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Genes Nov 2022tRNA-derived small RNAs (tsRNAs) as a novel non-coding RNA have been studied in many cardiovascular diseases, but the relationship between tsRNAs and septic...
tRNA-derived small RNAs (tsRNAs) as a novel non-coding RNA have been studied in many cardiovascular diseases, but the relationship between tsRNAs and septic cardiomyopathy has not been investigated. We sought to analyze changes of the expression profile of tsRNAs in septic cardiomyopathy and reveal an important role for tsRNAs. We constructed a sepsis model by cecal ligation and puncture (CLP) in mice, and microarray analysis was used to find differentially expressed tsRNAs. Quantitative real-time PCR was used to verify the expression of tsRNAs and the interference effect of angiogenin (ANG), a key nuclease producing tsRNAs. Bioinformatics analysis was used to predict target genes and functions. CCK-8 and LDH release assays were used to detect cell viability and cell death. A total of 158 tsRNAs were screened, of which 101 were up-regulated and 57 were down-regulated. A total of 8 tsRNAs were verified by qPCR, which was consistent with microarray results. Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses suggest that these tsRNAs may be associated with the Wnt signaling pathway and participate in cellular process. The expression of tsRNAs decreased after the interference of the key nuclease ANG, while CCK-8 suggested a corresponding decrease in cell viability and an increase in the release of LDH (cell death), indicating that tsRNAs can protect cardiomyocytes during the development of septic cardiomyopathy, reduced cardiomyocyte death. : A total of 158 tsRNAs changed significantly in septic cardiomyopathy, and these tsRNAs may play a protective role in the development of septic cardiomyopathy.
Topics: Mice; Animals; Sincalide; RNA, Transfer; MicroRNAs; Microarray Analysis; Cardiomyopathies
PubMed: 36553526
DOI: 10.3390/genes13122258 -
PeerJ 2022Recent studies have identified ribonucleotide reductase subunit M2 (RRM2) as a putative promoter of tumors. However, no systematic analysis of its carcinogenicity has...
BACKGROUND
Recent studies have identified ribonucleotide reductase subunit M2 (RRM2) as a putative promoter of tumors. However, no systematic analysis of its carcinogenicity has been conducted.
METHODS
The potential functions of RRM2 in various tumor types were investigated using data from the Genotype-Tissue Expression (GTEx), the Clinical Proteomic Tumor Analysis Consortium (CPTAC), the Cancer Genome Atlas (TCGA), the Human Protein Atlas (HPA), cBioPortal, GEPIA, String, and Gene Set Enrichment Analysis (GSEA). We analyzed the difference in mRNA and protein expression, pathological stage, survival, mutation, tumor microenvironment (TME), and immune cell infiltration in relation to RRM2. Meanwhile, using TCGA and the Tumor Immune Estimation Resource 2 (TIMER 2), the associations between RRM2 expression, immune infiltration, and immune-related genes were assessed. Additionally, CCK-8, Edu and RT-PCR assays were used to validate that RRM2 acts as an oncogene in liver cancer cells and its association with HBx. A cohort of liver hepatocellular carcinoma (LIHC) patients (n=154) from Huashan Hospital was analyzed for the expression of RRM2 and the association between RRM2 and immune infiltration.
RESULTS
Using the GTEx and TCGA databases, we discovered that 28 tumors expressed RRM2 at significantly higher levels than the corresponding normal tissues. Increased RRM2 expression may be predictive of a poor overall survival (OS) in patients with seven different cancers. GO, KEGG, and GSEA analyses revealed that the biological process of RRM2 was associated with the regulation of carcinogenic processes and immune pathways in a variety of tumor types. The expression of RRM2 was highly correlated with maker genes involved in immune activation and immunosuppression, immune checkpoints, DNA mismatch repair system (MMR), and the infiltration levels of Tregs and macrophages (TAMs), suggesting that the carcinogenic effect of RRM2 may be achieved by regulating immune related genes. Moreover, as demonstrated by CCK-8 and Edu assays, RRM2 was an oncogene in liver cancer cells. We confirmed for the first time that RRM2 was significantly upregulated by HBx, suggesting that RRM2 may be a key regulator of LIHC induced by HBV. IHC analysis validated the upregulated expression of RRM2 protein and its correlation with immune infiltration makers in a LIHC patient cohort.
CONCLUSION
RRM2 may be a valuable molecular biomarker for predicting prognosis and immunotherapeutic efficacy in pan-cancer, particularly in LIHC.
Topics: Humans; Proteomics; Sincalide; Carcinoma, Hepatocellular; Oncogenes; Liver Neoplasms; Tumor Microenvironment
PubMed: 36518297
DOI: 10.7717/peerj.14432 -
BMC Molecular and Cell Biology Nov 2022To establish castration-resistant prostate cancer (CRPC) - Lncap androgen-independent (AI) cell line from Lncap androgen-dependent (AD) cell line, and explore the...
BACKGROUND
To establish castration-resistant prostate cancer (CRPC) - Lncap androgen-independent (AI) cell line from Lncap androgen-dependent (AD) cell line, and explore the different molecular biological between these two cell lines.
METHODS
The Lncap-AD cell line was cultured and passaged 60 times over 16 months. The morphology of the Lncap-AI cell line was observed. AR levels identification were detected in qRT-PCR and Western Blot assay. CCK-8, EdU assay, wound healing assay and cell adhesion assays were used to observe the ability of proliferation, migration, and adhesion. SEM and TEM were used to observe microculture structure. At last, the PSA secrete ability was evaluated by Elisa assay.
RESULTS
The Lncap-AD cell line was cultured and passaged 60 times over 16 months. The Lncap-AI cell line showed a morphologic change at the end stage of culture, the cells turned slender and cell space turned separated compared to the Lncap-AD cell line. The relative levels of AR-related genes in the Lncap-AI cell line were up-regulation compared to the Lncap-AD cell line both in mRNA and protein levels. The expression of AR and HK2 proteins were influenced and down-regulation by Enzalutamide in the Lncap-AD cell line, but no obvious difference in Lncap-AI cell lines. Lncap-AI cell line showed strong viability of proliferation, migration, and adhesion by CCK-8, EdU assay, wound healing assay, and adhesion assay. The microstructure of Scanning Electron Microscopy (SEM) showed many synapses in the Lncap-AI cell line and PC3 cell line, but not in the Lncap-AD cell line. At last, the PSA secrete ability was evaluated by Elisa assay, and PCa cell lines showed no significant difference.
CONCLUSION
Simulation of CRPC progression, Lncap-AD cell line turned to Lncap-AI cell line with androgen deprivation therapy.
Topics: Male; Humans; Flutamide; Androgens; Androgen Antagonists; Prostate-Specific Antigen; Prostatic Neoplasms, Castration-Resistant; Sincalide; Cell Line
PubMed: 36443669
DOI: 10.1186/s12860-022-00453-2 -
Disease Markers 2022LINC01140 has been known to be involved in various cancers. However, its underlying molecular mechanism in breast cancer (BC) needs further exploration.
BACKGROUND
LINC01140 has been known to be involved in various cancers. However, its underlying molecular mechanism in breast cancer (BC) needs further exploration.
METHODS
The LINC01140, miR-452-5p, and RGS2 levels in BC cells and tissues were evaluated by means of RT-qPCR and western blotting. The variations in the biological functions of BC cells were analyzed through CCK-8, transwell, western blotting, and xenograft experiments to observe cell viability, migration, levels of apoptosis-related proteins (Bax and Bcl-2), and tumor growth. The correlations existing among LINC01140, miR-452-5p, and RGS2 were validated through luciferase reporter and RIP assays.
RESULTS
LINC01140 and RGS2 were remarkably downregulated in BC cells and tissues, whereas miR-452-5p was upregulated. LINC01140 overexpression diminished BC cell viability, migration, and tumor growth and facilitated apoptosis. MiR-452-5p upregulation enhanced cell viability and migration and suppressed apoptosis. Nevertheless, the additional upregulation of LINC01140 could reverse the promotive effects of miR-452-5p upregulation. Additionally, RGS2 overexpression inhibited the malignant phenotypes of BC cells, but miR-452-5p upregulation abolished this effect. In terms of mechanisms, LINC01140 acted as a miR-452-5p sponge. Moreover, RGS2 was determined to be miR-452-5p's downstream target gene in BC.
CONCLUSION
LINC01140 functioned as an antitumor agent in BC by sponging miR-452-5p to release RGS2. This hints that LINC01140 is a promising therapeutic target for BC.
Topics: Humans; Female; MicroRNAs; Cell Proliferation; Cell Movement; Cell Line, Tumor; Sincalide; bcl-2-Associated X Protein; Breast Neoplasms; Apoptosis Regulatory Proteins; Carcinogenesis; Cell Transformation, Neoplastic; Gene Expression Regulation, Neoplastic; RGS Proteins
PubMed: 36299824
DOI: 10.1155/2022/2434938 -
Oxidative Medicine and Cellular... 2022It has been reported that signaling from the nerve growth factor (NGF) pathway associated with peripheral nerves is able to contribute to perineural invasion (PNI) of...
Nerve Growth Factor (NGF) Encourages the Neuroinvasive Potential of Pancreatic Cancer Cells by Activating the Warburg Effect and Promoting Tumor Derived Exosomal miRNA-21 Expression.
BACKGROUND
It has been reported that signaling from the nerve growth factor (NGF) pathway associated with peripheral nerves is able to contribute to perineural invasion (PNI) of pancreatic cancer (PC). Nevertheless, the underlying mechanism by which NGF leads to PNI remained poorly understood.
METHODS
Western blotting was employed to determine NGF level in PC and paracarcinoma tissues and in PC cell lines as well as pancreatic ductal epithelial cells. MiaPaCa-2 and CFPAC-1 cells were treated with 100 ng/ml of NGF or the NGF inhibitor Tanezumab for 24 h, CCK-8 and Transwell assays were employed to test cell proliferation, invasion, and migration, respectively. TrkA expression was knocked down in MiaPaCa-2 and dorsal root ganglion (DRG) cells treated with NGF to determine its effect on the Warburg effect. To reveal that the NGF-TrkA signaling pathway was closely associated with PC PNI, neuroinvasion model was established by using MiaPaCa-2 cells via coculturing DRG cells in Matrigel. Further, exosomes were extracted from PC cells and identified by examining the levels of specific markers for exosomes. Then RT-qPCR was applied to test miR-21-5p level in tumor derived exosomal (TDE-miR-21-5p). RIP assay was performed to validate NGF and miR-21 binding ability in MiaPaCa-2 cells. Rescue experiments were performed by using coprocessing of Tanezumab and miR-21-5p mimic on MiaPaCa-2 cells, followed by coculture with DRG cells. Subsequently, we used a model of neuroinvasion in nude mice to assess the effect of NGF on tumor nerve invasion as well as on nociceptive transmission.
RESULTS
NGF level was preeminently higher in PC tissues and cell lines than in paracarcinoma tissues and normal pancreatic epithelial cell lines. NGF promoted MiaPaCa-2 and CFPAC-1 cell invasion and migration, while Tanezumab treatment showed the opposite results. Besides, NGF binding to TrkA receptors encouraged the intracellular Warburg effect in PC and DRG cells. TrkA blocking-up could restrain NGF induced PC cell migration and neural invasion. Mechanistically, NGF could upregulate TDE-miR-21-5p levels, and DRG cells took up TDE to activate the Warburg effect and stimulate nociceptor gene expression. miR-21-5p inhibitor could abolish the facilitative effect of NGF on PNI in MiaPaCa-2 cells. tumorigenesis experiments, Tanezumab markedly alleviated nerve invasion of PC cells as well as relieved nociceptive conduction in animal models.
CONCLUSIONS
These findings displayed that NGF/TrkA encouraged the neuroinvasive potential of PC cells by activating the Warburg effect in DRG cells through upregulation of TDE-miR-21-5p expression.
Topics: Animals; Mice; Cell Line, Tumor; Cell Proliferation; Gene Expression Regulation, Neoplastic; Mice, Nude; MicroRNAs; Neoplasm Invasiveness; Nerve Growth Factor; Pancreatic Neoplasms; Sincalide
PubMed: 36285300
DOI: 10.1155/2022/8445093 -
Journal of Experimental & Clinical... Oct 2022Beta-1,3-galactosyltransferase-4 (B3GALT4) plays a critical regulatory role in tumor biology. However, the role of B3GALT4 in modulating the tumor microenvironment (TME)...
BACKGROUND
Beta-1,3-galactosyltransferase-4 (B3GALT4) plays a critical regulatory role in tumor biology. However, the role of B3GALT4 in modulating the tumor microenvironment (TME) of neuroblastoma (NB) remains unknown.
METHODS
Public datasets and clinical NB samples were collected to evaluate the expression and clinical significance of GD2 and B3GALT4 in NB patients. CCK-8, colony formation, and transwell assays and experiments in tumor-bearing mouse models were conducted to investigate the function of B3GALT4. Flow cytometry, ELISA, immunohistochemistry, immunofluorescence, western blotting, and chemotaxis assays were conducted to ascertain the immunomodulatory mechanism of B3GALT4. The combined therapeutic effect of the lipid raft inhibitor MβCD and anti-GD2 mAb was validated in a murine model of NB.
RESULTS
GD2 was overexpressed in NB tissues and high expression of GD2 was associated with poor prognosis in NB patients. B3GALT4 was downregulated in NB tissues, and low expression of B3GALT4 indicated poor prognosis in NB patients. Silencing B3GALT4 significantly enhanced tumor progression both in vitro and in vivo. Meanwhile, the overexpression of B3GALT4 increased the recruitment of CD8 T lymphocytes via the chemokines CXCL9 and CXCL10. Additionally, B3GALT4 regulated NB-cell GD2 expression and lipid raft formation. Mechanistically, B3GALT4 regulated the expression of CXCL9 and CXCL10 via the c-Met signaling in the lipid rafts and the downstream AKT/mTOR/IRF-1 pathway. The lipid raft inhibitor, MβCD, attenuated B3GALT4 deficiency-induced tumor progression and immune evasion. Last, MβCD combined with anti-GD2 mAb treatment significantly enhanced the antitumor effect and the infiltration of CD8 T cells.
CONCLUSIONS
Upregulation of B3GALT4 promotes the secretion of CXCL9 and CXCL10 to recruit CD8 T lymphocytes via the GD2-mediated lipid rafts and the c-Met/AKT/mTOR/IRF-1 pathway. Moreover, lipid raft inhibitors may enhance the efficacy of anti-GD2 immunotherapy for NB.
Topics: Animals; Mice; CD8-Positive T-Lymphocytes; Cell Line, Tumor; Chemokines; Galactosyltransferases; Gangliosides; Membrane Microdomains; Neuroblastoma; Proto-Oncogene Proteins c-akt; Sincalide; TOR Serine-Threonine Kinases; Tumor Microenvironment; Proto-Oncogene Proteins c-met
PubMed: 36284313
DOI: 10.1186/s13046-022-02523-x -
Oxidative Medicine and Cellular... 2022This study investigated the possibility of exosomes loaded with si-PDGFR ability to suppress the progression of glioma. Common gliomas develop from neuroglial progenitor...
This study investigated the possibility of exosomes loaded with si-PDGFR ability to suppress the progression of glioma. Common gliomas develop from neuroglial progenitor cells. Many variables affect the survival rate and occurrence of gliomas. Understanding oxidative stress processes and creating new, efficient treatments are crucial because oxidative stress is linked to the development of brain tumors. For this purpose, selected clinical samples were subjected to various tests like quantitative real-time PCR, Cignal Finder RTK signaling 7-pathway reporter array analysis, CCK-8 analysis, flow cytometry, and immunoblotting. Here, we demonstrated that PDGFR expression was increased in glioma patients. Following that, cell-derived exosomes were extracted and collected and traced and selected tissue samples were subjected to immunohistochemical analysis. The results indicated that the knockdown of PDGFR (si-PDGFR) inhibited the proliferation of glioma cells. Besides this, si-PDGFR-loaded exosomes induced a similar antitumor effect in glioma cells. The anticancer effect of si-PDGFR-loaded exosomes was mediated by the inactivation of the PI3K/Akt/EZH2 pathway. Finally, we verified that this exosome delivery system, si-PDGFR-loaded exosomes, had robust targeting and no associated toxicity. In conclusion, the study confirmed that si-PDGFR-loaded exosomes inhibit glioma progression via inactivating the PI3K/Akt/EZH2 signaling pathway.
Topics: Humans; Cell Proliferation; Enhancer of Zeste Homolog 2 Protein; Exosomes; Glioma; Oxidative Stress; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Signal Transduction; Sincalide; Proto-Oncogene Proteins c-sis
PubMed: 36275907
DOI: 10.1155/2022/5081439 -
Oxidative Medicine and Cellular... 2022Thin endometrium remains a severe clinical challenge with no effective therapy to date. We aimed at exploring the role and molecular mechanism of human umbilical cord...
AIM
Thin endometrium remains a severe clinical challenge with no effective therapy to date. We aimed at exploring the role and molecular mechanism of human umbilical cord mesenchymal stem cell- (hucMSC-) derived exosomes (hucMSC-Ex) in repairing hypoxic injury of endometrial epithelial cells (EECs).
METHODS
Exosomes were harvested from the conditioned medium of hucMSC and characterized using western blot, transmission electron microscopy (TEM), flow cytometry, and nanoparticle tracking analysis (NTA). EECs were subjected to hypoxic conditions before cocultured with hucMSC-Ex. Cell viability, apoptosis, and migration were determined with CCK-8, flow cytometry, and wound healing assay, respectively. Apoptosis/EMT-related proteins were detected by western blot. The miRNA profiling was determined by RNA sequencing. The expression of miR-663a and CDKN2A was measured by qRT-PCR. MiR-663a in EECs was overexpressed by transfecting with miR-663a mimics.
RESULTS
Mesenchymal stem cells (MSCs) markers CD73, CD90, and CD106 were positively expressed in hucMSCs. Exosome isolated from hucMSC expressed CD63 and TSG101, and were 100-150 nm in diameter. HucMSC-Ex promoted cell proliferation inhibited by hypoxia. And hucMSC-Ex also inhibited hypoxia-induced apoptosis, migration, and EMT of EECs by upregulating the expression of Bcl-2 and E-cadherin and downregulating Bax and N-cadherin levels. Further, bioinformatics research found that hucMSC-Ex coculture can significantly upregulate the expression of miR-663a and decrease the expression of CDKN2A in hypoxia-induced EECs. Furthermore, miR-663a overexpression inhibited CDKN2A expression and increased the expression of Bcl-2 and E-cadherin in hypoxia-induced EECs.
CONCLUSIONS
HucMSC-Ex promoted cell proliferation, inhibited cell apoptosis, migration, and EMT in hypoxia-induced EECs, thereby alleviating hypoxia-induced EECs injury, which may be related to its regulation of miR-663a/CDKN2A expression. Our study indicated that hucMSC-Ex might benefit for repairing thin endometrium.
Topics: Female; Humans; Exosomes; Culture Media, Conditioned; Sincalide; bcl-2-Associated X Protein; Mesenchymal Stem Cells; Umbilical Cord; Endometrium; Epithelial Cells; Hypoxia; MicroRNAs; Cadherins; Cyclin-Dependent Kinase Inhibitor p16
PubMed: 36275892
DOI: 10.1155/2022/3082969 -
Oxidative Medicine and Cellular... 2022NHEKs, HaCaT cells, and HEK 293T cells were treated with IL-17A. CCK-8 assays were performed to detect cell activity, and immunofluorescence staining and Western...
METHODS
NHEKs, HaCaT cells, and HEK 293T cells were treated with IL-17A. CCK-8 assays were performed to detect cell activity, and immunofluorescence staining and Western blotting were performed to detect the protein expression of STAT3. After isolation of exosomes via ultracentrifugation, the contents of miR-124-3p and oxidative stress markers such as superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione peroxidase (GSH-Px) in keratinocytes were measured. Subsequently, transcriptomic analysis was performed using RNA-seq. Data were analysed by using the "edgeR" package within R. After verifying the abnormally expressed genes stimulated by IL-17A, a dual luciferase reporter assay was used to determine the interaction between miR-124-3p and STAT3. Finally, BALB/c mice were used to establish a psoriasis model for analysis. The effect of elevated miR-124-3p on the psoriasis mouse model was determined by exosomal delivery of miR-124-3p.
RESULTS
IL-17 intervention enhanced the cell activity of keratinocytes ( < 0.05). miR-124-3p was identified by RNA-seq as one of the differentially expressed miRNAs stimulated by IL-17A. miR-124-3p overexpression induced decreased STAT3 and MDA levels, increased SOD and GSH-Px levels in keratinocytes, and alleviated emergency responses of sclerosis damage ( < 0.05). The dual luciferase reporter assay results confirmed that STAT3 was regulated by miR-124-3p in a targeted manner ( < 0.05). Finally, miR-124-3p delivered by exosomes effectively alleviated the pathological manifestations and oxidative stress responses of psoriatic mice.
CONCLUSIONS
miR-124-3p regulates keratinocyte activity via STAT3 in response to IL-17A stimulation. The ectopic expression of miR-124-3p in psoriatic skin reduces IL-17A-induced inflammation and inhibits the STAT3 pathway, thus alleviating the symptoms of psoriasis. The findings of this study suggest that exosomes can be used to therapeutically deliver miR-124-3p to keratinocytes and psoriatic lesions, which may provide novel insight for psoriasis treatment.
Topics: Mice; Animals; Interleukin-17; Exosomes; Glutathione Peroxidase; Sincalide; Cell Proliferation; Keratinocytes; Psoriasis; MicroRNAs; Superoxide Dismutase; Malondialdehyde
PubMed: 36275890
DOI: 10.1155/2022/6264474 -
Frontiers in Immunology 2022Ti-5Cu alloy has been proved to have excellent mechanical properties and cell compatibility and has certain antibacterial properties due to the addition of Cu. However,...
Ti-5Cu alloy has been proved to have excellent mechanical properties and cell compatibility and has certain antibacterial properties due to the addition of Cu. However, there are few studies on the effects of Ti-5Cu alloy on macrophage polarization and immune-related bone formation. In this study, we prepared Ti-5Cu alloy by three-dimensional printing technology and found that Ti-5Cu alloy presented a much smoother surface compared with Ti. In addition, the CCK-8 results indicated the Ti-5Cu alloy had no cytotoxicity to RAW264.7 cells by co-culture. The results of inductively coupled plasma mass spectrometry showed that the concentration of Cu was 0.133 mg/L after 7 days of co-culture, and the CCK-8 results proved that Cu had no cytotoxicity to RAW264.7 at this concentration. Then, we studied the effects of Ti-5Cu alloy on macrophage polarization; it was shown that the Ti-5Cu alloy is more prone to modulate the RAW264.7 polarization towards the M2 phenotype and the conditioned medium derived from Ti-5Cu alloy significantly promoted the proliferation and osteogenic differentiation of MC3T3-E1 cells. However, when the expression of Oncostatin M (OSM) gene of RAW264.7 was knocked down, the osteogenic differentiation of MC3T3-E1 cells was decreased. This suggests that the OSM secreted by RAW264.7 co-cultured with Ti-5Cu alloy could accelerate the osteogenic differentiation of MC3T3-E1 cells by acting on OSMR/gp130 receptors.
Topics: Osteogenesis; Alloys; Titanium; Oncostatin M; Culture Media, Conditioned; Sincalide; Cytokine Receptor gp130; Macrophages; Phenotype; Printing, Three-Dimensional; Anti-Bacterial Agents
PubMed: 36275667
DOI: 10.3389/fimmu.2022.1001526