Adjacent Non-tumour Tissues Research Articles

The Association of Helicobacter pylori Infection and Virulence Factors in Gastric Cancer in Thi-Qar, Iraq

Background: The significance of Helicobacter pylori (H. pylori) infection in developing gastric cancer has been increasingly recognized. However, a noticeable gap exists in the literature regarding the prevalence of key virulence factors, particularly in Iraq. Therefore, Our study aimed to investigate the prevalence of H. pylori infection and the presence of virulence factors, namely the CagA and VacA genes, in gastric cancer patients. Methods: A case-control study was conducted on 45 gastric cancer patients (38 males and 7 females) from September 2020 to February 2021. During routine endoscopic procedures, gastric biopsy specimens were collected from the tumour and adjacent non-tumour tissues. The presence of H. pylori was detected using quantitative real-time PCR, and the virulence factors CagA and vacA were identified using specific primers. Results: H. pylori infection was detected in a significant 85% (38/45) of the patients, with a higher prevalence in males 89.5% (34/38) compared to females 10.5% (4/38) (P=0.021) (Chi-square test). No significant age-related differences were observed. The CagA gene was present in a substantial 58.8% (22/38) of H. pylori-positive patients, predominantly in tumor tissues at 81.81% (18/22), while the vacA gene was found in only one patient. These results underscore the importance of our research in understanding the prevalence of H. pylori infection and the presence of virulence factors in gastric cancer patients. Conclusion: The high prevalence of H. pylori infection and the significant presence of the CagA virulence factor in tumour tissues underscore the bacterium’s role and risk factors in gastric carcinogenesis in Thi-Qar, Iraq.

Biglycan promotes tumour proliferation and invasion in human colon cancers.

To investigate the role of biglycan (BGN) in colon cancer progression. The association between BGN mRNA levels and the survival time of patients with colon cancer was analysed by referencing data from the Gene Expression Omnibus and The Cancer Genome Atlas (TCGA). Gene Set Enrichment Analysis (GSEA) was conducted to explore gene sets and pathways associated with BGN. In vitro analyses were undertaken in HCT116 cells to analyse cell proliferation, migration, invasion, cell cycle control and apoptosis. GSEA, Western blot analysis and small interfering RNAs were used to explore the molecular mechanisms of BGN in colon cancer cells. Analysis of the pairs of tissues from colon cancer patients showed that BGN protein levels were higher in colon cancer tissues compared with adjacent non-tumour tissues. High BGN levels were significantly associated with shorter survival times. BGN knockdown inhibited cell proliferation, migration and invasion; and induced cell cycle arrest in the G0/G1 phase and promoted apoptosis in HCT116 cells. GSEA found that BGN might affect tumour progression via the mitogen-activated protein kinase (MAPK) signalling pathway. BGN might promote colon cancer progression via the MAPK signalling pathway and could be a potential target for future novel therapeutic strategies.

Existence and distribution of the microbiome in tumour tissues of children with hepatoblastoma

Cancer microbiota have recently been demonstrated in several cancer types. The microbiome enhances inflammation in the cancer microenvironment and affects the disease pathology by regulating tumourigenesis, cancer progression, and chemotherapy resistance. Hepatoblastoma (HB), the most common childhood malignant tumour, is a malignant embryonic tumour. However, the pathogenesis and molecular basis of HB remain poorly understood. In this study, to explore the existence and distribution of the microbiome in tumour tissues and adjacent non-tumour tissues of children with HB, we mainly performed 16S rDNA sequencing, and the results showed that the diversity and abundance of the microbiome in children with HB were significantly different between HB tumours and adjacent non-tumour tissues (p < 0.01). At the phylum level, the dominant microbiome in the tumour tissues were Proteobacteria, Bacteroidetes, and Firmicutes. At the genus level, Ruminococcus was more abundant in HB tumours than in the adjacent non-tumour tissues. Simultaneously, the abundances of Bacteroides, Parabacteroides, Lachnospiracea-NK4A136, and Alistipes in HB tumours were lower than those in the adjacent non-tumour tissues. In addition, Romboutsia strongly correlated with alpha-fetoprotein, an important indicator of HB. Sphingomonas was abundant in primary HB tumours, whereas Oscillibacter and Pandoraea were abundant in metastatic HB tumours. However, whether these bacteria are associated with HB needs further evaluation. Therefore, we identified the microbiome that correlated with the occurrence and development of HB. Ruminococcus and Romboutsia were identified as potential bacterial markers of HB tumours. To conclude, we found that HB also contains cancer microbiome, and it is necessary to shed light on the microbiome characteristics of HB in the future.

Integrative transcriptome analysis identifies a crotonylation gene signature for predicting prognosis and drug sensitivity in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) stands as the most prevalent and treatment-resistant malignant tumour, characterized by a dismal prognosis. Croton acylation (CA) has recently gained attention as a critical factor in cancer pathogenesis. This study sought to rapidly identify prognostic features of HCC linked to CA. Differential analysis was conducted between tumour tissues and adjacent non-tumour tissues in the TCGA-LIHC and GSE76427 datasets to uncover differentially expressed genes (DEG1 and DEG2). The intersection of DEG1 and DEG2 highlighted DEGs with consistent expression patterns. Single-sample gene set enrichment analysis scores were calculated for 18 lysine crotonylation-related genes (LCRGs) identified in prior research, showing significant differences between tumour and normal groups. Subsequently, weighted gene co-expression network analysis was employed to identify key module genes correlated with the LCRG score. Candidate genes were identified by overlapping consistently expressed DEGs with key module genes. Prognostic features were identified, and risk scores were determined via regression analysis. Patients were categorized into risk groups based on the optimal cutoff value. Gene set enrichment analysis (GSEA) and immunoassays were also performed. The prognostic features were further validated using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). A total of 88 candidate genes were identified from 1179 consistently expressed DEGs and 4200 key module genes. Seven prognostic features were subsequently identified: TMCO3, RAP2A, ITGAV, ZFYVE26, CHST9, HMGN4, and KLHL21. GSEA revealed that DEGs between risk groups were primarily associated with chylomicron metabolism, among other pathways. Additionally, activated CD4+ T cells demonstrated the strongest positive correlation with risk scores, and most immune checkpoints showed significant differences between risk groups, with ASXL1 exhibiting the strongest correlation with risk scores. The Tumour Immune Dysfunction and Exclusion score was notably higher in the high-risk group. Moreover, in both the TCGA-LIHC and ICGC-LIRI-JP datasets, the expression of other prognostic features was elevated in tumour tissues, with the exception of CHST9. RT-qPCR confirmed the increased expression of TMCO3, RAP2A, ITGAV, ZFYVE26, and HMGN4. This study establishes a risk model for HCC based on seven crotonylation-associated prognostic features, offering a theoretical framework for the diagnosis and treatment of HCC.

PUS7-dependent pseudouridylation of ALKBH3 mRNA inhibits gastric cancer progression.

RNA pseudouridylation is a critical post-transcriptional modification that influences gene expression and impacts various biological functions. Despite its significance, the role of mRNA pseudouridylation in cancer remains poorly understood. This study investigates the impact of pseudouridine synthase 7 (PUS7)-mediated pseudouridylation of Alpha-ketoglutarate-dependent Dioxygenase alkB Homolog 3 (ALKBH3) mRNA in gastric cancer. Immunohistochemistry and Western blotting were used to assess PUS7 protein levels in human gastric cancer tissues. The relationship between PUS7 and gastric cancer progression was examined using 3D colony formation assays and subcutaneous xenograft models. Real-time quantitative PCR (RT-qPCR), Western blotting, and polysome profiling assays were conducted to investigate how PUS7 regulates ALKBH3. A locus-specific pseudouridine (Ψ) detection assay was used to identify Ψ sites on ALKBH3 mRNA. Our findings indicate a significant reduction of PUS7 in gastric cancer tissues compared to adjacent non-tumour tissues. Functional analyses reveal that PUS7 inhibits gastric cancer cell proliferation and tumour growth via its catalytic activity. Additionally, PUS7 enhances the translation efficiency of ALKBH3 mRNA by modifying the U696 site with pseudouridine, thereby attenuating tumour growth. Importantly, ALKBH3 functions as a tumour suppressor in gastric cancer, with its expression closely correlated with PUS7 levels in tumour tissues. PUS7-dependent pseudouridylation of ALKBH3 mRNA enhances its translation, thereby suppressing gastric cancer progression. These findings highlight the potential significance of mRNA pseudouridylation in cancer biology and suggest a therapeutic target for gastric cancer. PUS7 enhances the translation efficiency of ALKBH3 through its pseudouridylation activity on ALKBH3 mRNA, thereby inhibiting gastric tumourigenesis. The expression levels of PUS7 and ALKBH3 are significantly correlated in gastric tumours, which may be potential prognostic predictors and therapeutic targets for patients with gastric cancer.

CXCL6-CXCR2 axis-mediated PD-L2+ mast cell accumulation shapes the immunosuppressive microenvironment in osteosarcoma

Osteosarcoma (OS) is the most common primary bone malignancy and has a high propensity for local invasion and metastasis. The tumour microenvironment of OS is infiltrated by a large number of immune cells, which play a crucial role in its progression and prognosis. Mast cells are important innate immune cells in the tumour stroma and exhibit different phenotypes in diverse tumour microenvironments. However, the underlying mechanisms of mast cell accumulation and the phenotypic characteristics of mast cells in OS remain poorly understood. In this article, we found for the first time that mast cell accumulation in osteosarcoma tissue was modulated by the CXCL6-CXCR2 axis and that the number of infiltrating mast cells was significantly greater in tumour tissues than in adjacent nontumour tissues. These tumour-infiltrating mast cells express high levels of the immunosuppressive molecule PD-L2, and survival analyses revealed that patients in the PD-L2+ high-expression group had a worse prognosis. In vitro, mast cells were induced to express PD-L2 in a time- and dose-dependent manner using OS tissue culture supernatants to mimic the tumour microenvironment. Mechanistic studies revealed that tumour cell-derived G-CSF significantly induced mast cell PD-L2 expression by activating STAT3. Importantly, mast cells overexpressing PD-L2 inhibit tumour-specific CD8+ T-cell proliferation and tumour-killing cytokine secretion, which is reversed by blocking PD-L2 on mast cells. Therefore, our findings provide new insight into the immunosuppressive and tumorigenic roles of mast cells, as well as a novel mechanism by which PD-L2-expressing mast cells mediate immune tolerance.

Pan-cancer analysis of DCTN2 and its tumour-promoting role in HCC by modulating the AKT pathway.

Dynactin subunit 2 (DCTN2) has been reported to play a role in progression of several tumours; however, the involvement of DCTN2 in potential mechanism or the tumour immune microenvironment among various cancers still remains largely unknown. Therefore, the objective of this study was to comprehensively investigate the expression status and potential function of DCTN2 in various malignancies through different database, such as The Cancer Genome Atlas, the Genotype-Tissue Expression and Gene Expression Omnimus databases. We discovered that DCTN2 expression was high in many type of tumours tissues compared to adjacent non-tumour ones. High DCTN2 signified poor prognosis for patients with tumours. Additionally, Gene Set Enrichment Analysis (GSEA) analysis revealed that DCTN2 was positively correlated with oncogenic pathways, including cell cycle, tumour metastasis-related pathway, while it was negatively with anti-tumour immune signalling pathway, such as INF-γ response. More importantly, we elucidated the functional impact of DCTN2 on hepatocellular carcinoma (HCC) progression and its underlying mechanisms. DCTN2 expression was much higher in HCC tissues than in adjacent non-tumour tissues. Silencing DCTN2 dramatically suppressed the proliferative and metastasis capacities of tumour cell invitro. Mechanistically, DCTN2 exerted tumour-promoting effects by modulating the AKT signalling pathway. DCTN2 knockdown in HCC cells inhibited AKT phosphorylation and its downstream targets as well. Rescue experiments revealed that the anti-tumour effects of DCTN2 knockdown were partially reversed upon AKT pathway activation. Overall, DCTN2 may be a potent biomarker signifying tumour prognosis and a promising therapeutic target for tumour treatment, particularly in HCC.

GALNT14 in association with GDF-15 promotes stemness and drug resistance through β-catenin signalling pathway in breast cancer.

Altered glycosylation plays a role in carcinogenesis. GALNT14 promotes cancer stem-like properties and drug resistance. GDF-15 is known to induces drug resistance and stemness markers for maintenance of breast cancer (BC) stem-like cell state. Currently there is lack of data on association of GDF-15 and GALNTs. In this study, the expression and interaction of GALNT14 and GDF-15 with stemness (OCT4 and SOX2) and drug resistance (ABCC5) markers were evaluated in BC. We investigated tumour tissue from 30 BC patients and adjacent non-tumour tissues. Expression of serum GALNT14 from BC patients and matched healthy controls was evaluated. Expression of GALNT14, GDF-15, OCT4, SOX2, ABCC5, and β-catenin in BC tissue was determined by RT-PCR. Knockdown of GALNT14 and GDF-15 in the MCF-7 cell line was done through siRNA, gene expression and protein expression of β-catenin by western blot were determined. A significant increase in the expression of GALNT14, GDF-15, OCT4, SOX2, ABCC5, and β-catenin was observed in BC tumour tissues compared to adjacent non-tumour tissues. The serum level of GALNT14 was significantly high in BC patients (80.7 ± 65.3 pg/ml) compared to healthy controls (12.2 ± 9.12 pg/ml) (p < 0.000). To further analyse the signalling pathway involved in BC stemness and drug resistance, GALNT14 and GDF-15 were knocked down in the MCF-7 cell line, and it was observed that after knockdown, the expression level of OCT4, SOX2, ABCC5, and β-catenin was decreased, and co-knockdown with GALNT14 and GDF-15 further decreased the expression of genes. It can be concluded that GALNT14, in association with GDF-15, promotes stemness and intrinsic drug resistance in BC, possibly through the β-catenin signalling pathway.

MicroRNA 211-5p inhibits cancer cell proliferation and migration in pancreatic cancer by targeting BMP2.

MicroRNAs (miRNAs) are essential to the tumour growth and metastasis of several cancers. However, the implied functions of miR-211-5p in pancreatic cancer (PC) remains poorly known. In the present study, we discovered that miR-211-5p was a significantly downregulated miRNA in PC tissues compared to adjacent non-tumour tissues. Moreover, we revealed that miR-211-5p overexpression suppressed the proliferation and metastasis of PC cells. Mechanistically, miR-211-5p directly bond to 3'UTR of bone morphogenetic protein-2 (BMP2) and negatively regulated its expression. Rescue experiments showed that the biological function of miR-211-5p was reversed by BMP-2 overexpression in PC cells. Clinical data indicated that BMP2 expression was negatively correlated with miR-211-5p levels in PC patients. Our study provided evidence that miR-211-5p served as a significant suppressor in PC, provided potential targets for prognosis and treatment of patients with PC.

ZNF746 promotes M2 macrophage polarisation and favours tumour progression in breast cancer via the Jagged1/Notch pathway

Breast cancer (BC) is a major threat to women's health. BC is a heterogeneous disease and treatment strategies and outcomes differ between subtypes. Investigating the molecular mechanisms of BC will help to identify potential therapeutic targets and develop new therapies. Here we report that zinc finger protein 746 (ZNF746), a Krüppel-associated box and zinc finger protein, exhibits tumour-promoting properties in BC. Functional experiments (cell growth, colony formation, cell cycle analysis, and transwell analysis) were used to evaluate the proliferation, migration, and invasion capacity of BC cells. Immunohistochemistry was performed to detect the expression of ZNF746, CD163 (M2 macrophage marker), and HES1 (Notch target) in BC tissues. ZNF746 was highly expressed in BC tissues compared to adjacent paired non-tumour tissues. Patients with M1 BC had higher expression of ZNF746 compared to patients with non-metastatic (M0) BC, and higher expression of ZNF746 was associated with poorer overall survival. The immunohistochemical results showed a positive correlation between the expression of ZNF746 and the expression of CD163 or HES1. ZNF746 promoted BC cell proliferation, migration, and invasion and increased the expression of molecules essential for monocyte recruitment and differentiation (CCL2 and CSF1). Furthermore, THP-1 monocytes cultured in the conditioned medium derived from BC cells overexpressing ZNF746 exhibited enhanced M2 polarisation. In contrast, ZNF746 knockdown reduced BC cell proliferation, migration, and invasion and suppressed M2 polarisation. Mechanistically, ZNF746 promoted the activation of the Jagged1/Notch pathway, and the Jagged1 siRNA-mediated blockade of this pathway prevented the tumour-promoting functions of ZNF746. In conclusion, this study uncovers the role of ZNF746 in promoting M2 macrophage polarisation and suggests that ZNF746 may be a promising therapeutic target for limiting BC progression.

Identification of the γ-glutamyl cycle as a novel therapeutic target and 5-oxoproline as a new biomarker for diagnosing pancreatic cancer

Background Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignant solid tumours, and abnormal metabolic reprogramming in the tumour microenvironment is regarded as an important contributor to its pathogenesis. Objectives As there is an urgency to identify new targets based on the metabolic features that are highly refractory to PDAC treatment, this study aimed to identify suitable therapeutic targets for PDAC. Methods In this study, gene set enrichment and Kyoto Encyclopedia of Genes and Genomes analyses were performed on 163 PDAC tissue samples and 165 normal pancreatic tissue samples from The Cancer Genome Atlas and Genotype-Tissue Expression databases to identify alterations in critical metabolites that may contribute to PDAC pathogenesis. Furthermore, ultra-performance liquid chromatography–tandem mass spectrometry was performed to identify significant metabolic pathways between 24 pairs of tumour and adjacent non-tumour tissues and between serum samples from PDAC patients and healthy donors. Results Fifty-one tissue metabolites and 26 serum metabolites were altered in PDAC. Among them, those in the γ-glutamyl cycle were the most substantially changed, and 5-oxoproline was the biomarker of PDAC with the most significantly decreased levels. Conclusions The γ-glutamyl cycle and 5-oxoproline might be potential biomarkers and therapeutic targets to improve the diagnosis, therapy, and prognosis of PDAC.

ABCG2 Gene and ABCG2 Protein Expression in Colorectal Cancer-In Silico and Wet Analysis.

ABCG2 (ATP-binding cassette superfamily G member 2) is a cell membrane pump encoded by the ABCG2 gene. ABCG2 can protect cells against compounds initiating and/or intensifying neoplasia and is considered a marker of stem cells responsible for cancer growth, drug resistance and recurrence. Expression of the ABCG2 gene or its protein has been shown to be a negative prognostic factor in various malignancies. However, its prognostic significance in colorectal cancer remains unclear. Using publicly available data, ABCG2 was shown to be underexpressed in colon and rectum adenocarcinomas, with lower expression compared to both the adjacent nonmalignant lung tissues and non-tumour lung tissues of healthy individuals. This downregulation could result from the methylation level of some sites of the ABCG2 gene. This was connected with microsatellite instability, weight and age among patients with colon adenocarcinoma, and with tumour localization, population type and age of patients for rectum adenocarcinoma. No association was found between ABCG2 expression level and survival of colorectal cancer patients. In wet analysis of colorectal cancer samples, neither ABCG2 gene expression, analysed by RT-PCR, nor ABCG2 protein level, assessed by immunohistochemistry, was associated with any clinicopathological factors or overall survival. An ABCG2-centered protein-protein interaction network build by STRING showed proteins were found to be involved in leukotriene, organic anion and xenobiotic transport, endodermal cell fate specification, and histone methylation and ubiquitination. Hence, ABCG2 underexpression could be an indicator of the activity of certain signalling pathways or protein interactors essential for colorectal carcinogenesis.

Ubiquitin‐dependent regulation of MYC: A promising therapeutic target

Ubiquitin‐dependent regulation of MYC: A promising therapeutic target

IL-36 expression is increased in NSCLC with IL-36 stimulation of lung cancer cells promoting a pro-tumorigenic phenotype

The IL-36 cytokines are a recently described subset of the IL-1 family of cytokines, and have been shown to play a role in the pathogenesis of respiratory diseases such as asthma and COPD. Given the common aetiological links between COPD and lung cancer development, as well as the involvement of other IL-1 family members in lung tumorigenesis, the aim of this work was to investigate the role of IL-36 cytokines in the pathogenesis of lung cancer. In this study we demonstrate that expression of IL-36 cytokines and receptor mRNA and protein are significantly increased in lung cancer tissue compared to adjacent non-tumour tissue. In vitro assays showed that stimulation of two lung cancer cell lines, SKMES-1 human squamous cell and LLC murine lung cancer, with IL-36R agonists resulted in increased cellular migration and proliferation. All IL-36 cytokines induced the expression of pro-inflammatory chemokines in both lung cancer cell lines with synergistic effects identified upon co-stimulation of cells with IL-17, IL-22 and TNFα. Furthermore, we report that IL-36 cytokines induce protein expression of the immune checkpoint inhibitor protein PD-L1 on lung cancer cells. Taken together, this data indicates that targeting IL-36R signalling may be a useful targeted therapy for lung cancer patients with IL-36R+ cancer cells.

Rapid, label-free histopathological diagnosis of liver cancer based on Raman spectroscopy and deep learning

Biopsy is the recommended standard for pathological diagnosis of liver carcinoma. However, this method usually requires sectioning and staining, and well-trained pathologists to interpret tissue images. Here, we utilize Raman spectroscopy to study human hepatic tissue samples, developing and validating a workflow for in vitro and intraoperative pathological diagnosis of liver cancer. We distinguish carcinoma tissues from adjacent non-tumour tissues in a rapid, non-disruptive, and label-free manner by using Raman spectroscopy combined with deep learning, which is validated by tissue metabolomics. This technique allows for detailed pathological identification of the cancer tissues, including subtype, differentiation grade, and tumour stage. 2D/3D Raman images of unprocessed human tissue slices with submicrometric resolution are also acquired based on visualization of molecular composition, which could assist in tumour boundary recognition and clinicopathologic diagnosis. Lastly, the potential for a portable handheld Raman system is illustrated during surgery for real-time intraoperative human liver cancer diagnosis.

Characterisation of a novel transcript LNPPS acting as tumour suppressor in bladder cancer via PDCD5-mediated p53 degradation blockage.

Long non-coding RNAs (lncRNAs) play a crucial role in tumour initiation and progression. However, little is known about their contributions to p53-related bladder cancer (BC) inhibition. By using high-throughput sequencing, we screened the expression profiles of lncRNAs in BC and adjacent non-tumour tissues. The roles of a novel lncRNA, named LNPPS [a lncRNA for programmed cell death 5 (PDCD5) and p53 stability], were determined by gain- and loss-of-function assays. RNA pull-down followed by mass spectrometry analysis, RNA immunoprecipitation assays and other immunoprecipitation assays were performed to reveal the interactions among LNPPS, PDCD5 and p53, and the regulatory effect of LNPPS on the complex ubiquitination network comprising PDCD5, p53 and mouse double minute 2 homologue (MDM2). LNPPS was downregulated in BC and markedly inhibited the viability of BC cells by inducing PDCD5/p53-related apoptosis in vivo and in vitro. Mechanistically, LNPPS, serving as a scaffold, connected PDCD5 and p53 with nucleotides (nt) located at 121-251 nt and 251-306nt of LNPPS, respectively. This process allowed LNPPS to protect PDCD5 from proteasomal degradation by blocking its K20 site ubiquitination. On the other hand, the increased interaction between PDCD5 and p53 displaced p53 from the MDM2-p53 ubiquitination complex, resulting in an increase in p53 expression and related apoptosis levels. Moreover, LNPPS could induce the accumulation of PDCD5 and p53 in the nucleus and exert a synergistic effect on the prevention of protein degradation. In addition, we confirmed that the downregulation of LNPPS in BC was mediated by the decreased N6-methyladenosine (m6 A) modification. Our findings highlight a novel cross-talk between LNPPS and the PDCD5/p53/MDM2 ubiquitination axis in BC development, indicating its potential as a therapeutic target for BC patients.

Elevated Notch-1 expression promotes the lymph node metastasis of gastric cancer and the Notch-1-PTEN-ERK1/2 signalling axis promotes the progression of gastric cancer.

Gastric cancer (GC) is one of the most common malignant tumours and has a high fatality rate worldwide. This study investigated the role of the Notch-1 signalling pathway in the pathogenesis and progression of GC. A total of 64 patients with GC were included in this study. Immunohistochemistry staining was used to detect Notch-1 expression in tumour tissues and adjacent non-tumour tissues, and Notch-1 knockdown in GC cells was identified using short hairpin RNA. A cell scratch assay, transwell assay and flow cytometry analysis were used to analyse the effect of Notch-1 knockdown on cell proliferation, migration and cell cycle distribution. The expression of Notch-1, PTEN, Akt, ERK1/2, E-cadherin and other proteins was detected using Western blotting. The expression level of Notch-1 in GC tissues was higher than that in adjacent non-tumour tissues (P<0.05). High levels of Notch-1 were also found to be associated with sex (male) and lymph node metastasis (P<0.05). Notch-1 knockdown in the AGS and BGC-823 GC cell lines inhibited the migration and proliferation of GC cells, and Notch-1 knockdown arrested the cell cycle in the G0/G1 phase. PTEN protein expression was elevated in the presence of Notch-1 knockdown, resulting in the inhibition of phosphorylated Akt protein expression. In addition, phosphorylated ERK protein levels decreased in the presence of Notch-1 knockdown. Further inhibition of ERK1/2 signalling by the MEK1/2 inhibitor U0126 decreased the proliferation of AGS cells. The results of in vivo experiments with xenotransplantation in nude mice are consistent with these results. Notch-1 plays a key role in the development of GC and was found to promote the lymph node metastasis of GC. Notch-1 knockdown can effectively attenuate the progression of GC cells, which may function in part through the Notch-1-PTEN-ERK1/2 signalling axis.

Hsa-microRNA-27b-3p inhibits hepatocellular carcinoma progression by inactivating transforming growth factor-activated kinase-binding protein 3/nuclear factor kappa B signalling

BackgroundMicroRNAs (miRNAs) play crucial roles in the development of hepatocellular carcinoma (HCC). Hsa-microRNA-27b-3p (hsa-miR-27b) is involved in the formation and progression of various cancers, but its role and clinical value in HCC remain unclear.MethodsThe expression of hsa-miR-27b in HCC was examined by quantitative real-time PCR (qRT-PCR) and in situ hybridization (ISH) assays of clinical samples. Cell Counting Kit-8 assays (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU) incorporation assays, Transwell assays, filamentous actin (F-actin) staining and western blot analyses were used to determine the effects of hsa-miR-27b on HCC cells in vitro. Subcutaneous xenograft and lung metastatic animal experiments were conducted to verify the role of hsa-miR-27b in HCC in vivo. In silico prediction, qRT-PCR, western blot, anti-Argonaute 2 (AGO2) RNA immunoprecipitation (RIP) and dual luciferase reporter assays were applied to identify the target genes of hsa-miR-27b. To detect the impacts of hsa-miR-27b on nuclear factor kappa B (NF-кB) signalling cascades mediated by transforming growth factor-activated kinase-binding protein 3 (TAB3), we performed qRT-PCR, western blot assays, immunofluorescence staining, immunohistochemistry (IHC) and dual-luciferase reporter assays. Recombinant oncolytic adenovirus (OncoAd) overexpressing hsa-miR-27b was constructed to detect their therapeutic value in HCC.ResultsThe expression of hsa-miR-27b was lower in HCC than in adjacent non-tumourous tissues (ANTs), and the reduced expression of hsa-miR-27b was associated with worse outcomes in patients with HCC. Hsa-miR-27b significantly inhibited the proliferation, migration, invasion, subcutaneous tumour growth and lung metastasis of HCC cells. The suppression of hsa-miR-27b promoted the nuclear translocation of NF-κB by upregulating TAB3 expression. TAB3 was highly expressed in HCC compared with ANTs and was negatively correlated with the expression of hsa-miR-27b. The impaired cell proliferation, migration and invasion by hsa-miR-27b overexpression were recovered by ectopic expression of TAB3. Recombinant OncoAd with overexpression of hsa-miR-27b induced anti-tumour activity compared with that induced by negative control (NC) OncoAd in vivo and in vitro.ConclusionsBy targeting TAB3, hsa-miR-27b acted as a tumour suppressor by inactivating the NF-кB pathway in HCC in vitro and in vivo, indicating its therapeutic value against HCC.Graphical

FDX1 expression predicts favourable prognosis in clear cell renal cell carcinoma identified by bioinformatics and tissue microarray analysis.

Ferredoxin 1 (FDX1), an iron-sulphur protein, is responsible for electron transfer in a range of metabolic redox reactions. Clear cell renal cell carcinoma (ccRCC) is an aggressive cancer characterised by metabolic reprogramming, and FDX1 is a critical regulator of cuproptosis. However, the expression profile and prognostic value of FDX1 associated with clinicopathological features in ccRCC remain largely unelucidated. In this study, we integrated a series of public bioinformatic analysis to explore the mRNA and protein profiles of FDX1 across human cancers and cell lines and validated its expression and prognostic value, especially in ccRCC. In this study, FDX1 mRNA and protein expression were aberrantly downregulated and associated with ccRCC grade, stage, and nodal metastasis, whereas in adjacent non-tumour kidney tissue, it was abundantly expressed and cytoplasmically localised in renal tubular epithelial cells. Multivariate analysis indicated that low FDX1 expression contributed to unfavourable overall and disease-free survival. The functional enrichment of FDX1 co-expressed genes in ccRCC involved mainly mitochondrial dysfunction in various metabolic processes and biological oxidation, besides iron-sulphur cluster biogenesis. Furthermore, FDX1 modulates immunological infiltration to affect prognosis. Thus, FDX1 downregulation is mechanistically because of ccRCC tumourigenesis and is a promising prognostic biomarker to stratify patients with ccRCC.

Sphingosine 1-phosphate lyase facilitates cancer progression through converting sphingolipids to glycerophospholipids.

BackgroundIn addition to potent agonist properties for sphingosine 1‐phosphate (S1P) receptors, intracellularly, S1P is an intermediate in metabolic conversion pathway from sphingolipids to glycerolysophospholipids (glyceroLPLs). We hypothesized that this S1P metabolism and its products might possess some novel roles in the pathogenesis of cancer, where S1P lyase (SPL) is a key enzyme.MethodsThe mRNA levels of sphingolipid‐related and other cancer‐related factors were measured in human hepatocellular carcinoma (HCC), colorectal cancer, and esophageal cancer patients’ tumours and in their adjacent non‐tumour tissues. Phospholipids (PL) and glyceroLPLs were measured by using liquid chromatography‐tandem mass spectrometry (LC‐MS/MS). In‐vitro experiments were performed in Colon 26 cell line with modulation of the SPL and GPR55 expressions. Xenograft model was used for determination of the cancer progression and for pharmacological influence.ResultsBesides high SPL levels in human HCC and colon cancer, SPL levels were specifically and positively linked with levels of glyceroLPLs, including lysophosphatidylinositol (LPI). Overexpression of SPL in Colon 26 cells resulted in elevated levels of LPI and lysophosphatidylglycerol (LPG), which are agonists of GPR55. SPL overexpression‐enhanced cell proliferation was inhibited by GPR55 silencing. Conversely, inhibition of SPL led to the opposite outcome and reversed by adding LPI, LPG, and metabolites generated during S1P degradation, which is regulated by SPL. The xenograft model results suggested the contribution of SPL and glyceroLPLs to tumour progression depending on levels of SPL and GPR55. Moreover, the pharmacological inhibition of SPL prevented the progression of cancer. The underlying mechanisms for the SPL‐mediated cancer progression are the activation of p38 and mitochondrial function through the LPI, LPG‐GPR55 axis and the suppression of autophagy in a GPR55‐independent manner.ConclusionA new metabolic pathway has been proposed here in HCC and colon cancer, SPL converts S1P to glyceroLPLs, mainly to LPI and LPG, and facilitates cancer development.