TNF-α Signaling Pathway Research Articles

The therapeutic effect of Fufang Zhenshu Tiaozhi (FTZ) on osteoclastogenesis and ovariectomized-induced bone loss: evidence from network pharmacology, molecular docking and experimental validation.

Fufang Zhenshu Tiaozhi (FTZ) has been widely used in clinical practice and proven to be effective against aging-induced osteoporosis in mice. This study aimed to explore the mechanism of FTZ against osteoclastogenesis and ovariectomized-induced (OVX) bone loss through the network pharmacology approach. The ingredients of FTZ were collected from the previous UPLC results, and their putative targets were obtained through multiple databases. Differentially expressed genes (DEGs) during osteoclastogenesis were identified through multi-microarrays analysis. The common genes between FTZ targets and DEGs were used to perform enrichment analyses through the clusterProfier package. The affinity between all FTZ compounds and enriched genes was validated by molecular docking. The effects of FTZ on osteoclastogenesis and bone resorption were evaluated by TRAP staining, bone resorption assay and RT-qPCR in vitro, while its effects on bone loss by ELISA and Micro-CT in vivo. Enrichment analyses indicated that the inhibitory effects of FTZ may primarily involve the regulation of inflammation, osteoclastogenesis, as well as TNF-α signaling pathway. 130 pairs docking results confirmed FTZ ingredients have good binding activities with TNF-α pathway enriched genes. FTZ treatment significantly reduced TRAP, TNF-α, IL-6 serum levels and increased bone volume in OVX mice. Consistently, in vitro experiments revealed that FTZ-containing serum significantly inhibited osteoclast differentiation, bone resorption, and osteoclast related mRNA expression. This study revealed the candidate targets of FTZ and its potential mechanism in inhibiting osteoclastogenesis and bone loss induced by OVX, which will pave the way for the application of FTZ in the postmenopausal osteoporosis treatment.

Visual analysis on regulation of necroptosis with Chinese medicine based on VOSviewer and CiteSpace knowledge graphs

The study was conducted by searching the literature related to the regulation of necroptosis with Chinese medicine from January 1, 2005 to December 31, 2021 in CNKI, VIP, Wanfang, Web of Science(WoS), and PubMed. The obtained literature were imported into NoteExpress for eliminating duplicates and screening, and the final included articles were imported into Excel to plot the publication trend. The core authors were identified according to Price's law, and VOSviewer 1.6.17 was used to draw a collaborative view of the core authors and sort the high-frequency keywords. Then CiteSpace 5.8.R3 was employed to analyze keywords clustering, burst, and timeline view. Finally, 98 Chinese articles and 72 English articles were included in the study. The number of publications on the regulation of necroptosis with Chinese medicine has been increasing year by year. China ranked among the top in the world in terms of the number of publications, and Chinese authors played a central role in this field. Specifically, LIU Hua published the most Chinese literature while CHEN X P had the most English publications. The collaborative view of the core authors showed more intra-team cooperation and less inter-team cooperation. The Chinese and English keywords formed ten clusters separately, indicating that the research hotspots of regulation of necroptosis with Chinese medicine mainly focused on disease, prescription, related factors, and mecha-nism. Further, the analysis of Chinese and English keywords revealed that regarding disease treatment, tumor, ischemia-reperfusion injury, neurodegenerative diseases, and inflammatory diseases were studied most. The Chinese medicines that received much attention in this field were curcumin, shikonin and tanshinone. The main protein factors involved were Ripk1, Ripk3, Mlkl, and TNF-α, and Ripk1/Ripk3/Mlkl and p53 signaling pathways were predominant. Moreover, single herbs and herbal monomers were the hotspots of the included articles. In the future, scholars need to expand the study of classical Chinese herbal compounds and explore their mechanism of action in the occurrence and development of various diseases, to provide new ideas and experimental basis for the treatment of clinical diseases with Chinese medicine.

Downregulation of TNFR2 decreases survival gene expression, promotes apoptosis and affects the cell cycle of gastric cancer cells.

BACKGROUNDChronic inflammation due to Helicobacter pylori (H. pylori) infection promotes gastric carcinogenesis. Tumour necrosis factor-α (TNF-α), a key mediator of inflammation, induces cell survival or apoptosis by binding to two receptors (TNFR1 and TNFR2). TNFR1 can induce both survival and apoptosis, while TNFR2 results only in cell survival. The dysregulation of these processes may contribute to carcinogenesis.AIMTo evaluate the effects of TNFR1 and TNFR2 downregulation in AGS cells treated with H. pylori extract on the TNF-α pathway.METHODSAGS cell lines containing TNFR1 and TNFR2 receptors downregulated by specific shRNAs and nonsilenced AGS cells were treated with H. pylori extract for 6 h. Subsequently, quantitative polymerase chain reaction with TaqMan® assays was used for the relative quantification of the mRNAs (TNFA, TNFR1, TNFR2, TRADD, TRAF2, CFLIP, NFKB1, NFKB2, CASP8, CASP3) and miRNAs (miR-19a, miR-34a, miR-103a, miR-130a, miR-181c) related to the TNF-α signalling pathway. Flow cytometry was employed for cell cycle analysis and apoptosis assays.RESULTSIn nonsilenced AGS cells, H. pylori extract treatment increased the expression of genes involved in cell survival and inhibited both apoptosis (NFKB1, NFKB2 and CFLIP) and the TNFR1 receptor. TNFR1 downregulation significantly decreased the expression of the TRADD and CFLIP genes, although no change was observed in the cellular process or miRNA expression. In contrast, TNFR2 downregulation decreased the expression of the TRADD and TRAF2 genes, which are both important downstream mediators of the TNFR1-mediated pathway, as well as that of the NFKB1 and CFLIP genes, while upregulating the expression of miR-19a and miR-34a. Consequently, a reduction in the number of cells in the G0/G1 phase and an increase in the number of cells in the S phase were observed, as well as the promotion of early apoptosis.CONCLUSIONOur findings mainly highlight the important role of TNFR2 in the TNF-α pathway in gastric cancer, indicating that silencing it can reduce the expression of survival and anti-apoptotic genes.

P548: BEMCENTINIB COMBINED WITH LOW-DOSE CYTARABINE IS EFFICACIOUS AND WELL TOLERATED IN RELAPSED AML PATIENTS UNFIT FOR INTENSIVE CHEMOTHERAPY. UPDATES FROM THE ONGOING PHASE II TRIAL (NCT02488408)

Background: The new standard of care (SOC) in newly-diagnosed AML patients (pts) unfit for intensive chemotherapy (IC) due to age or co-morbidities yields favourable efficacy. However, beyond 1st line, these pts have limited treatment options, with a dismal mOS of 2.4 months at relapse, highlighting a significant unmet need for new treatments in this pt population. Bemcentinib (BEM) is a first-in-class, orally bioavailable, highly selective AXL-inhibitor. AXL (a receptor tyrosine kinase) represents an important target in AML, as its expression on AML cells and stem cell compartment is associated with poor prognosis, resistance to chemotherapy and decreased antitumor immune response. Aims: The ongoing BGBC003 PhII trial studied the safety and efficacy of the BEM+LDAC combination in relapsed (REL) AML pts unfit for IC; in addition, a comprehensive translational biomarker analysis was pursued. Here, we present preliminary efficacy and safety data and include initial translational multiomics data from bone marrow mononuclear cells (BMMNC). Methods: Pts received BEM at 200 mg/OD with 3 loading doses at 400 mg/OD and LDAC at SOC schedule. Study endpoints included overall survival (OS), objective response (OR), clinical benefit rate (CBR) (OR+unchanged [UC]) and exploratory biomarker analyses. Longitudinal BMMNC samples (n=32) from 13 pts underwent scRNA-seq and CITEseq (Chromium 10x genomics; TotalSeq, Biolegend). Pts were stratified by best response: CR, CRi, PR for Responders; UC, PD for Non-Responders. Cell type annotations were inferred from expression of known markers at RNA and protein level and for identification of malignant cells copy number variation was included. Results: As of 13 Dec 2021, 22 REL AML pts (10/22 in 1st relapse) were treated with BEM+LDAC. Median age was 75.5yrs [66-86], median prior lines of therapy 1 [1-8] and adverse cytogenetic risk in 7/22 (32%). Only 19/22 pts were evaluable for efficacy. The overall REL population (n=19) demonstrated a composite CR (CRc=CR+CRi) of 26% (5/19) and CBR of 79% (14/19); AML pts in 1st relapse (10/19) showed a CRc of 30% (3/10) and CBR of 70% (7/10) whereas REL pts with time on treatment (ToT) > 3 months (11/19) achieved CRc of 45% (5/11) and CBR 91% (9/11); median ToT of 12.9 months for CR/CRi pts; 1 pt remains on treatment. Median OS was 6.2 months in the overall REL pts (n=19), 7.4 months in the AML pts in 1st relapse (10/19) and 11.3 months in REL pts with ToT >3 months. Late onset responses suggest an immunological mechanism of action (iMOA) and may contribute to a longer ToT and survival. CITEseq analysis identified differences in the immune compartment, underscoring an iMOA associated with response to BEM+LDAC. Furthermore, increased gene set enrichment of TNFα signalling via NfkB at screening in malignant blasts of responders emphasizes the influence of the TNFα signalling pathway as an interesting field of further research. The safety profile of BEM+LDAC in the REL AML pts (n=19) is comparable with the known safety profile of LDAC. TRAEs of ≥G3 observed in ≥10% of pts were anaemia (33% BEM+LDAC; 22% BEM), and ECG QT prolonged (11% BEM+LDAC; 11% Bem). No G5 TRAEs reported. Summary/Conclusion: BEM+LDAC is efficacious and well tolerated in REL AML pts unfit for IC, with promising survival benefit compared to historical data. Translational research showed activation of CD8+T cells and B/Plasma cells in response to treatment, indicating that BEM elicits activation of two major adaptive immune cell populations responsible for anti-AML immune responses. BEM+LDAC warrants further evaluation in this population.

Abstract 255: Peroxidasin deficiency recruits pro-healing macrophages into the liver and inhibits NAFLD progression to HCC

Abstract Introduction: During liver fibrosis, tissue repair mechanisms replace necrotic tissue with highly stabilized extracellular matrix (ECM) proteins. ECM stabilization influences the speed of tissue recovery. Here, we used a mouse model of nonalcoholic fatty liver disease (NAFLD) to study the function of peroxidasin (PXDN), a peroxidase that uses H2O2 to cross-link collagen IV, during liver fibrosis progression to hepatocellular carcinoma (HCC). Method: Pxdn-/- and Pxdn+/+ mice were fed with a choline-deficient L-amino acid-defined high-fat diet (CDAHFD) for 16 weeks to create a NAFLD-HCC preclinical model. Liver histology, collagen content, flow cytometry, immunostaining of immune cells, RNA-seq, and liver function tests were analyzed. In vivo imaging of liver reactive oxygen species (ROS) was performed using a redox-active iron complex, Fe-PyC3A. Results: Using Fe-PyC3A as an MRI contrast agent, we detected a higher content of ROS in Pxdn-/- livers (healthy) that was not necessarily directly toxic but could activate hypoxia-related molecular pathways. Genome-wide expression analysis of liver tissue and differential gene expression (DGE) combined with Gene Ontology (GO) analysis identified significant upregulation of genes associated with hypoxia and TNFα signaling pathways already in Pxdn-/- sham livers (without injury). In addition, we observed an upregulation of genes involved in the innate immune response, leukocyte activation, and chemotaxis. After 16 weeks of CDAHFD, gross analysis of collected liver showed no HCC nodule formation in Pxdn-/- mice while 60% of the WT mice had HCC tumors. Collagen deposition showed less collagen accumulation in Pxdn-/- mice. Flow cytometry of macrophages showed Pxdn-/- mice had increased pro-healing M2 macrophages recruitment in early- and mid-stage NAFLD (4 weeks and 8 weeks on CDAHFD) compared to WT controls. In addition, we observed a significant decrease in the number of CD3+ T cells and CD8+ cytotoxic T cells in the late-stage of NAFLD in Pxdn KO mice. DGE analysis revealed that IL-12 is highly expressed in Pxdn-/- injured livers. Additionally, multiple other T cell-related molecules such as IL-10, IL-6, CCL2, IL-7, and CD4 were elevated in Pxdn-/- injured liver. Elevation of these cytokines is an indicator for higher recruitment of pro-healing and anti-HCC macrophage to the site of injury. Conclusion: Our findings demonstrate that PXDN deficiency is associated with the induction of the hypoxia and TNFα signaling pathways and the recruitment of pro-healing and anti-HCC macrophages to the liver. This results in significantly decreased collagen stabilization during liver fibrosis and accelerates fibrosis reversal. In addition, recruited macrophages-controlled T cell response and inhibited HCC formation in Pxdn-/- mice. Citation Format: Mozhdeh Sojoodi, Stephen C. Barrett, Derek J. Erstad, Shadi Salloum, Shijia Zhu, Tongqi Qian, Selene Colon, Eric Gale, Veronica Clavijo Jordan, Yongtao Wang, Shen Li, Michael Lanuti, Lawrence Zukerberg, Peter Caravan, Yujin Hoshida, Raymond T. Chung, Gautam Bhave, Georg M. Lauer, Bryan C. Fuchs, Kenneth K. Tanabe. Peroxidasin deficiency recruits pro-healing macrophages into the liver and inhibits NAFLD progression to HCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 255.

Abstract 5827: Increased miR150 expression in tumors enhances lymphocytes infiltration causing immunosuppression through inflammatory chemokines

Abstract Dysregulated miRNAs play key roles in multiple biological processes and cancer. miR-150-5p, a cancer-related miRNA, has been reported to be aberrantly expressed in various cancers. We found that triple-negative and HER2+ breast cancer patients have elevated expression of miR150 in the TCGA and METABRIC patient cohort. Both cohorts showed significantly increased expression of miR150 in grade IV patients. Further studies explained that miR150 is strongly correlated with CD8+ T cells and B cells in accord with increased expression of immune-checkpoint markers PD-1, PDL-1, and CTLA4. We, therefore, hypothesized that higher expression of miR150 in tumors attracted more immune cells in the tumor microenvironment. We use two triple negative breast cancer cell lines MDA-231 and BT-549 cells and overexpressed miR150 followed by co-culture them with Jurkart and B cells. Our data revealed that both jurkart and B cells had increased migration towards cancer cells overexpressed miR150 mimic compared to miR150 mimic negative control. Interestingly, overexpressed miR150 mimic in cancer cells inhibited migration in both triple-negative breast cancer cells. Overexpression of miR150 mimic in cancer cells did not alter cell proliferation. To understand further the role of miR150 in breast cancer, we overexpressed miR150 mimic and its corresponding negative control in MDA-231 and did RNA seq analysis. RNA seq data explained that approximately 4927 genes are upregulated with miR150 mimic overexpression. Interestingly several chemokines and mediators of metastasis (CXCL2, CXCL1 CCL20, IGBP3, and ANGPTL4) are upregulated among the top 50 upregulated genes. In addition, gene enrichment analysis explained that TNF-α signaling pathways, Hypoxia, epithelial-mesenchymal pathways, E2F signaling, and Inflammatory pathways are a few upregulated pathways with miR150 mimic overexpression compared to the negative control. We also found miR150 high tumor enriched correlated with interferon-alpha and gamma signaling, inflammatory response pathways similar with cohort analysis data. In conclusion, miR150 in cancer plays a key role in interferon and inflammatory signaling pathways attracting more immune cells and suppressing the tumor microenvironment. Citation Format: Aparna Maiti, Masanori Oshi, Rongrong Wu, Nitai Hait, Kazuaki Takabe. Increased miR150 expression in tumors enhances lymphocytes infiltration causing immunosuppression through inflammatory chemokines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5827.

Abstract 3724: PBRM1 loss in RCC: Implications for immunotherapy

Abstract Introduction The association between mutational status of the polybromo-1 (PBRM1) gene in clear cell renal cell carcinomas (ccRCCs) and papillary RCCs (pRCCs) in the response to immune checkpoint blockade is debatable. Here, we aim to characterize the biological changes that PBRM1 loss causes and how these changes might modulate the immune response to cancer. Methods and Results Gene set enrichment analysis (GSEA) of PBRM1-mutant RCC tumors from TCGA data sets reveals NF-kB signaling via TNFα, hypoxia, and p53 signaling pathways are enriched compared to PBRM1 WT. PBRM1-mutant tumors also have increased number of insertion/deletion mutations and non-synonymous mutations that are directly proportional to neoantigen burden. CIBERSORT analysis of PBRM1-mutant ccRCC tumors reveals decreased transcriptional signature associated with CD8+ T-cell infiltration and lower levels of two cytolytic signatures. PBRM1-mutant ccRCC cell lines have constitutively activated NF-kB signaling, as well as a greater induction of NF-kB in response to TNFα via western blot when compared with PBRM1 reconstituted cells. Additionally, PD-L1 is more strongly expressed on the cell surface of PBRM1-mutant cells, which may modulate their interactions with the immune system. However, MHC-I expression was not found to be significantly different between PBRM1 mutant and reconstituted cells. Conclusions These results shed light on changes in biology and immunogenicity associated with or caused by PBRM1 loss. They also implicate NF-kB as a potential moderator of immune response, which should be studied further to characterize its effects on immune checkpoint blockade therapy (ICB). Citation Format: Alexander Metz, Rahmat Sikder, Moataz Ellithi, Micheal Slifker, Philip Abbosh. PBRM1 loss in RCC: Implications for immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3724.

Qingfeiyin Decoction Inhibits H1N1 Virus Infection via Modulation of Gut Microbiota and Inflammatory Pathways in a Murine Model

Influenza virus-caused lung infection and its pandemic outbreaks are a persistent public health challenge. The H1N1 subtype is the most common type of influenza infection observed in humans. Maxingshigantang decoction, a classic formula of Chinese herbal medicine, has been used for the prevention and treatment of respiratory infection for many centuries. Qingfeiyin decoction, based on Maxingshigantang, has been used in the clinic for decades. To explore the underlying mechanisms, according to the traditional Chinese medicine theory “the lung and the large intestine are interior–exterior,” which can be translated to the “gut–lung axis” in a contemporary term, the composition of gut microbiota was determined using 16S rRNA and the transcriptome of the colon was determined by RNA sequencing. The results showed that Qingfeiyin decoction decreased the viral load, alleviated the lung injury, increased the survival rate, partly restored the shortening of the colon caused by the H1N1 virus, and downregulated inflammatory pathways including MAPK, TNFα, and JAK-STAT signaling pathways. Qingfeiyin decoction increased the relative abundance of the genera of Coprococcus , Ruminococcus, Lactobacillus, and Prevotella and prevented the H1N1 virus-induced decrease in the abundance of the genera of Escherichia, Parabacteroides, Butyricimonas, and Anacrotruncus. These results will help better understand the mechanisms for Qingfeiyin decoction’s protective effect against influenza virus infection.

Gpr174 Knockout Alleviates DSS-Induced Colitis via Regulating the Immune Function of Dendritic Cells.

BackgroundDysfunction of the immune system would disturb the intestinal homeostasis and lead to inflammatory bowel disease (IBD). Dendritic cells (DCs) help maintain intestinal homeostasis and immediately respond to pathogens or injuries once the mucosa barriers are destroyed during IBD. G protein-coupled receptors(GPR)174 is an essential regulator of immunity that is widely expressed in most immune cells, including DCs. However, the role of GPR174 in regulating the immune function of DC in colitis has not been investigated.MethodsDextran sodium sulfate (DSS) was administered to establish the mice colitis model. Data of weight, length of colon, disease activity index (DAI), and macroscopic scores were collected. The flow cytometry was used to detect the infiltrations of T cells and DCs, the mean fluorescence intensity (MFI) of CD80, CD86, CD40, and major histocompatibility complex-II (MHC-II). And T cells proliferataion was measured by carboxyfluorescein diacetate succinimidyl ester (CFSE). The expression of cytokines (tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-10 (IL-10), interferon-γ (IFN-γ), interleukin -4 (IL-4)) and GPR174 mRNA were measured by Elisa, quantitative polymerase chain reaction (qPCR), and immunofluorescence. RNA of bone-marrow-derived dendritic cells (BMDCs) was extracted for sequencing. Adoptive transfer of BMDCs was administrated intravenously.ResultsGpr174-/- mice exposed to 3% DSS showed significant alleviation characterized by reduced loss of weight, more minor colon damage, and better DAI and macroscopic scores. The expression of pro-inflammatory cytokines (TNF-α, IL-6) decreased, while anti-inflammatory cytokine (IL-10) increased compared with WT mice. In vitro, Gpr174-/- BMDCs showed less maturity, with a declined expression of MHC-II, CD80, CD86 and reduced TNF-α, higher IL-10 after LPS stimulation. Gpr174-/- BMDCs were less capable of activating OT-II naïve CD4+ T cells than WT BMDCs and induced more Th0 cells to differentiate into Treg while less into Th1. Furthermore, the transcriptome sequencing analysis exhibited that Gpr174 participated in TNF-α (NF-κB) signaling, leukocyte transendothelial migration, and Th1/Th2 cell differentiation pathways. Adoptive transfer of Gpr174-/- BMDCs to WT mice ameliorated DSS-induced colitis.ConclusionOur study indicated that GPR174 was involved in the pathogenesis of IBD by regulating the maturation of the dendritic cells to maintain immune homeostasis. TNF-α (NF-κB) signaling pathway, leukocyte transendothelial migration, and Th1/Th2 cell differentiation pathways may be the target pathway.

Comprehensive Testing of Chemotherapy and Immune Checkpoint Blockade in Preclinical Cancer Models Identifies Additive Combinations

Antibodies that target immune checkpoints such as cytotoxic T lymphocyte antigen 4 (CTLA‐4) and the programmed cell death protein 1/ligand 1 (PD-1/PD-L1) are now a treatment option for multiple cancer types. However, as a monotherapy, objective responses only occur in a minority of patients. Chemotherapy is widely used in combination with immune checkpoint blockade (ICB). Although a variety of isolated immunostimulatory effects have been reported for several classes of chemotherapeutics, it is unclear which chemotherapeutics provide the most benefit when combined with ICB. We investigated 10 chemotherapies from the main canonical classes dosed at the clinically relevant maximum tolerated dose in combination with anti‐CTLA-4/anti-PD-L1 ICB. We screened these chemo-immunotherapy combinations in two murine mesothelioma models from two different genetic backgrounds, and identified chemotherapies that produced additive, neutral or antagonistic effects when combined with ICB. Using flow cytometry and bulk RNAseq, we characterized the tumor immune milieu in additive chemo-immunotherapy combinations. 5-fluorouracil (5-FU) or cisplatin were additive when combined with ICB while vinorelbine and etoposide provided no additional benefit when combined with ICB. The combination of 5-FU with ICB augmented an inflammatory tumor microenvironment with markedly increased CD8+ T cell activation and upregulation of IFNγ, TNFα and IL-1β signaling. The effective anti‐tumor immune response of 5-FU chemo-immunotherapy was dependent on CD8+ T cells but was unaffected when TNFα or IL-1β cytokine signaling pathways were blocked. Our study identified additive and non-additive chemotherapy/ICB combinations and suggests a possible role for increased inflammation in the tumor microenvironment as a basis for effective combination therapy.

TRIM47 is a novel endothelial activation factor that aggravates lipopolysaccharide-induced acute lung injury in mice via K63-linked ubiquitination of TRAF2.

Endothelial activation plays an essential role in the pathogenesis of sepsis-induced acute lung injury, however, the detailed regulatory mechanisms remain largely unknown. Here, we reported that TRIM47, an E3 ubiquitin ligase of the tripartite motif-containing protein family, was highly expressed in vascular endothelial cells. TRIM47-deficient mice were effectively resistant to lipopolysaccharide (LPS)-induced acute lung injury and death by attenuating pulmonary inflammation. TRIM47 was upregulated during TNFα-induced endothelial activation in vitro. Knockdown of TRIM47 in endothelial cells inhibited the transcription of multiple pro-inflammatory cytokines, reduced monocyte adhesion and the expression of adhesion molecules, and suppressed the secretion of IL-1β and IL-6 in endothelial cells. By contrast, overexpression of TRIM47 promoted inflammatory response and monocyte adhesion upon TNFα stimulation. In addition, TRIM47 was able to activate the NF-κB and MAPK signaling pathways during endothelial activation. Furthermore, our experiments revealed that TRIM47 resulted in endothelial activation by promoting the K63-linked ubiquitination of TRAF2, a key component of the TNFα signaling pathway. Taken together, our studies demonstrated that TRIM47 as a novel activator of endothelial cells, promoted LPS-induced pulmonary inflammation and acute lung injury through potentiating the K63-linked ubiquitination of TRAF2, which in turn activates NF-κB and MAPK signaling pathways to trigger an inflammatory response in endothelial cells.

Ethanol Consumption Promotes TNF-α Signaling Pathway in Rat Kidney: Rescue Effect of Curcumin

Ethanol Consumption Promotes TNF-α Signaling Pathway in Rat Kidney: Rescue Effect of Curcumin

Computational systems pharmacology analysis of Tong-Jing-Yi formula in the treatment of dysmenorrhea

Background: Tong-Jing-Yi (TJY) formula consists of Leonurus, fried Toosendan and processed Cyperus, etc. The therapeutic effect of TJY on dysmenorrhea has been clinically validated, but the underlying mechanism remains unclear. The present study aimed to explore the possible molecular targets of TJY and the potential mechanisms. Methods: The components of TJY formula were identified by ultra performance liquid chromatography–quadrupole-time of flight/mass spectrometry. SwissTargetPrediction database was used to predict the targets of TJY formula, and targets associated with primary dysmenorrhea were also collected through other databases. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were conducted. Results: A total of 91 compounds with identified structures were screened, including 3 groups of isomers. The results predicted 854 TJY formula-related targets and 363 disease-related targets. GO and KEGG analysis showed that the top 5 target genes were PIK3CA, AKT1, EGFR, AKT2 and CYP19A1. PI3K-Akt signaling, chemokine signaling, focal adhesion, and Rap1 signaling were ranked in the top 15 pathways. Conclusion: TJY formula might play roles in the treatment of dysmenorrhea underlying mechanisms relating to the involvement of TNF-α, interleukin and PI3K-Akt signaling pathway. Potential pathways have been identified that need mechanistic confirmation in a laboratory setting in the future.

Network Pharmacological Study on Mechanism of the Therapeutic Effect of Modified Duhuo Jisheng Decoction in Osteoporosis.

BackgroundModified Duhuo Jisheng Decoction (MDHJSD) is a traditional Chinese medicine prescription for the treatment of osteoporosis (OP), but its mechanism of action has not yet been clarified. This study aims to explore the mechanism of MDHJSD in OP through a combination of network pharmacology analysis and experimental verification.MethodsThe active ingredients and corresponding targets of MDHJSD were acquired from the Traditional Chinese Medicine System Pharmacology (TCMSP) database. OP-related targets were acquired from databases, including Genecards, OMIM, Drugbank, CTD, and PGKB. The key compounds, core targets, major biological processes, and signaling pathways of MDHJSD that improve OP were identified by constructing and analysing the relevant networks. The binding affinities between key compounds and core targets were verified using AutoDock Vina software. A rat model of ovariectomized OP was used for the experimental verification.ResultsA total of 100 chemical constituents, 277 targets, and 4734 OP-related targets of MDHJSD were obtained. Subsequently, five core components and eight core targets were identified in the analysis. Pathway enrichment analysis revealed that overlapping targets were significantly enriched in the tumour necrosis factor-alpha (TNF-α) signaling pathway, an inflammation signaling pathway, which contained six of the eight core targets, including TNF-α, interleukin 6 (IL-6), transcription factor AP-1, mitogen-activated protein kinase 3, RAC-alpha serine/threonine-protein kinase, and caspase-3 (CASP3). Molecular docking analysis revealed close binding of the six core targets of the TNF signaling pathway to the core components. The results of experimental study show that MDHJSD can protect bone loss, inhibit the inflammatory response, and downregulate the expression levels of TNF-α, IL-6, and CASP3 in ovariectomized rats.ConclusionThe mechanism of MDHJSD in the treatment of OP may be related to the regulation of the inflammatory response in the bone tissue.

HSP90 inhibitor RGRN-305 for oral treatment of plaque-type psoriasis: efficacy, safety and biomarker results in an open-label proof-of-concept study.

HSP90 is a downstream regulator of tumour necrosis factor (TNF)-α and interleukin (IL)-17A signalling and may therefore serve as a novel target in the treatment of psoriasis. This phase Ib proof-of-concept study was undertaken to evaluate the safety and efficacy of a novel HSP90 inhibitor (RGRN-305) in the treatment of plaque psoriasis. We conducted an open-label, single-arm, dose-selection, single-centre proof-of-concept study. Patients with plaque psoriasis were treated with 250 mg or 500 mg RGRN-305 daily for 12 weeks. Efficacy was evaluated clinically using the Psoriasis Area and Severity Index (PASI), body surface area (BSA), Physician's Global Assessment (PGA) scores and the Dermatology Life Quality Index (DLQI). Skin biopsies collected at baseline and at 4, 8 and 12 weeks after initiation of treatment were used for immunohistochemical staining and for gene expression analysis. Safety was monitored via laboratory tests, vital signs, electrocardiogram and physical examinations. Six of the 11 patients who completed the study responded to RGRN-305 with a PASI improvement between 71% and 94%, whereas five patients were considered nonresponders with a PASI response < 50%. No severe adverse events were reported. Four of seven patients treated with 500 mg RGRN-305 daily experienced a mild-to-moderate exanthematous drug-induced eruption owing to the study treatment. Two patients chose to discontinue the study because of this exanthematous eruption. RGRN-305 treatment resulted in pronounced inhibition of the IL-23, TNF-α and IL-17A signalling pathways and normalization of both histological changes and psoriatic lesion gene expression profiles in patients who responded to treatment. Treatment with RGRN-305 showed acceptable safety, especially in the low-dose group, and was associated with clinically meaningful improvement in a subset of patients with plaque psoriasis, indicating that HSP90 may serve as a novel future target in psoriasis treatment.

Transcriptomic Profiling of MSI-H/dMMR Gastrointestinal Tumors to Identify Determinants of Responsiveness to Anti-PD-1 Therapy.

Transcriptomic profiling was performed for microsatellite instability-high (MSI-H)/mismatch repair-deficient (dMMR) gastrointestinal tumors to determine the predictors of response to PD-1 blockade. Thirty-six patients with MSI-H/dMMR gastrointestinal tumors, including gastric cancer, colorectal cancer, cholangiocarcinoma, small intestine cancer, and pancreatic cancer, being treated with PD-1 blockade were analyzed. We conducted the transcriptomic analysis of gastrointestinal tumors using RNA sequencing data, including the consensus molecular subtypes (CMS) of colorectal cancer. Gene set enrichment analysis (GSEA) demonstrated that non-responders had upregulations of epithelial-mesenchymal transition, angiogenesis, hypoxia, mTORC1, TNF-α, KRAS, Wnt/β-catenin, TGF-β, and various metabolism-related signaling pathways. Meanwhile, the IFNγ pathway was enriched in responders. On the basis of the leading-edge analysis of GSEA, VEGF-A was significantly correlated with enriched pathways in non-responders. Patients with high VEGF-A expression, compared with those with low expression, had significantly shorter progression-free survival [PFS; median 4.8 months vs. not reached (NR), P = 0.032] and overall survival (median 11.1 months vs. NR, P = 0.045). Among 13 patients with colorectal cancer evaluable for CMS classification, the objective response rate was 100%, 0%, 0%, and 16.7% in CMS1, CMS2, CMS3, and CMS4, respectively. Patients with CMS1 had significantly longer PFS (NR vs. 4.8 months, P = 0.017) than those with CMS2, CMS3, or CMS4. Several transcriptomic features, including CMS classification and related genes, were associated with response to PD-1 blockade in MSI-H/dMMR gastrointestinal tumors. These findings can help develop predictive biomarkers or combination immunotherapies.

Exosomes Secreted from Adipose-Derived Stem Cells Are a Potential Treatment Agent for Immune-Mediated Alopecia.

Background Alopecia has become an exceedingly prevalent dermatological disorder. Etiologically, infection (bacterial and fungal infection), inflammation, and immune dysregulation are the main causes of immune-mediated hair loss. Treating hair loss has remained challenging as the available therapies are limited. Exosomes from adipose-derived stem cells (ADSC-Exos) have been used for treating neurodegenerative diseases and autoimmune diseases and in wound-healing treatments. However, the function and mechanism of ADSC-Exos in alopecia treatment remain unclear. This study is aimed at investigating the effects of ADSC-Exos on hair growth in vitro and in vivo for potentially treating immune-mediated alopecia and further exploring the underlying mechanism. Methods Cell proliferation, migration, and apoptosis of dermal papilla cells (DPCs) that were treated with ADSC-Exos were detected using the cell counting kit-8 (CCK-8) assay, scratch wound-healing assay, and flow cytometry assay, respectively. A C57BL/6 hair-depilated mouse model was established in vivo; then, ADSC-Exos were subcutaneously injected alone or in combined with minoxidil. The effects of ADSC-Exos on hair growth, pathological changes, and the related mechanism were investigated by HE staining, quantitative real‐time PCR (qRT-PCR), western blotting, and RNA sequencing (RNA-seq). Results ADSC-Exos significantly promoted DPC proliferation and migration while also reducing apoptosis. In addition, compared with the control group, ADSC-Exos-treated mice had better hair growth, more hair follicles (HFs) and thicker dermis. RNA-seq revealed that the miR-22 and TNF-α signaling pathways were markedly downregulated in DPCs after ADSC-Exos treatment. In addition, according to qRT-PCR and western blotting results, the Wnt/β-catenin signaling pathway was activated in the skin of ADSC-Exos-treated mice. Conclusion ADSC-Exos therapy positively affected the promotion of hair regrowth by regulating miR-22, the Wnt/β-catenin signaling pathway, and the TNF-α signaling pathway, implying that ADSC-Exos could be a promising cell-free therapeutic strategy for immune-mediated alopecia.

Network Pharmacology and Molecular Docking Study of the Chinese Miao Medicine Sidaxue in the Treatment of Rheumatoid Arthritis.

PurposeThis study aimed to investigate the molecular mechanisms of Compound Sidaxue (SX), a prescription of Chinese Miao medicine, in treating rheumatoid arthritis (RA) using network pharmacology and in vivo experimental approaches.MethodsNetwork pharmacology was adopted to detect the active components of four Traditional Chinese herbal medicine (TCM) of SX, and the key targets and signaling pathways in the treatment of RA were predicted, and the key components and targets were screened for molecular docking. The predicted targets and pathways were validated in bovine type II collagen and incomplete Freund’s adjuvant emulsifier-induced rat RA model.ResultsIn this study, we identified 33 active components from SX, predicted to act on 44 RA-associated targets by network pharmacology. PPI network demonstrated that TNF-α, VEGF-A, IL-2, IL-6, AKT, PI3K, STAT1 may serve as the key targets of SX for the treatment of RA. The main functional pathways involving these key targets include PI3K-AKT signaling pathway, TNF signaling pathway, NF-κB signaling pathway. Molecular docking analysis found that the active components β-amyrin, cajanin, eleutheroside A have high affinity for TNF-α, VEGFA, IL-2, AKT, and PI3K, etc. SX can improve joint swelling in Collagen-induced arthritis (CIA) rats, reduce inflammatory cell infiltration and angiogenesis in joint synovial tissue, and down-regulate IL-2, IL-6, TNF-α, VEGF, PI3K, AKT, p-AKT, NF-κBp65, the expression of p-NF-κBp65, STAT1, and PTGS2 are used to control the exacerbation of inflammation and alleviate the proliferation of synovial pannus, and at the same time play the role of cartilage protection to achieve the effect of treating RA.ConclusionThrough a network pharmacology approach and animal study, we predicted and validated the active compounds of SX and their potential targets for RA treatment. The results suggest that SX can markedly alleviate CIA rat by modulating the VEGF/PI3K/AKT signaling pathway, TNF-α signaling pathway, IL/NF-κB signaling pathway.

Vancomycin Pretreatment on MPTP-Induced Parkinson's Disease Mice Exerts Neuroprotection by Suppressing Inflammation Both in Brain and Gut.

A growing body of evidence implies that gut microbiota was involved in pathogenesis of Parkinson's disease (PD), but the mechanism is still unclear. The aim of this study is to investigate the effects of antibiotics pretreatment on the 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced PD mice. In this study, vancomycin pretreatment was given by gavage once daily with either vancomycin or distilled water for 14days to mice, then mice were administered with MPTP (20mg/kg, i.p) for four times in one day to establish an acute PD model. Results show that vancomycin pretreatment significantly improved motor dysfunction of mice in pole and traction tests. Although vancomycinpretreatment had no effect on dopamine (DA) or the process of DA synthesis, it inhibited the metabolism of DA by suppressing the expression of striatal monoamine oxidase B (MAO-B). Furthermore, vancomycin pretreatment reduced the number of astrocytes and microglial cells in the substantia nigra pars compacta (SNpc) to alleviate neuroinflammation, decreased the expression of TLR4/MyD88/NF-κB/TNF-α signaling pathway in both brain and gut. Meanwhile, vancomycin pretreatment changed gut microbiome composition and the levels of fecal short chain fatty acids (SCFAs). The abundance of Akkermansia and Blautia increased significantly after vancomycin pretreatment, which might be related to inflammation and inhibition of TLR4 signaling pathway. In summary, these results demonstrate that the variation of gut microbiota and its metabolites induced by vancomycin pretreatment might decrease dopamine metabolic rate and relieve inflammation in both gut and brain via the microbiota-gut-brain axis in MPTP-induced PD mice. The neuroprotection of vancomycin pretreatment on MPTP-induced Parkinson's disease mice The alterations of gut microbiota and SCFAs induced by vancomycin pretreatment might not only improve motor dysfunction, but also decrease dopamine metabolism and relieve inflammation in both brain and gut via TLR4/MyD88/NF-κB/TNF-α pathway in MPTP-induced PD mice.

Identification of Master Regulators Driving Disease Progression, Relapse, and Drug Resistance in Lung Adenocarcinoma.

Backgrounds: Lung cancer is the leading cause of cancer related death worldwide. Current treatment strategies primarily involve surgery, chemotherapy, radiotherapy, targeted therapy, and immunotherapy, determined by TNM stages, histologic types, and genetic profiles. Plenty of studies have been trying to identify robust prognostic gene expression signatures. Even for high performance signatures, they usually have few shared genes. This is not totally unexpected, since a prognostic signature is associated with patient survival and may contain no upstream regulators. Identification of master regulators driving disease progression is a vital step to understand underlying molecular mechanisms and develop new treatments. Methods: In this study, we have utilized a robust workflow to identify potential master regulators that drive poor prognosis in patients with lung adenocarcinoma. This workflow takes gene expression signatures that are associated with poor survival of early-stage lung adenocarcinoma, EGFR-TKI resistance, and responses to immune checkpoint inhibitors, respectively, and identifies recurrent master regulators from seven public gene expression datasets by a regulatory network-based approach. Results: We have found that majority of the master regulators driving poor prognosis in early stage LUAD are cell-cycle related according to Gene Ontology annotation. However, they were demonstrated experimentally to promote a spectrum of processes such as tumor cell proliferation, invasion, metastasis, and drug resistance. Master regulators predicted from EGFR-TKI resistance signature and the EMT pathway signature are largely shared, which suggests that EMT pathway functions as a hub and interact with other pathways such as hypoxia, angiogenesis, TNF-α signaling, inflammation, TNF-β signaling, Wnt, and Notch signaling pathways. Master regulators that repress immunotherapy are enriched with MYC targets, E2F targets, oxidative phosphorylation, and mTOR signaling. Conclusion: Our study uncovered possible mechanisms underlying recurrence, resistance to targeted therapy, and immunotherapy. The predicted master regulators may serve as potential therapeutic targets in patients with lung adenocarcinoma.