Alcohol inducing macrophage M2b polarization in colitis by modulating the TRPV1-MAPK/NF-κB pathways.

BACKGROUND: Macrophages exhibit different phenotypes in inflammatory bowel disease (IBD) and promote inflammation or tissue repair depending on their polarization state. Alcohol is a widely used solvent in pharmaceutical formulations, and its consumption is associated with an increased risk of colitis; however, its effects on macrophages in IBD remain poorly understood. PURPOSE: This study aimed to investigate the effect of alcohol on macrophages in dextran sodium sulfate (DSS)-induced colitis and understand the underlying mechanisms. METHODS: DSS-treated C57BL/6 mice were exposed to varying concentrations of alcohol, transient receptor potential vanilloid 1 (TRPV1) antagonist, and 5-aminosalicylic acid. The distal colon was resected, fixed, stained, and histologically analyzed, through hematoxylin and eosin (H&E) staining and immunofluorescence staining. Ratio [Ca2+]i measurements, western blotting, quantitative polymerase chain reaction, cytokine measurements, and RNA sequencing analyses were also performed. Peritoneal macrophages and RAW264.7 cells were used for in vitro experiments, and various assays were performed to evaluate cellular responses, gene expression, and signaling pathways. RESULTS: Alcohol exacerbated DSS-treated mice colitis and promoted the secretion of various inflammatory cytokines from colonic macrophages. Alcohol enhances the calcium ion influx induced by lipopolysaccharide (LPS) in peritoneal macrophages, while the TRPV1 antagonist capsazepine (CPZ) inhibits LPS- and/or alcohol- induced calcium influx in macrophages. Alcohol and LPS activate the MAPK/P38, MAPK/ERK, and NF-κB signaling pathways and induce the macrophage M2b polarization, resulting in the increased expression level of inflammatory cytokines such as Tnf, Il1b, and Il10. Additionally, CPZ can inhibit the facilitatory effects of alcohol or LPS on the abovementioned pathways and inflammatory factors, reversing macrophage M2b polarization and promoting alcohol-induced colitis. The inhibition of nucleotide binding oligomerization domain containing 2 (NOD2) partially suppressed the alcohol and LPS effects on macrophages. CONCLUSION: Alcohol exacerbates experimental colitis and induces M2b polarization of macrophage via TRPV1-MAPK/NF-κB. Our study provides new insights into the potential therapeutic targets for IBD treatment by elucidating the role of TRPV1 in alcohol-exacerbated colitis, using CPZ as a potential therapeutic option. The identification of transient receptor potential ankyrin subtype 1 (TRPA1) as a therapeutic target expands the scope of future research.

Impact of Hepatic Pedicle Clamping on Long-Term Survival Following Hepatectomy for Hepatocellular Carcinoma: Stratified Analysis Based on Intraoperative Blood Transfusion Status.

BACKGROUND: Hepatic pedicle clamping (HPC) is frequently utilized during hepatectomy to reduce intraoperative bleeding and diminish the need for intraoperative blood transfusion (IBT). The long-term prognostic implications of HPC following hepatectomy for hepatocellular carcinoma (HCC) remain under debate. This study aims to elucidate the association between HPC and oncologic outcomes after HCC resection, stratified by whether IBT was administered. PATIENTS AND METHODS: Prospectively collected data on patients with HCC who underwent curative resection from a multicenter database was studied. Patients were stratified into two cohorts on the basis of whether IBT was administered. The impact of HPC on long-term overall survival (OS) and recurrence-free survival (RFS) between the two cohorts was assessed by univariable and multivariable Cox regression analyses. RESULTS: Of 3362 patients, 535 received IBT. In the IBT cohort, using or not using HPC showed no significant difference in OS and RFS outcomes (5-year OS and RFS rates 27.9% vs. 24.6% and 13.8% vs. 12.0%, P = 0.810 and 0.530). However, in the non-IBT cohort of 2827 patients, the HPC subgroup demonstrated significantly decreased OS (5-year 45.9% vs. 56.5%, P < 0.001) and RFS (5-year 24.7% vs. 33.3%, P < 0.001) when compared with the subgroup without HPC. Multivariable Cox regression analysis identified HPC as an independent risk factor of OS and RFS [hazard ratios (HR) 1.16 and 1.12, P = 0.024 and 0.044, respectively] among patients who did not receive IBT. CONCLUSIONS: The impact of HPC on the oncological outcomes following hepatectomy for patients with HCC differed significantly whether IBT was administered, and HPC adversely impacted on long-term survival for patients without receiving IBT during hepatectomy.

The Ongoing Necessity of Sentinel Lymph Node Biopsy for cT1-2N0 Breast Cancer Patients.

Background: Recent clinical trials attempt to determine whether it is appropriate to omit axillary lymph node surgery in patients with cT1-2N0 breast cancer. The study aimed to investigate the true extent of axillary node disease in patients with clinically negative nodes and explore the differences between negative axillary ultrasound (AUS-cN0) and suspicious axillary ultrasound with negative fine-needle aspiration (FNA-cN0). Methods: Pathologically identified T1-2 invasive breast cancer patients with clinically negative nodes were retrospectively analyzed at our center between January 2019 and December 2022. Patients who received any systematic treatment before surgery were excluded from this study. Results: A total of 538 patients were enrolled in this study. 134 (24.9%) patients had pathologically positive nodes, and 404 (75.1%) patients had negative nodes. Univariate analysis revealed that tumor size, T stage, Ki67 level, and vascular invasion (VI) were strongly associated with pathological axillary lymph node positivity. In multivariate analysis, VI was the only independent risk factor for node positivity in patients with cT1-2N0 disease (OR: 3.723, confidence interval [CI]: 2.380-5.824, p < 0.001). Otherwise, pathological node positivity was not significantly different between AUS-cN0 and FNA-cN0 groups (23.4% vs. 28.8%, p = 0.193). However, the rate of high nodal burden (≥3 positive nodes) was significantly higher in FNA-cN0 group. Further investigation revealed that FNA-cN0 and VI were independently associated with a high nodal burden (OR: 2.650, CI: 1.081-6.496, p = 0.033; OR: 3.521, CI: 1.249-9.931, p = 0.017, respectively). Conclusions: cT1-2 breast cancer patients with clinically negative axillary lymph nodes may have pathologically positive lymph nodes and even a high nodal burden. False negatives in AUS and AUS-guided FNA should not be ignored, and sentinel lymph node biopsy remains an ongoing necessity for cT1-2N0 breast cancer patients.

TAOK3 Facilitates Esophageal Squamous Cell Carcinoma Progression and Cisplatin Resistance Through Augmenting Autophagy Mediated by IRGM.

Esophageal squamous cell carcinoma (ESCC) is one of the deadliest cancers because of its robust aggressive phenotype and chemoresistance. TAO kinase belongs to mitogen-activated protein kinases, which mediate drug resistance in multiple cancers. However, the role of TAO kinase in ESCC progression and chemoresistance has never been explored. Here, it is reported that TAOK3 augments cell autophagy and further promotes ESCC progression and chemoresistance. Mechanistically, TAOK3 phosphorylates KMT2C at S4588 and strengthens the interaction between KMT2C and ETV5. Consequently, the nuclear translocation of KMT2C is increased, and the transcription of autophagy-relevant gene IRGM is further upregulated. Additionally, the inhibitor SBI-581 can significantly suppress cell autophagy mediated by TAOK3 and synergizes with cisplatin to treat ESCC in vitro and in vivo.

Homeobox A7 promotes esophageal squamous cell carcinoma progression through C-C motif chemokine ligand 2-mediated tumor-associated macrophage recruitment.

Homeobox A7 (HOXA7) plays essential roles in multiple malignancies and was reported to be overexpressed in esophageal squamous cell carcinoma (ESCC). However, its functions in the ESCC tumor microenvironment remain to be explored. In this study, we showed that HOXA7 was overexpressed in ESCC among HOXA family members and correlated with tumor-associated macrophage (TAM) infiltration both in The Cancer Genome Atlas database and ESCC clinical samples. Moreover, transactivation of C-C motif chemokine ligand 2 (CCL2) by HOXA7 was identified (real-time quantitative PCR [RT-qPCR], western blot analysis, ELISA, and ChIP-qPCR), which was detected to drive chemotaxis and M2 polarization of macrophages both in vitro (Transwell assay) and in vivo (xenograft tumors models). In addition, CCL2 triggers macrophage expression of epidermal growth factor (EGF) (RT-qPCR and ELISA), which promotes tumor proliferation and metastasis by activating its receptor EGFR. In addition, EGF-induced ESCC cell proliferation and migration can be abrogated by HOXA7 knockdown (CCK-8 proliferation assay, EdU fluorescence, and Transwell assay). These results indicate a novel mechanistic role of HOXA7 in the cross-talk between ESCC and TAMs, which could be an underlying therapeutic target for ESCC.

CD1C is associated with breast cancer prognosis and immune infiltrates.

BACKGROUND: The tumor microenvironment (TME) in breast cancer plays a vital role in occurrence, development, and therapeutic responses. However, immune and stroma constituents in the TME are major obstacles to understanding and treating breast cancer. We evaluated the significance of TME-related genes in breast cancer. METHODS: Invasive breast cancer (BRCA) samples were retrieved from the TCGA and GEO databases. Stroma and immune scores of samples as well as the proportion of tumor infiltrating immune cells (TICs) were calculated using the ESTIMATE and CIBERSORT algorithms. TME-related differentially expressed genes (DEGs) were analyzed by a protein interaction (PPI) network and univariate Cox regression to determine CD1C as a hub gene. Subsequently, the prognostic value of CD1C, its response to immunotherapy, and its mechanism in the TME were further studied. RESULTS: In BRCA, DEGs were determined to identify CD1C as a hub gene. The expression level of CD1C in BRCA patients was verified based on the TCGA database, polymerase chain reaction (PCR) results, and western blot analysis. Immunohistochemical staining (IHC) results revealed a correlation between prognosis, clinical features, and CD1C expression in BRCA. Enrichment analysis of GSEA and GSVA showed that CD1C participates in immune-associated signaling pathways. CIBERSORT showed that CD1C levels were associated with tumor immune infiltrating cells (TILs), such as different kinds of T cells. Gene co-expression analysis showed that CD1C and the majority of immune-associated genes were co-expressed in BRCA. In renal cell carcinoma, patients with a high expression of CD1C had a better immunotherapy effect. CONCLUSION: CD1C is an important part of the TME and participates in immune activity regulation in breast tumors. CD1C is expected to become a prognostic marker and a new treatment target for breast cancer.

Cloning, Expression, and Characterization of a GHF 11 Xylanase from Alteromonas macleodii HY35 in Escherichia coli.

A xylanase gene xynZT-1 from Alteromonas macleodii HY35 was cloned and expressed in Escherichia coli (E. coli). The sequencing results showed that the ORF of xynZT-1 was 831 bp. The xylanase DNA sequence encoded a 29 amino acids (aa) signal peptide and a 247-aa mature peptide. The XynZT-1 has been a calculated molecular weight (MW) of 27.93 kDa, isoelectric point (pI) of 5.11 and the formula C1266H1829N327O384S5. The amino acid sequence of the xynZT-1 had a high similarity with that of glycosyl hydrolase family 11 (GHF11) reported from other microorganisms. The DNA sequence encoding mature peptide was subcloned into pET-28a(+) expression vector. The resulted plasmid pET-28a-xynZT-1 was transformed into E. coli BL21(DE3), and the recombinant strain BL21(DE3)/xynZT-1 was obtained. The optimum temperature and pH of the recombinant XynZT-1 were 45 ℃ and 5.0, respectively.

Long non-coding RNA SNHG8 enhances triple-negative breast cancer cell proliferation and migration by regulating the miR-335-5p/PYGO2 axis.

BACKGROUND: Growing evidence has demonstrated that long non-coding RNAs (lncRNAs) can function as modulators in the development of triple-negative breast cancer (TNBC). However, the function of lncRNA small nucleolar RNA host gene 8 (SNHG8) in TNBC remains unclear. Therefore, our study aimed at investigating the role of SNHG8 in the proliferation and migration of TNBC cells. METHODS: SNHG8 expression was evaluated using RT-qPCR assay. Cell proliferation and migration were assessed by EdU, colony formation and Transwell assays. The levels of proteins related to EMT process were examined by western blot assay. The interaction among SNHG8, miR-335-5p and pygopus family PHD finger 2 (PYGO2) was detected by RIP assay, RNA pull down assay and luciferase reporter assay. RESULTS: SNHG8 expression was significantly up-regulated in TNBC cells. SNHG8 silencing obviously inhibited TNBC cell proliferation, migration and EMT process. Moreover, SNHG8 acted as a sponge to sequester miR-335-5p in TNBC cells. Besides, PYGO2 was proven as a target gene of miR-335-5p, and SNHG8 promoted TNBC cell proliferation, migration and EMT process through regulating miR-335-5p and PYGO2. CONCLUSIONS: Totally, our study indicated that SNHG8 promoted TNBC cell proliferation and migration by regulating the miR-335-5p/PYGO2 axis.

TNF-α/NF-κB signaling epigenetically represses PSD4 transcription to promote alcohol-related hepatocellular carcinoma progression.

BACKGROUND: Chronic alcohol consumption is more frequently associated with advanced, aggressive hepatocellular carcinoma (HCC) tumors. Alcohol adversely impacts ER/Golgi membrane trafficking and Golgi protein N-glycosylation in hepatocytes; these effects have been attributed (in part) to dysregulated adenosine diphosphate-ribosylation factor (ARF) GTPase signaling. Here, we investigated the role of the ARF GTPase guanine exchange factor PSD4 in HCC progression. METHODS: R-based bioinformatics analysis was performed on publicly available array data. Modulating gene expression was accomplished via lentiviral vectors. Gene expression was analyzed using quantitative real-time PCR and immunoblotting. PSD4 promoter methylation was assessed using quantitative methylation-specific PCR. Phospho-p65(S276)/DNMT1 binding to the PSD4 promoter was analyzed via chromatin immunoprecipitation. We constructed ethanol/DEN-induced and DEN only-induced transgenic murine models of HCC. RESULTS: We identified PSD4 as a hypermethylated, suppressed gene in alcohol-related HCC tumors; however, PSD4 was not dysregulated in all-cause HCC tumors. Certain HCC cell lines also displayed varying degrees of PSD4 downregulation. PSD4 overexpression or knockdown decreased and increased cell migration and invasiveness, respectively. Mechanistically, PSD4 transcription was repressed by TNF-α-induced phospho-p65(S276)'s recruitment of DNA methyltransferase 1 (DNMT1), resulting in PSD4 promoter methylation. PSD4 inhibited pro-EMT CDC42 activity, resulting in downregulation of E-cadherin and upregulation of N-cadherin and vimentin. Hepatocyte-specific PSD4 overexpression reduced ethanol/DEN-induced HCC tumor progression and EMT marker expression in vivo. CONCLUSIONS: PSD4 is a hypermethylated, suppressed gene in alcohol-related HCC tumors that negatively modulated pro-EMT CDC42 activity. Furthermore, we present a novel phospho-NF-κB p65(S276)/DNMT1-mediated promoter methylation mechanism by which TNF-α/NF-κB signaling represses PSD4 transcription in HCC cells.

Gene expression signature for detection of gastric cancer in peripheral blood.

Gastric cancer (stomach cancer) is the fifth most common malignancy and the third leading cause of cancer-associated mortality worldwide. Identifying gastric cancer patients at an early and curable stage of the disease is essential if mortality rates for this disease are to decrease. A non-invasive blood-based test that is an indicator of gastric cancer risk would likely be of benefit in identifying gastric cancer patients at an early stage, and such a test may enhance clinical decision making. This study identified a four-gene expression signature in peripheral blood samples associated with gastric cancer. A total of 216 blood samples were collected, including those from 36 gastric cancer patients, 55 healthy controls and 125 patients with other carcinomas, and gene expression profiles were examined using an Affymetrix Gene Profiling microarray. Blood gene expression profiles were compared between patients with stomach cancer, healthy controls and patients affected with other malignancies. A four-gene panel was identified comprising purine-rich element binding protein B, structural maintenance of chromosomes 1A, DENN/MADD domain containing 1B and programmed cell death 4. The four-gene panel discriminated gastric cancer with an area under the receiver-operating-characteristic curve of 0.99, an accuracy of 95%, sensitivity of 92% and specificity of 96%. The non-invasive nature of the blood test, together with the relatively high accuracy of the four-gene panel may assist physicians in gastric cancer screening decision making.

Risk factors of central lymph node metastasis in papillary thyroid carcinoma: A retrospective cohort study.

OBJECTIVE: The aim of this study was to explore the risk factors that were associated with central lymph node metastases (CLNM) in papillary thyroid carcinoma (PTC) patients. METHODS: A total of 180 patients with PTC who underwent surgery in our hospital between January 2014 and December 2016 were identified retrospectively. The relationship between clinicopathological factors and CLNM were analyzed by univariate and multivariate logistic regression. RESULTS: The incidence of CLNM was 67.8% (122/180) in PTC patients. Univariate analysis showed that multifocality (p = 0.002), HT (p < 0.001) and LVI (p < 0.001) were significant associated with CLNM. No significant associations were found between factors and CLNM, including age, gender, tumor size and ETE. Multivariate logistic regression analysis showed that multifocality (p = 0.011), HT (p < 0.001) and LVI (p < 0.001) were independent predictors of CLNM in PTC patients. CONCLUSIONS: Our study identified several independent risk factors predicting CLNM in PTC patients, such as multifocality, HT and LVI. The CLNM is very common in PTC patients, and routine prophylactic central neck dissection (PCND) may recommended in PTC patients with those risk factors of CLNM.

[Equol induced apoptosis of human breast cancer MDA-MB-231 cell by inhibiting the expression of nuclear factor-kappaB].

OBJECTIVE: To study the inhibition effect of equol on MDA-MB-231 cells, a human breast cancer cell line with estrogen-independent receptor. METHODS: Cultured MDA-MB-231 cells were treated with different contents of equol. The cell viability was assessed with MTT method. The apoptosis was detected by flow cytometry and immunofluorescence. Western Blot was used to assess the expression of NF-kappaB. RESULTS: The growth of MDA-MB-231 cells was inhibited by equol in a does-and time-dependent manner. The apoptosis of MDA-MB-231 cells induced by equol was the result of inhibiting the expression of NF-kappaB. CONCLUSION: The apoptosis of MDA-MB-231 cells might be the reduced expression of NF-kappaB induced by equol.