Calculus bovis inhibits M2 tumor-associated macrophage polarization via Wnt/ß-catenin pathway modulation to suppress liver cancer.

BACKGROUND: Calculus bovis (CB), used in traditional Chinese medicine, exhibits anti-tumor effects in various cancer models. It also constitutes an integral component of a compound formulation known as Pien Tze Huang, which is indicated for the treatment of liver cancer. However, its impact on the liver cancer tumor microenvironment, particularly on tumor-associated macrophages (TAMs), is not well understood. AIM: To elucidate the anti-liver cancer effect of CB by inhibiting M2-TAM polarization via Wnt/ß-catenin pathway modulation. METHODS: This study identified the active components of CB using UPLC-Q-TOF-MS, evaluated its anti-neoplastic effects in a nude mouse model, and elucidated the underlying mechanisms via network pharmacology, transcriptomics, and molecular docking. In vitro assays were used to investigate the effects of CB-containing serum on HepG2 cells and M2-TAMs, and Wnt pathway modulation was validated by real-time reverse transcriptase-polymerase chain reaction and Western blot analysis. RESULTS: This study identified 22 active components in CB, 11 of which were detected in the bloodstream. Preclinical investigations have demonstrated the ability of CB to effectively inhibit liver tumor growth. An integrated approach employing network pharmacology, transcriptomics, and molecular docking implicated the Wnt signaling pathway as a target of the antineoplastic activity of CB by suppressing M2-TAM polarization. In vitro and in vivo experiments further confirmed that CB significantly hinders M2-TAM polarization and suppresses Wnt/ß-catenin pathway activation. The inhibitory effect of CB on M2-TAMs was reversed when treated with the Wnt agonist SKL2001, confirming its pathway specificity. CONCLUSION: This study demonstrated that CB mediates inhibition of M2-TAM polarization through the Wnt/ß-catenin pathway, contributing to the suppression of liver cancer growth.

Three-dimensional cultured human umbilical cord mesenchymal stem cells attenuate pulmonary fibrosis by improving the balance of mitochondrial fusion and fission.

Pulmonary fibrosis is a kind of fibrotic interstitial pneumonia with poor prognosis. Aging, environmental pollution, and coronavirus disease 2019 are considered as independent risk factors for pulmonary fibrogenesis. Consequently, the morbidity and mortality striking continues to rise in recent years. However, the clinical therapeutic efficacy is very limited and unsatisfactory. So it is necessary to develop a new effective therapeutic approach for pulmonary fibrosis. Human umbilical cord mesenchymal stem cells (hucMSCs) are considered as a promising treatment for various diseases because of their multiple differentiation and immunomodulatory function. The key bottleneck in the clinical application of hucMSCs therapy is the high-quality and large-scale production. This study used FloTrix miniSpin bioreactor, a three-dimensional (3D) cell culture system, for large-scale expansion of hucMSCs in vitro, and proved 3D cultured hucMSCs inhibited the differentiation of fibroblasts into myofibroblasts and myofibroblasts proliferation and migration, leading to slow down the development of pulmonary fibrosis. Further mechanistic studies clarified that hucMSCs reduced the amount of binding between circELP2 and miR-630, resulting in blocking YAP/TAZ translocation from cytoplasm to nucleus. This condition inhibited mitochondrial fusion and promoted mitochondrial fission, and ultimately improved fusion/fission balance and cellular homeostasis. To sum up, this work clarified the anti-fibrosis and mechanism of hucMSCs cultured from the 3D FloTrix miniSpin bioreactor. We hope to provide new ideas and new methods for the clinical transformation and industrialization of hucMSCs therapy.

circELP2 reverse-splicing biogenesis and function as a pro-fibrogenic factor by targeting mitochondrial quality control pathway.

Idiopathic pulmonary fibrosis (IPF) is considered as a chronic, fibrosing interstitial pneumonia with unknown mechanism. The present work aimed to explore the function, biogenesis and regulatory mechanism of circELP2 in pulmonary fibrosis and evaluate the value of blocking circELP2-medicated signal pathway for IPF treatment. The results showed that heterogeneous nuclear ribonucleoprotein L initiated reverse splicing of circELP2 resulting in the increase of circELP2 generation. The biogenetic circELP2 activated the abnormal proliferation and migration of fibroblast and extracellular matrix deposition to promote pulmonary fibrogenesis. Mechanistic studies demonstrated that cytoplasmic circELP2 sponged miR-630 to increase transcriptional co-activators Yes-associated protein 1 (YAP1) and transcriptional co-activator with PDZ-binding motif (TAZ). Then, YAP1/TAZ bound to the promoter regions of their target genes, such as mTOR, Raptor and mLST8, which in turn activated or inhibited the genes expression in mitochondrial quality control pathway. Finally, the overexpressed circELP2 and miR-630 mimic were packaged into adenovirus vector for spraying into the mice lung to evaluate therapeutic effect of blocking circELP2-miR-630-YAP1/TAZ-mitochondrial quality control pathway in vivo. In conclusion, blocking circELP2-medicated pathway can alleviate pulmonary fibrosis, and circELP2 may be a potential target to treat lung fibrosis.

Proteomic analysis reveals the aging-related pathways contribute to pulmonary fibrogenesis.

Aging usually causes lung-function decline and susceptibility to chronic lung diseases, such as pulmonary fibrosis. However, how aging affects the lung-fibrosis pathways and leads to the occurrence of pulmonary fibrosis is not completely understood. Here, mass spectrometry-based proteomics was used to chart the lung proteome of young and old mice. Micro computed tomography imaging, RNA immunoprecipitation, dual-fluorescence mRFP-GFP-LC3 adenovirus monitoring, transmission electron microscopy, and other experiments were performed to explore the screened differentially expressed proteins related to abnormal ferroptosis, autophagy, mitochondria, and mechanical force in vivo, in vitro, and in healthy people. Combined with our previous studies on pulmonary fibrosis, we further demonstrated that these biological processes and underlying molecular players were also involved in the aging process. Our work depicted a comprehensive cellular and molecular atlas of the aging lung and attempted to explain why aging is a risk factor for pulmonary fibrosis and the role that aging plays in the progression of pulmonary fibrosis. The abnormalities of aging triggered an increase in mechanical force and ferroptosis, autophagy blockade, and mitochondrial dysfunction, which often appear during pulmonary fibrogenesis. We hope that the elucidation of these anomalies will help to enhance our understanding of senescence-inducing pulmonary fibrosis, thereby guiding the use of anti-senescence as an entry point for early intervention in pulmonary fibrosis and age-related diseases.

hucMSCs Treatment Ameliorated Pulmonary Fibrosis via Downregulating the circFOXP1-HuR-EZH2/STAT1/FOXK1 Autophagic Axis.

This study was performed to determine the effect of human umbilical cord mesenchymal stem cells (hucMSCs) treatment on pulmonary fibrosis and investigate the circFOXP1-mediated autophagic mechanism of hucMSCs treatment. Pulmonary fibrosis models were established by spraying bleomycin in mice and TGF-ß1 treatment of MRC-5 cells. Results showed that hucMSCs were retained in lung and hucMSCs treatment alleviated pulmonary fibrosis. Morphological staining indicated that hucMSCs-treated mice had thinner alveolar walls, effectively improved alveolar structure, significantly reduced alveolar inflammation, and decreased collagen deposition than control mice. Fibrotic proteins, including vimentin, α-SMA, collagens I and III, and the differentiation-related protein S100 calcium-binding protein A4 was reduced considerably in the hucMSCs-treated group. The mechanistic study revealed that the inhibition of hucMSCs treatment on pulmonary fibrogenesis depended on downregulating circFOXP1, in which hucMSCs treatment promoted circFOXP1-mediated autophagy process via blocking the nuclear human antigen R (HuR) translocation and promoting the HuR degradation, leading to a marked decrease in autophagy negative regulators EZH2, STAT1, and FOXK1. In conclusion, hucMSCs treatment significantly improved pulmonary fibrosis by downregulating the circFOXP1-HuR-EZH2/STAT1/FOXK1 autophagic axis. hucMSCs can act as an effective treatment for pulmonary fibrosis.

hucMSCs treatment prevents pulmonary fibrosis by reducing circANKRD42-YAP1-mediated mechanical stiffness.

Idiopathic pulmonary fibrosis (IPF) is a fibrosing interstitial pneumonia of unknown cause. The most typical characteristic of IPF is gradual weakening of pulmonary elasticity and increase in hardness/rigidity with aging. This study aims to identify a novel treatment approach for IPF and explore mechanism of mechanical stiffness underlying human umbilical cord mesenchymal stem cells (hucMSCs) therapy. Target ability of hucMSCs was examined by labeling with cell membrane dye Dil. Anti-pulmonary fibrosis effect of hucMSCs therapy by reducing mechanical stiffness was evaluated by lung function analysis and MicroCT imaging system and atomic force microscope in vivo and in vitro. Results showed that stiff environment of fibrogenesis caused cells to establish a mechanical connection between cytoplasm and nucleus, initiating expression of related mechanical genes such as Myo1c and F-actin. HucMSCs treatment blocked force transmission and reduced mechanical force. For further exploration of mechanism, ATGGAG was mutated to CTTGCG (the binding site of miR-136-5p) in the full-length sequence of circANKRD42. Wildtype and mutant plasmids of circANKRD42 were packaged into adenovirus vectors and sprayed into lungs of mice. Mechanistic dissection revealed that hucMSCs treatment repressed circANKRD42 reverse splicing biogenesis by inhibiting hnRNP L, which in turn promoted miR-136-5p binds to 3'-Untranslated Region (3'-UTR) of YAP1 mRNA directly, thus inhibiting translation of YAP1 and reducing YAP1 protein entering nucleus. The condition repressed expression of related mechanical genes to block force transmission and reduce mechanical forces. The mechanosensing mechanism mediated directly by circANKRD42-YAP1 axis in hucMSCs treatment, which has potential general applicability in IPF treatment.

AhR activation promotes Treg cell generation by enhancing Lkb1-mediated fatty acid oxidation via the Skp2/K63-ubiquitination pathway.

Several aryl hydrocarbon receptor (AhR) agonists have been reported to promote the generation of regulatory T cells (Treg cells), and the action mechanisms need to be identified. In this study, we addressed the underlying mechanism of AhR activation to induce the generation of Treg cells in the view of cellular metabolism. Naïve CD4+ T cells were purified with mouse CD4+ CD62L+ T Cells Isolation Kits. The proportions of Treg cells were detected by flow cytometry. The value of oxygen consumption rate (OCR) of CD4+ T cells was detected by the Seahorse XFe 96 analyzer. The activation of fatty acid oxidation (FAO)-related metabolic pathways was detected by Western blotting. Intracellular localization of Lkb1 was detected by immunofluorescence. The Strad-Mo25-Lkb1 complex formation and K63 chain ubiquitination modification of Lkb1 were detected by co-immunoprecipitation. The binding of AhR to the Skp2 promoter was detected by constructing luciferase reporter gene. AhR or carnitine palmitoyltransferases 1 was knockdown in dextran sulphate sodium (DSS)-induced colitis or collagen-induced arthritis (CIA) mice by infecting mice with adeno-associated virus via the tail vein injection. Compared to the control group, exogenous and endogenous AhR agonists 3,3'-diindolylmethane (DIM) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) were shown to preferentially upregulate the mRNA expression of FAO-related enzymes and the value of OCR. Consistently, pharmacological or genetic inhibition of FAO markedly diminished the induction of DIM and ITE on the differentiation of Treg cells. DIM and ITE functioned mainly through activating the liver kinase B1 (Lkb1)-AMPK pathway via promotion of Lkb1-Strad-Mo25 complex formation and Lkb1 K63 ubiquitination. DIM and ITE were also shown to upregulate the mRNA expression of Skp2, a ubiquitination-related enzyme, and facilitate the binding of AhR to the xenobiotic responsive element of Skp2 promoter region by luciferase reporter gene assay. Furthermore, the contribution of Skp2/K63 ubiquitination/Lkb1/FAO axis was verified in (DSS)-induced colitis or CIA mice. In summary, these findings indicate that AhR activation promotes Treg cell generation by enhancing Lkb1-mediated FAO via the Skp2/K63-ubiquitination pathway, and AhR agonists may be used as inducers of Treg cells to prevent and treat autoimmune diseases.

Evaluating the Effect of Hydrogen Sulfide in the Idiopathic Pulmonary Fibrosis Model with a Fluorescent Probe.

Hydrogen sulfide (H2S), a gaseous signaling molecule, is involved in a wide range of physiological and pathological processes. H2S has been proven to play a beneficial role in lung diseases, and the relationship between perturbations in endogenous H2S synthesis and degree with idiopathic pulmonary fibrosis (IPF) has attacted increasing attention. However, the changes in endogenous lung H2S levels in the pathological progression of chronic pulmonary diseases remain unclear. To this end, we synthesized a fluorescent probe (Bcy-HS) for the selective imaging of H2S in living cells and mice. This probe was mainly used for in situ in vivo and cellular imaging as well as a systematic assessment of intrapulmonary H2S levels at different stages of IPF. In addition, we also discussed the potential of H2S supplementation in the treatment of pulmonary fibrotic diseases. Our results confirmed the key role of H2S in pulmonary fibrosis. In cellular and mice models of pulmonary fibrosis, intracellular H2S levels are reduced. However, the severity of oxidative damage and pulmonary fibrosis decreased after NaSH (H2S donor). Therefore, we concluded that increasing the H2S content in vivo may be a novel strategy for IPF treatment.

Development and validation of a nomogram for the early prediction of acute kidney injury in hospitalized COVID-19 patients.

Introduction: Acute kidney injury (AKI) is a prevalent complication of coronavirus disease 2019 (COVID-19) and is closely linked with a poorer prognosis. The aim of this study was to develop and validate an easy-to-use and accurate early prediction model for AKI in hospitalized COVID-19 patients. Methods: Data from 480 COVID-19-positive patients (336 in the training set and 144 in the validation set) were obtained from the public database of the Cancer Imaging Archive (TCIA). The least absolute shrinkage and selection operator (LASSO) regression method and multivariate logistic regression were used to screen potential predictive factors to construct the prediction nomogram. Receiver operating curves (ROC), calibration curves, as well as decision curve analysis (DCA) were adopted to assess the effectiveness of the nomogram. The prognostic value of the nomogram was also examined. Results: A predictive nomogram for AKI was developed based on arterial oxygen saturation, procalcitonin, C-reactive protein, glomerular filtration rate, and the history of coronary artery disease. In the training set, the nomogram produced an AUC of 0.831 (95% confidence interval [CI]: 0.774-0.889) with a sensitivity of 85.2% and a specificity of 69.9%. In the validation set, the nomogram produced an AUC of 0.810 (95% CI: 0.737-0.871) with a sensitivity of 77.4% and a specificity of 78.8%. The calibration curve shows that the nomogram exhibited excellent calibration and fit in both the training and validation sets. DCA suggested that the nomogram has promising clinical effectiveness. In addition, the median length of stay (m-LS) for patients in the high-risk group for AKI (risk score ≥ 0.122) was 14.0 days (95% CI: 11.3-16.7 days), which was significantly longer than 8.0 days (95% CI: 7.1-8.9 days) for patients in the low-risk group (risk score <0.122) (hazard ratio (HR): 1.98, 95% CI: 1.55-2.53, p < 0.001). Moreover, the mortality rate was also significantly higher in the high-risk group than that in the low-risk group (20.6 vs. 2.9%, odd ratio (OR):8.61, 95%CI: 3.45-21.52). Conclusions: The newly constructed nomogram model could accurately identify potential COVID-19 patients who may experience AKI during hospitalization at the very beginning of their admission and may be useful for informing clinical prognosis.

Madecassic acid alleviates colitis-associated colorectal cancer by blocking the recruitment of myeloid-derived suppressor cells via the inhibition of IL-17 expression in γδT17 cells.

INTRODUCTION: Madecassic acid (MA), a triterpene compound isolated from Centella Asiatica herbs, has previously been shown to attenuate colitis induced by DSS in mice. In the present study, we address whether and how MA ameliorates colitis-associated colorectal cancer (CAC), which accounts for a considerable proportion of colorectal cancer. METHODS: CAC was induced by AOM/DSS in mice, and MA was administered orally once a day. To identify the source cells of IL-17 and the target cells for MA reducing the expression of IL-17 in the colons of CAC mice, single-cell suspensions were prepared from the colons of CAC mice and analyzed by flow cytometry. An adoptive transfer experiment was performed to verify the importance of the decreasing γδT17 cell population in the anti-CAC effect of MA. RESULTS: Oral administration of MA reduced the burden and incidence of tumors in the CAC mice. MA decreased the number of MDSCs in the colon tissues of CAC mice and ameliorated anti-tumor immune responses. MA could prevent the migration of MDSCs by inhibiting the activation of γδT17 cells and the expression of chemokines. The population of activated-γδT17 cells in the tumor microenvironment of CAC mice positively correlated with the number of MDSCs and tumors as well as tumor load. Moreover, the anti-CAC effect of MA was significantly counteracted by the adoptive transfer of γδT17 cells. CONCLUSIONS: MA alleviates CAC by blocking the recruitment of MDSCs to increase the population of anti-tumor immune cells in tumor microenvironment via inhibition of the activation of γδT17 cells.

Tetrandrine, an immunosuppressive alkaloid isolated from Steohania tetrandra S. Moore, induces the generation of Treg cells through enhancing fatty acid oxidation.

Our previous studies have demonstrated that tetrandrine can induce the generation of regulatory T (Treg) cells in vitro and in vivo. But, the underlying mechanism of tetrandrine remains obscure. Naïve CD4+ T cells are isolated from the mesenteric lymph nodes of mice for the differentiation of Treg cells. Flow cytometry is used to detect the frequencies of Treg cells. Non-targeted metabolomics analysis based on UHPLC-QTOF/MS is performed to assess the intracellular metabolic profiles. ChIP-PCR analysis is conducted to detect the level of H3K27ac at Foxp3 promoter and CNS regions. Tetrandrine treatment alters the metabolic profile of Treg cells, and pathway enrichment of differential metabolites mainly involves fatty acid oxidation (FAO). Tetrandrine promotes the mRNA expression of carnitine palmitoyl transferase-1, and increases the level of acetyl coenzyme A (acetyl-CoA) and the intracellular oxygen consumption rate. Either CPT1 inhibitor (etomoxir) or siRNA markedly diminishes the promotion of tetrandrine on Treg cell differentiation. Furthermore, tetrandrine enhances the acetylation of H3K27 in the promoter and CNS1 regions of Foxp3 through the acetyl-CoA derived from FAO. In the mice with collagen-induced arthritis, tetrandrine also induces Treg cell generation through FAO pathway. In addition, tetrandrine enhances the immunosuppressive function of Treg cells both in vitro and in vivo. The findings indicate that tetrandrine promotes Treg cell differentiation by enhancing FAO-mediated Foxp3 acetylation, and the CPT1-mediated FAO can serve as the target for the discovery of novel inducers of Treg cell generation.

Phytoestrogen arctigenin preserves the mucus barrier in inflammatory bowel diseases by inhibiting goblet cell apoptosis via the ERß/TRIM21/PHB1 pathway.

Intestinal mucus barrier dysfunction is closely involved in the pathogenesis of inflammatory bowel diseases (IBD). To investigate the protective effect and underlying mechanism of arctigenin, a phytoestrogen isolated from the fruits of Arctium lappa L., on the intestinal mucus barrier under colitis condition. The role of arctigenin on the intestinal mucus barrier and the apoptosis of goblet cells were examined by using both in vitro and in vivo assays. Arctigenin was demonstrated to promote the mucus secretion and maintain the integrity of mucus barrier, which might be achieved by an increase in the number of goblet cells via inhibiting apoptosis. Arctigenin selectively inhibited the mitochondrial pathway-mediated apoptosis. Moreover, arctigenin elevated the protein level of prohibitin 1 (PHB1) through blocking the ubiquitination via activation of estrogen receptor ß (ERß) to competitively interact with PHB1 and disrupt the binding of tripartite motif 21 (TRIM21) with PHB1. ERß knock down in the colons of mice with DSS-induced colitis resulted in significant reduction of the protection of arctigenin and DPN against the mucosal barrier. Arctigenin can maintain the integrity of the mucus barrier by inhibiting the apoptosis of goblet cells through the ERß/TRIM21/PHB1 pathway.

The neuropeptide cortistatin attenuates Th17 cell response through inhibition of glycolysis via GHSR1.

The neuropeptide cortistatin (CST) has been reported to attenuate Th17 cell response in multiple disease models, but the mechanism of action remains obscure. Here, we show that either overexpression or exogenous addition of CST obviously restricts Th17 cell differentiation. As metabolic reprogramming plays an important role in Th17 cell development, we explore whether CST inhibits Th17 cell differentiation by regulating glycolysis. The results show that CST substantially attenuates the glycolysis during Th17 differentiation and down-regulates the mRNA expression of myelocytomatosis oncogene (Myc) and hexokinase 2 (HK2), the glycolysis rate-limiting enzyme. Following the overexpression of Myc and HK2, the inhibitory effect of CST on Th17 differentiation nearly disappears, suggesting that Myc-HK2 pathway is deeply involved. Furthermore, growth hormone secretagogue receptor 1 (GHSR1) is identified as the key receptor subtype for CST attenuating glycolysis and Th17 cell differentiation by the combined uses of CST with various receptor subtype blockers. The knockdown of GHSR1 abrogates the inhibition of CST on Th17 differentiation and glycolysis. Finally, in the colitis mice induced by dextran sulfate sodium, an intraperitoneal injection of CST markedly inhibits Th17 cell response and down-regulates the expression of HK2 in the Th17 cells, which is reversed by the combined use of GHSR1 antagonist. These findings suggest that inhibition of glycolysis is the key pathway for CST attenuating Th17 cell response, and GHSR1, Myc and HK2 are potential therapeutic targets of Th17 cell-related diseases.

ATF3 -activated accelerating effect of LINC00941/lncIAPF on fibroblast-to-myofibroblast differentiation by blocking autophagy depending on ELAVL1/HuR in pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is characterized by lung scarring and has no effective treatment. Fibroblast-to-myofibroblast differentiation and myofibroblast proliferation and migration are major clinical manifestations of this disease; hence, blocking these processes is a practical treatment strategy. Here, highly upregulated LINC00941/lncIAPF was found to accelerate pulmonary fibrosis by promoting fibroblast-to-myofibroblast differentiation and myofibroblast proliferation and migration. Assay for transposase-accessible chromatin using sequencing and chromatin immunoprecipitation experiments elucidated that histone 3 lysine 27 acetylation (H3K27ac) activated the chromosome region opening in the LINC00941 promoter. As a consequence, the transcription factor ATF3 (activating transcription factor 3) bound to this region, and LINC00941 transcription was enhanced. RNA affinity isolation, RNA immunoprecipitation (RIP), RNase-RIP, half-life analysis, and ubiquitination experiments unveiled that LINC00941 formed a RNA-protein complex with ELAVL1/HuR (ELAV like RNA binding protein 1) to exert its pro-fibrotic function. Dual-fluorescence mRFP-GFP-MAP1LC3/LC3 (microtubule associated protein 1 light chain 3) adenovirus monitoring technology, human autophagy RT2 profiler PCR array, and autophagic flux revealed that the LINC00941-ELAVL1 axis inhibited autophagosome fusion with a lysosome. ELAVL1 RIP-seq, RIP-PCR, mRNA stability, and rescue experiments showed that the LINC00941-ELAVL1 complex inhibited autophagy by controlling the stability of the target genes EZH2 (enhancer of zeste 2 polycomb repressive complex 2 subunit), STAT1 (signal transducer and activators of transcription 1) and FOXK1 (forkhead box K1). Finally, the therapeutic effect of LINC00941 was confirmed in a mouse model and patients with IPF. This work provides a therapeutic target and a new effective therapeutic strategy related to autophagy for IPF.Abbreviations: ACTA2/a-SMA: actin alpha 2, smooth muscle; ATF3: activating transcription factor 3; ATG: autophagy related; Baf-A1: bafilomycin A1; BLM: bleomycin; CDKN: cyclin dependent kinase inhibitor; CLN3: CLN3 lysosomal/endosomal transmembrane protein, battenin; COL1A: collagen type I alpha; COL3A: collagen type III alpha; CXCR4: C-X-C motif chemokine receptor 4; DRAM2: DNA damage regulated autophagy modulator 2; ELAVL1/HuR: ELAV like RNA binding protein 1; EZH2: enhancer of zeste 2 polycomb repressive complex 2 subunit; FADD: Fas associated via death domain; FAP/FAPα: fibroblast activation protein alpha; FOXK1: forkhead box K1; FVC: forced vital capacity; GABARAP: GABA type A receptor-associated protein; GABARAPL2: GABA type A receptor associated protein like 2; IGF1: insulin like growth factor 1; IPF: idiopathic pulmonary fibrosis; LAMP: lysosomal associated membrane protein; lncRNA: long noncoding RNA; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; NPC1: NPC intracellular cholesterol transporter 1; RGS: regulator of G protein signaling; RPLP0: ribosomal protein lateral stalk subunit P0; ROC: receiver operating characteristic; S100A4: S100 calcium binding protein A4; SQSTM1/p62: sequestosome 1; STAT1: signal transducers and activators of transcription 1; TGFB1/TGF-ß1: transforming growth factor beta 1; TNF: tumor necrosis factor; UIP: usual interstitial pneumonia; ULK1: unc-51 like autophagy activating kinase 1; VIM: vimentin.

hnRNPL-activated circANKRD42 back-splicing and circANKRD42-mediated crosstalk of mechanical stiffness and biochemical signal in lung fibrosis.

Increasing circular RNAs (circRNAs) are involved in the progression of idiopathic pulmonary fibrosis (IPF). However, circRNA biogenesis and circRNA-mediated crosstalk between mechanical stiffness and biochemical signals in IPF remain obscure. In this study, a novel circRNA-ankyrin repeat domain 42 (ANKRD42) from peripheral blood of patients with IPF, which participated in pulmonary fibrosis through the close communication of mechanical stiffness and biochemical signals, was identified. Mechanistic studies revealed that the heterogeneous nuclear ribonucleoprotein L (hnRNP L) activated the circANKRD42 reverse splicing biogenesis. The biogenetic circANKRD42 sponged miR-324-5p to promote the AJUBA expression, which blocked the binding between phosphorylated yes-associated protein 1 (YAP1) and large tumor suppressor kinase 1/2 (LATS1/2), leading to increased YAP1 entering the nucleus. circANKRD42 also sponged miR-136-5p to promote the YAP1 translation. Accumulating YAP1 in nucleus bound to TEAD, which initiated the transcription of genes related to mechanical stiffness. Finally, the therapeutic effect of circANKRD42 was evaluated in mice and the association between circANKRD42 and clinicopathological features was analyzed in IPF patients. Our findings supported that circANKRD42 is a promising biomarker and a potential therapeutic target related to cytoskeleton tension for IPF treatment.

Pterostilbene pre-treatment reduces LPS-induced acute lung injury through activating NR4A1.

CONTEXT: Pterostilbene (PTE), a common polyphenol compound, exerts an anti-inflammatory effect in many diseases, including acute lung injury (ALI). OBJECTIVE: This study explores the potential mechanism of PTE pre-treatment against lipopolysaccharide (LPS)-induced ALI. MATERIALS AND METHODS: Sixty Sprague-Dawley rats were divided into control, ALI, 10 mg/kg PTE + LPS, 20 mg/kg PTE + LPS, and 40 mg/kg PTE + LPS groups. At 24 h before LPS instillation, PTE was administered orally. At 2 h before LPS instillation, PTE was again administered orally. After 24 h of LPS treatment, the rats were euthanized. The levels of inflammatory cells and inflammatory factors in the bronchoalveolar lavage fluid (BALF), the expression of nuclear receptor subfamily 4 group A member 1 (NR4A1), and the nuclear factor (NF)-κB pathway-related protein levels were detected. NR4A1 agonist was used to further investigate the mechanism of PTE pre-treatment. RESULTS: After PTE pre-treatment, the LPS induced inflammation was controlled and the survival rate was increased to 100% from 70% after LPS treatment 24 h. For lung injury score, it decreased to 1.5 from 3.5 after treating 40 mg/kg PTE. Compared with the control group, the expression of NR4A1 in the ALI group was decreased by 20-40%. However, the 40 mg/kg PTE pre-treatment increased the NR4A1 expression by 20-40% in the lung tissue. The results obtained with pre-treatment NR4A1 agonist were similar to those obtained by pre-treatment 40 mg/kg PTE. CONCLUSIONS: PTE pre-treatment might represent an appropriate therapeutic target and strategy for preventing ALI induced by LPS.

Mechanism of SMND-309 against lung injury induced by chronic intermittent hypoxia.

INTRODUCTION: Obstructive sleep apnea-hypopnea syndrome (OSAHS) is a common sleep disorder that causes severe physiological disturbance. Evidence showed that OSAHS is an important associated comorbidity that can affect the survival of patients with pulmonary fibrosis. Until now, the potential mechanisms by which OSAHS accelerates the progression of lung fibrosis remain unclear. By constructing a pathological model of chronic intermittent hypoxia (CIH), the present study aimed to explore the pathological progress and potential mechanism of lung injury caused by OSAHS. Meanwhile, SMND-309 was given for treatment to evaluate its potential therapeutic role in CIH-induced lung injury. METHODS: Mice were randomly divided into (C57BL/6 wild-type) WT+(room air) RA, WT + CIH, SMND-309 + RA, and SMND-309 + CIH groups. The WT + CIH and SMND-309 + CIH groups were exposed to CIH condition for 12 weeks, while the other groups were processed in normal oxygen at the same time. The SMND-309 + RA and SMND-309 + CIH groups were intraperitoneally injected with SMND-309 at the last week of the modeling period. After 12 weeks of treatment, three mice from each group were perfused through the heart. Lung tissues were isolated, fixed, sectioned, and stained with H&E, Masson, and immunofluorescence stain. The rest of the lung tissues were harvested for Western blot and ELISA assays. RESULTS: CIH treatment increased the expression of pro-inflammatory factors (TNF-α and IL-6), resulting in lung tissue structure disorder, inflammatory cell infiltration, increased pulmonary capillary permeability, and pulmonary edema. The activation of the NF-κB signaling pathway played a crucial role in the process of inflammation. Noticeably, we observed M2 macrophage accumulation in the lung after CIH exposure, which promoted epithelial-mesenchymal transition (EMT) and pulmonary tissue fibrosis. ELISA assays showed the increased expression of TGF-ß, IL-10, and IL-4 in the CIH group. SMND-309 inhibited pulmonary inflammation, reduced the accumulation of M2 macrophage, alleviated collagen deposition andlung damage. CONCLUSION: CIH could induce chronic lung inflammation, promote the activation of M2 macrophages, trigger the occurrence of EMT, and accelerate the deposition of lung collagen, eventually leading to lung tissue damage. This study presents a possible explanation by which interstitial lung diseases, particularly idiopathic pulmonary fibrosis (IPF) with OSAHS, are usually associated with fast progress and poor prognosis. SMND-309 showed a good protective effect on CIH-induced lung damage.

Evaluation of cyclooxygenase-2 fluctuation via a near-infrared fluorescent probe in idiopathic pulmonary fibrosis cell and mice models.

Idiopathic pulmonary fibrosis (IPF) is a devastating and fatal interstitial lung disease due to various challenges in diagnosis and treatment. Due to its complicated pathogenesis and difficulty in early diagnosis, there is no effective cure. Cyclooxygenase-2 (COX-2) is inextricably associated with pulmonary fibrosis. The abnormal level of COX-2 leads to extremely exacerbated pulmonary fibrosis. Therefore, we reported a near-infrared fluorescent probe Cy-COX to detect the fluctuation of COX-2 levels during pulmonary fibrosis and explain its important protective effect. The probe Cy-COX showed a significant enhancement of fluorescence signal to COX-2 with excellent selectivity and sensitivity. In order to clarify the relationship between COX-2 and pulmonary fibrosis, we used the probe Cy-COX to detect COX-2 fluctuation in organisms with pulmonary fibrosis. The results showed that the COX-2 level increased in the early stage and decreased in the late stage with the aggravation of pulmonary fibrosis. Furthermore, up-regulation of COX-2 levels can effectively alleviate the severity of pulmonary fibrosis. Therefore, Cy-COX is a fast and convenient imaging tool with great potential to predict the early stage of pulmonary fibrosis and evaluate the therapeutic effects.

Near-Infrared Fluorescent Probe for Imaging and Evaluating the Role of Vanin-1 in Chemotherapy.

Pantetheinase (also known as Vanin-1) is highly expressed in the liver, kidneys, and intestine and is closely associated with a number of diseases. Vanin-1 can hydrolyze pantetheine to pantothenic acid (vitamin B5) and cysteamine and participate in the synthesis of glutathione (GSH). GSH is highly expressed in tumor cells and plays a major role in the resistance of tumor cells to cisplatin. Therefore, we urgently need a method to monitor the activity level of Vanin-1 in tumor cells and tissues and elucidate the relationship between the role of Vanin-1 in GSH synthesis and tumor resistance. Herein, we report a Cy-Pa fluorescent probe for imaging Vanin-1 in cells and in vivo that can qualitatively and quantitatively detect the fluctuation of Vanin-1 concentrations in HepG2 and HepG2/DDP cells or tumor tissues of tumor-bearing mice. This probe shows excellent potential in in situ real-time monitoring of endogenous Vanin-1. Moreover, we proved that Vanin-1 can inhibit GSH synthesis using the probe. When the Vanin-1 inhibitor RR6 was used in combination with cisplatin, HepG2 and HepG2/DDP cells showed increased resistance to cisplatin, while the therapeutic efficiency of cisplatin was reduced in HepG2 and HepG2/DDP xenografts. In this study, Vanin-1 was shown to play an important role in the treatment of cancer, and the study of Vanin-1 may provide an idea for the treatment of cancer in the future.

Pulmonary metastasis of distal cholangiocarcinoma with multiple cavities in bilateral lungs: A case report.

Cholangiocarcinoma is a type of malignant tumor derived from the epithelium of the bile duct. Cases of cholangiocarcinoma metastasis to the lung are rare, especially those with imaging features of multiple cavities in bilateral lungs. Here, we report a case of a patient who had previously undergone radical resection of primary distal cholangiocarcinoma 18 months ago. Transbronchoscopic lung biopsy of the right lung and biopsy of the left supraclavicular lymph node were performed for pathology confirmation, as well as immunohistochemistry. Multiple cavity shadows in bilateral lungs and enlarged lymph nodes were found on the computed tomography (CT) scan obtained 18 months postoperatively. No obviously enlarged lymph nodes were observed under the carina and beside the aortic arch, whereas enlarged lymph nodes were found above the left clavicle. Biopsy of lung and supraclavicular lymph nodes confirmed metastatic adenocarcinoma. Immunohistochemistry showed that it originated from the digestive tract and had the same homology as cholangiocarcinoma (CK19 +, Villin +). Cholangiocarcinoma can be transferred to the lung and the left supraclavicular lymph nodes through the lymphatic pathway by characteristic jumping lymph node metastases. Diffuse cystic change is a specific CT manifestation of the lymphatic lung metastasis of cholangiocarcinoma.