronic obstructive pulmonary disease (COPD) patients. Surprisingly, the intestinal microbiome has also been implicated in the modulation of pulmonary inflammatory diseases. Therefore, dysbiosis of the lung and intestinal microbiomes pose significant negative effects on human health. This dataset describes the microbial community profiles of the lungs and intestines of ageing rats exposed to ambient unconcentrated traffic-related air pollution for three months. The whole-body exposure system was equipped with and without high efficiency particulate air (HEPA) filtration (gaseous vs. PM2.5 pollution). The data can provide valuable information on lung and intestinal microbiome changes, including that which was only found after traffic-related air pollution exposure.

A Giant Compressive Mesenteric Lipoblastoma Initially Suspected to Be Abdominal Malignancy: A Report of a Rare Case in a Nine-Month-Old Infant.

Lipoblastoma is a rare benign soft tissue neoplasm rising from embryonic white adipose tissue known as lipoblast that keeps proliferating during the postnatal period. Although lipoblastomas are benign, they often grow rapidly. Most lipoblastomas are asymptomatic at presentation; they can present as a growing painless palpable mass and progressive symptoms of various organ compression depending on localization. A giant mesenteric lipoblastoma is a rare case with only a few cases reported. An infant with large intraabdominal masses may present preoperative diagnostic difficulties. Differential diagnoses are broad and may include sarcomas, germ-cell tumors, lipomas, lymphomas, hepatoblastomas, Wilm's tumors, and neuroblastomas. Thorough clinical, radiological, and pathological investigations are ultimately required to obtain a definitive diagnosis. Regardless of location, the treatment of choice for lipoblastoma is complete surgical resection. All patients should be followed up for a minimum of five years We report a rare case of a giant compressive mesenteric lipoblastoma that was initially suspected as abdominal malignancy in a nine-month-old infant. As physicians, we must always consider the underlying cause as well as the malignant or benign nature of a growing mass to treat the patient appropriately.

Alteration in branching morphogenesis via YAP/TAZ in fibroblasts of fetal lungs in an LPS-induced inflammation model.

BACKGROUND: Chorioamnionitis is a common cause of preterm birth and leads to serious complications in newborns. The objective of this study was to investigate the role of the Hippo signaling pathway in lung branching morphogenesis under a lipopolysaccharide (LPS)-induced inflammation model. MATERIALS AND METHODS: IMR-90 cells and ex vivo fetal lungs were treated with 0, 10, 30, or 50 µg/ml LPS for 24 and 72 h. Supernatant levels of lactate dehydrogenase (LDH), interleukin (IL)-6, IL-8, Chemokine (C-X-C motif) ligand 1(CXCL1), branching and the surface area ratio, Yes-associated protein (YAP), transcription coactivator with PDZ-binding motif (TAZ), fibroblast growth factor 10 (FGF10), fibroblast growth factor receptor II (FGFR2), SRY-box transcription factor 2 (SOX2), SOX9, and sirtuin 1 (SIRT1) levels were examined. Differentially expressed genes in fetal lungs after LPS treatment were identified by RNA-sequencing. RESULTS: LPS at 50 µg/ml increased IL-6 and IL-8 in IMR-90 cells and increased IL-6, CXCL1 and LDH in fetal lungs. The branching ratio significantly increased by LPS at 30 µg/ml compared to the control but the increased level had decreased by 50 µg/ml LPS exposure. Exposure to 50 µg/ml LPS increased phosphorylated (p)-YAP, p-YAP/YAP, and p-TAZ/TAZ in IMR-90 cells, whereas 50 µg/ml LPS decreased FGF10 and SOX2. Consistently, p-YAP/YAP and p-TAZ/TAZ were increased in fibronectin+ cells of fetal lungs. Moreover, results of RNA-sequencing in fetal lungs showed that SMAD, FGF, IκB phosphorylation, tissue remodeling and homeostasis was involved in branching morphogenesis following exposure to 50 µg/ml LPS. The p-SIRT1/SIRT1 ratio increased in IMR-90 cells by LPS treatment. CONCLUSIONS: This study showed that regulation of the Hippo pathway in fibroblasts of fetal lungs was involved in branching morphogenesis under an inflammatory disease such as chorioamnionitis.

Diesel exhaust particles inhibit lung branching morphogenesis via the YAP/TAZ pathway.

Prenatal exposure to air pollution may associated with inhibition of lung development in the child, however the possible mechanism is unclear. We investigated the effects of traffic-related diesel exhaust particle (DEP) exposure on fetal lung branching morphogenesis and elucidate the possible mechanism. Ex vivo fetal lungs collected from ICR mice at an age of 11.5 embryonic (E) days were exposed to DEPs at 0 (control), 10, and 50 µg/mL and branching morphogenesis was measured for 3 days. Normal IMR-90 human fetal lung fibroblast cells were exposed to DEPs at 0 (control), 10, and 50 µg/mL for 24 h. We observed that DEP exposure significantly inhibited lung branching morphogenesis with reduced lung branching ratios and surface areas on day 3. RNA sequencing (RNA-Seq) showed that DEP increased the inflammatory response and impaired lung development-related gene expressions. DEPs significantly decreased Yes-associated protein (YAP), phosphorylated (p)-YAP, transcriptional coactivator with a PDZ-binding motif (TAZ), and p-TAZ in IMR-90 cells at 10 and 50 µg/mL. Treatment of fetal lungs with the YAP inhibitor, PFI-2, also demonstrated restricted lung branching development similar to that of DEP exposure, with a significantly decreased lung branching ratio on day 3. DEP exposure significantly decreased the lung branching modulators fibroblast growth factor receptor 2 (FGFR2), sex-determining region Y-box 2 (SOX2), and SOX9 in IMR-90 cells at 10 and 50 µg/mL. Fetal lung immunofluorescence staining showed that DEP decreased SOX2 expression in fibronectin+ fibroblasts. DEP exposure decreased the cellular senescence regulator, p-sirtuin 1 (SIRT1)/SIRT1 in IMR-90 cells, with RNA-Seq showing impaired telomere maintenance. DEP exposure impaired fetal lung growth during the pseudoglandular stage through dysregulating the Hippo signaling pathway, causing fibroblast lung branching restriction and early senescence. Prenatal exposure to traffic-related air pollution has adverse effects on fetal lung development.

High frequency of KRAS and EGFR mutation profiles in BRAF-negative thyroid carcinomas in Indonesia.

OBJECTIVE: Thyroid cancer incidence has steadily increased in Indonesia. However, data on Kirsten rat sarcoma virus (KRAS) and EGFR mutations in thyroid cancer in Indonesia remain unavailable, except for BRAF-V600E, the most common BRAF gene mutation. This study aimed to analyze KRAS and EGFR mutation profiles in BRAF-V600E negative thyroid cancer samples. RESULTS: BRAF-V600E mutations were found in papillary thyroid carcinomas in 40.3% patients with mean age of 53 years old. In BRAF-V600E-negative samples, 41.3% had KRAS mutations with mean age of 55.5 years old. KRAS mutation was found in 52.6% of follicular carcinomas and 47.4% of papillary thyroid carcinomas. Additionally, 45.7% had EGFR mutations in patients with mean age of 50.5 years old. EGFR mutation was found in 71.4% of papillary thyroid carcinoma and 28.6% of follicular carcinoma. Nearly half of the BRAF-V600E negative thyroid carcinoma samples harbored either KRAS or EGFR mutations. This finding suggests that in BRAF-V600E negative thyroid carcinoma samples, testing for RAS and EGFR mutation may be warranted for further therapeutic consideration.

Effects of antibiotics and metals on lung and intestinal microbiome dysbiosis after sub-chronic lower-level exposure of air pollution in ageing rats.

We investigated the effects of antibiotics, drugs, and metals on lung and intestinal microbiomes after sub-chronic exposure of low-level air pollution in ageing rats. Male 1.5-year-old Fischer 344 ageing rats were exposed to low-level traffic-related air pollution via whole-body exposure system for 3 months with/without high-efficiency particulate air (HEPA) filtration (gaseous vs. particulate matter with aerodynamic diameter of ≤2.5 µm (PM2.5) pollution). Lung functions, antibiotics, drugs, and metals in lungs were examined and linked to lung and fecal microbiome analyses by high-throughput sequencing analysis of 16 s ribosomal (r)DNA. Rats were exposed to 8.7 µg/m3 PM2.5, 10.1 ppb NO2, 1.6 ppb SO2, and 23.9 ppb O3 in average during the study period. Air pollution exposure decreased forced vital capacity (FVC), peak expiratory flow (PEF), forced expiratory volume in 20 ms (FEV20), and FEF at 25∼75% of FVC (FEF25-75). Air pollution exposure increased antibiotics and drugs (benzotriazole, methamphetamine, methyl-1 H-benzotriazole, ketamine, ampicillin, ciprofloxacin, pentoxifylline, erythromycin, clarithromycin, ceftriaxone, penicillin G, and penicillin V) and altered metals (V, Cr, Cu, Zn, and Ba) levels in lungs. Fusobacteria and Verrucomicrobia at phylum level were increased in lung microbiome by air pollution, whereas increased alpha diversity, Bacteroidetes and Proteobacteria and decreased Firmicutes at phylum level were occurred in intestinal microbiome. Lung function decline was correlated with increasing antibiotics, drugs, and metals in lungs as well as lung and intestinal microbiome dysbiosis. The antibiotics, drugs, and Cr, Co, Ca, and Cu levels in lung were correlated with lung and intestinal microbiome dysbiosis. The lung microbiome was correlated with intestinal microbiome at several phylum and family levels after air pollution exposure. Our results revealed that antibiotics, drugs, and metals in the lung caused lung and intestinal microbiome dysbiosis in ageing rats exposed to air pollution, which may lead to lung function decline.

Sacrococcygeal Yolk Sac Tumor in a Two-Year-Old Girl With Multiple Metastases: A Case Report.

Sacrococcygeal yolk sac tumor (YST) is an infrequent extra-gonadal malignant germ cell tumor (GCT) that occurs exclusively within the first two years of life. A two-year-old girl came with a massive mass on her left buttock, which continued to grow, and within three months had become extremely large and hindered her from walking. Physical examination revealed a sacrococcygeal mass of 15 cm in diameter. Multislice CT showed an intraluminal inferior cava vein mass extending into the pelvic cavity with coccygeal osseous destruction, pulmonary metastasis, and multiple hepatic metastases. Laboratory data revealed elevated tumor marker values for alpha-feto-protein (AFP), lactate dehydrogenase (LDH), and Ca-125. Cytopathology following fine needle aspiration biopsy evaluation of the smear sample revealed a cellular tumor with pseudo glandular, microcystic, and solid patterns. The cytopathology did not show pathognomic findings. An immunocytochemistry (IHC) examination of the cell block showed a positive result for anti-AFP antibody. The patient was diagnosed and treated with chemotherapy for a sacrococcygeal YST. Clinical follow-up on the fourth month showed that the tumor had shrunk to 4 cm in size. Laboratory follow-up data after four months showed significant improvement. Unfortunately, the patient passed away on the seventh cycle of chemotherapy due to lung and hepatic metastases.

Reduction of Emphysema Severity by Human Umbilical Cord-Derived Mesenchymal Stem Cells in Mice.

Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity and mortality in chronic lung disease patients throughout the world. Mesenchymal stem cells (MSCs) have been shown to regulate immunomodulatory, anti-inflammatory, and regenerative responses. However, the effects of human-umbilical-cord-derived mesenchymal stem cells (hUC-MSCs) on the lung pathophysiology of COPD remain unclear. We aimed to investigate the role of hUC-MSCs in emphysema severity and Yes-associated protein (Yap) phosphorylation (p-Yap) in a porcine-pancreatic-elastase (PPE)-induced emphysema model. We observed that the emphysema percentages (normalized to the total lung volume) measured by chest computed tomography (CT) and exercise oxygen desaturation were significantly reduced by hUC-MSCs at 107 cells/kg body weight (BW) via intravenous administration in emphysematous mice (p < 0.05). Consistently, the emphysema index, as assessed by the mean linear intercept (MLI), significantly decreased with hUC-MSC administration at 3 × 106 and 107 cells/kg BW (p < 0.05). Changes in the lymphocytes, monocytes, and splenic cluster of differentiation 4-positive (CD4+) lymphocytes by PPE were significantly reversed by hUC-MSC administration in emphysematous mice (p < 0.05). An increasing neutrophil/lymphocyte ratio was reduced by hUC-MSCs at 3 × 106 and 107 cells/kg BW (p < 0.05). The higher levels of tumor necrosis factor (TNF)-α, keratinocyte chemoattractant (KC), and lactate dehydrogenase (LDH) in bronchoalveolar lavage fluid (BALF) were significantly decreased by hUC-MSC administration (p < 0.05). A decreasing p-Yap/Yap ratio in type II alveolar epithelial cells (AECII) of mice with PPE-induced emphysema was significantly increased by hUC-MSCs (p < 0.05). In conclusion, the administration of hUC-MSCs improved multiple pathophysiological features of mice with PPE-induced emphysema. The effectiveness of the treatment of pulmonary emphysema with hUC-MSCs provides an essential and significant foundation for future clinical studies of MSCs in COPD patients.

A Potent Histone Deacetylase Inhibitor MPT0E028 Mitigates Emphysema Severity via Components of the Hippo Signaling Pathway in an Emphysematous Mouse Model.

Background: Chronic obstructive pulmonary disease (COPD) is a major cause of chronic mortality. The objective of this study was to investigate the therapeutic potential of a novel potent histone deacetylase (HDAC) inhibitor MPT0E028 on emphysema. Materials and Methods: A mouse model of porcine pancreatic elastase (PPE)-induced emphysema was orally administered 0, 25, or 50 mg/kg body weight (BW) of the MPT0E028 five times/week for 3 weeks. Pulmonary function, mean linear intercept (MLI), chest CT, inflammation, yes-associated protein (YAP), transcriptional coactivator with PDZ-binding motif (TAZ), surfactant protein C (SPC), T1-α, p53, and sirtuin 1 (SIRT1) levels were examined. Results: 50 mg/kg BW of the MPT0E028 significantly decreased the tidal volume in emphysematous mice (p < 0.05). Emphysema severity was significantly reduced from 26.65% (PPE only) to 13.83% (50 mg/kg BW of the MPT0E028). Total cell counts, neutrophils, lymphocytes, and eosinophils significantly decreased with both 25 and 50 mg/kg BW of the MPT0E028 (p < 0.05). Also, 50 mg/kg BW of the MPT0E028 significantly decreased the levels of KC, TNF-α, and IL-6 in lung tissues and serum (p < 0.05). Expressions of p-TAZ/TAZ in lung tissues significantly decreased with 50 mg/kg BW of the MPT0E028 (p < 0.05). Expressions of p53 significantly decreased in alveolar regions with 50 mg/kg BW of the MPT0E028 (p < 0.05), and the expression of SPC increased in alveolar regions with 50 mg/kg BW of the MPT0E028 (p < 0.05). Conclusions: Our study showed that the potent HDAC inhibitor MPT0E028 reduced the severity and inflammation of emphysema with improvement in lung function, which could be regulated by Hippo signaling pathway. The MPT0E028 may have therapeutic potential for emphysema.

Air pollution-regulated E-cadherin mediates contact inhibition of proliferation via the hippo signaling pathways in emphysema.

Air pollution has been linked to emphysema in chronic obstruction pulmonary disease (COPD). However, the underlying mechanisms in the development of emphysema due to air pollution remain unclear. The objective of this study was to investigate the role of components of the Hippo signaling pathway for E-cadherin-mediated contact inhibition of proliferation in the lungs after air pollution exposure. E-Cadherin-mediated contact inhibition of proliferation via the Hippo signaling pathway was investigated in Sprague-Dawley (SD) rats whole-body exposed to air pollution, and in alveolar epithelial A549 cells exposed to diesel exhaust particles (DEPs), E-cadherin-knockdown, and high-mobility group box 1 (HMGB1) treatment. Underlying epithelial differentiation, apoptosis, and senescence were also examined, and the interaction network among these proteins was examined. COPD lung sections were used to confirm the observations in rats. Expressions of HMGB1 and E-cadherin were negatively regulated in the lungs and A549 cells by air pollution, and this was confirmed by knockdown of E-cadherin and by treating A549 cells with HMGB1. Depletion of phosphorylated (p)-Yap occurred after exposure to air pollution and E-cadherin-knockdown, which resulted in decreases of SPC and T1α. Exposure to air pollution and E-cadherin-knockdown respectively downregulated p-Sirt1 and increased p53 levels in the lungs and in A549 cells. Moreover, the protein interaction network suggested that E-cadherin is a key activator in regulating Sirt1 and p53, as well as alveolar epithelial cell differentiation by SPC and T1α. Consistently, downregulation of E-cadherin, p-Yap, SPC, and T1α was observed in COPD alveolar regions with particulate matter (PM) deposition. In conclusion, our results indicated that E-cadherin-mediated cell-cell contact directly regulates the Hippo signaling pathway to control differentiation, cell proliferation, and senescence due to air pollution. Exposure to air pollution may initiate emphysema in COPD patients.

The Prevalence of the EML4-ALK Fusion Gene in Cytology Specimens from Patients with Lung Adenocarcinoma.

BACKGROUND: Under the National Comprehensive Cancer Network (NCCN) guidelines for non-small-cell lung carcinoma (NSCLC), anaplastic lymphoma kinase (ALK) gene rearrangement is required to be assessed. However, data showing the prevalence of the ALK rearrangement is still deficient and is not yet available in Indonesia. This study used direct smear preparation from transthoracic needle specimens that are minimally invasive. The main objective of the study is to identify the prevalence of the ALK fusion rearrangement gene in cytological specimens. MATERIALS AND METHODS: A total of 35 direct smear preparations diagnosed as lung adenocarcinoma and EGFR mutation negative were involved in this study. The samples were taken between 2017 and 2019. These samples were examined for EML4-ALK fusion rearrangement gene using qRT-PCR. The EML4-ALK rearrangement status was determined by qRT-PCR with high-resolution melting (HRM) analysis. RESULTS: A total of 28 (80%) samples were from males, and 7 samples were from females. Seven (20% 95% CI: 8.4%-36.9%) samples were EML4-ALK rearrangement positive. The average age of the patients was 63.5 years old. The most common sites of metastasis in this study were pleural cavity, bone, liver, and CNS. CONCLUSIONS: qRT-PCR successfully identified EML4-ALK fusion rearrangement in direct smear preparations of lung adenocarcinoma. Direct smear samples can be used for EML4-ALK rearrangement detection using qRT-PCR. The EML4-ALK rearrangement gene has high prevalence in selected lung adenocarcinoma and EGFR mutation-negative populations. ALK inhibitors in lung cancer can be openly considered for use in Indonesian patients to improve the outcome of this subset of patients.