Stem Cells International最新文献

Human bone marrow-derived mesenchymal stem cells (hBMSCs) are promising candidates for stem cell therapy in clinical trials. Applications of hBMSCs in clinical therapy are limited by cellular senescence due to long-term ex vivo expansion. Metformin, an oral hypoglycemic drug for type 2 diabetes, has been shown to have antiaging effects. However, the mechanisms of metformin in antiaging treatment remain controversial. Here, we used D-galactose (D-gal) to establish an appropriate model of senescent hBMSCs to explore the antiaging effects of metformin. Following metformin treatment with a low concentration range, senescence phenotypes induced by D-gal significantly changed, including generation of reactive oxygen species (ROS), loss of mitochondrial membrane potential (MMP), and cell cycle arrest. In contrast, no apparent change was found in unsenescent hBMSCs. Furthermore, the results show that activation of 5'AMP-activated protein kinase (AMPK) by metformin enhances cell autophagy in senescent hBMSCs. These findings suggest that metformin exerts antiaging function within the low concentration range by enhancing autophagy and exhibits potential benefits for clinical stem cell therapy by ameliorating the ex vivo replicative senescence of hBMSCs.

Aging is an inevitable process that biological changes accumulate with time and results in increased susceptibility to different tumors. But currently, aging-related genes (ARGs) in osteosarcoma were not clear. We investigated the potential prognostic role of ARGs and established an ARG-based prognostic signature for osteosarcoma. The transcriptome data and corresponding clinicopathological information of patients with osteosarcoma were obtained from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Molecular subtypes were generated based on prognosis-related ARGs obtained from univariate Cox analysis. With ARGs, a risk signature was built by univariate, least absolute shrinkage and selection operator (LASSO), and multivariate Cox regression analyses. Differences in clinicopathological features, immune infiltration, immune checkpoints, responsiveness to immunotherapy and chemotherapy, and biological pathways were assessed according to molecular subtypes and the risk signature. Based on risk signature and clinicopathological variables, a nomogram was established and validated. Three molecular subtypes with distinct clinical outcomes were classified based on 36 prognostic ARGs for osteosarcoma. A nine-ARG-based signature in the TCGA cohort, including BMP8A, CORT, SLC17A9, VEGFA, GAL, SSX1, RASGRP2, SDC3, and EVI2B, has been created and developed and could well perform patient stratification into the high- and low-risk groups. There were significant differences in clinicopathological features, immune checkpoints and infiltration, responsiveness to immunotherapy and chemotherapy, cancer stem cell, and biological pathways among the molecular subtypes. The risk signature and metastatic status were identified as independent prognostic factors for osteosarcoma. A nomogram combining ARG-based risk signature and metastatic status was established, showing great prediction accuracy and clinical benefit for osteosarcoma OS. We characterized three ARG-based molecular subtypes with distinct characteristics and built an ARG-based risk signature for osteosarcoma prognosis, which could facilitate prognosis prediction and making personalized treatment in osteosarcoma.

Background: Obesity-related diseases have important implications for the occurrence, severity, and outcome of ischemic heart disease. Patients with obesity, hyperlipidemia, and diabetes mellitus (metabolic syndrome) are at increased risk of heart attack with decreased plasma lipocalin levels, and lipocalin is negatively correlated with heart attack incidence. APPL1 is a signaling protein with multiple functional structural domains and plays an important role in the APN signaling pathway. There are two known subtypes of lipocalin membrane receptors, AdipoR1 and AdipoR2. AdioR1 is mainly distributed in skeletal muscle while AdipoR2 is mainly distributed in the liver.

Objective: To clarify whether the AdipoR1-APPL1 signaling pathway mediates the effect of lipocalin in reducing myocardial ischemia/reperfusion injury and its mechanism will provide us with a new approach to treat myocardial ischemia/reperfusion injury using lipocalin as an intervention and therapeutic target.

Methods: (1) Induction of hypoxia/reoxygenation in SD mammary rat cardiomyocytes to simulate myocardial ischemia/reperfusion; (2) downregulation of APPL1 expression in cardiomyocytes to observe the effect of lipocalin on myocardial ischemia/reperfusion and its mechanism of action.

Results: (1) Primary mammary rat cardiomyocytes were isolated and cultured and induced to simulate MI/R by hypoxia/reoxygenation; (2) lipocalin inhibited H/R-induced apoptosis in cardiomyocytes; and (3) APN attenuated MI/R injury through AdipoR1-APPL1 and the possible mechanism.

Conclusion: This study demonstrates for the first time that lipocalin can attenuate myocardial ischemia/reperfusion injury through the AdipoR1-APPL1 signaling pathway and that the reduction of AdipoR1/APPL1 interaction plays an important role in cardiac APN resistance to MI/R injury in diabetic mice.

Background: The mechanisms underlying M2 macrophage polarization induced by nucleus pulposus (NP) cells are unclear. The effects that M2-polarized macrophages have on NP cells are also controversial.

Methods: Transcriptome sequencing was performed to detect the gene change profiles between NP cells from ruptured intervertebral disc (IVD) and normal IVD. The main difference on biological activities between the two cell groups were analyzed by GO analysis and KEGG analysis. Virus transduction, flow cytometry, immunofluorescence, RT-PCR, western blot, CCK-8, TUNEL staining, and AO/EB staining were performed to explore the interactions between NP cells and RAW264.7 macrophages. Statistics were performed using SPSS26.

Results: 801 upregulated and 276 downregulated genes were identified in NP cells from ruptured IVD in mouse models. According to GO and KEGG analysis, we found that the differentially expressed genes (DEG) were dominantly enriched in inflammatory response, extracellular matrix degradation, blood vessel morphogenesis, immune effector process, ossification, chemokine signaling pathway, macrophage activation, etc. CX3CL1 was one of the top 20% DEG, and we confirmed that both NP tissue and cells expressed remarkably higher level of CX3CL1 in mouse models (p < 0.001^∗). Besides, we further revealed that both the recombinant CX3CL1 and NP cells remarkably induced M2 polarization of RAW264.7 (p < 0.001^∗), respectively, while this effect was significantly reversed by si-CX3CL1 or JMS-17-2 (p < 0.001^∗). Furthermore, we found that M2 macrophages significantly decreased the apoptosis rate (p < 0.001^∗) and the catabolic gene levels (p < 0.001^∗) of NP cells, while increased the viability, proliferation as well as the anabolic gene levels of NP cells (p < 0.01^∗).

Conclusions: Via regulating CX3CL1/CX3CR1 pathway, NP cells can induce the M2 macrophage polarization. M2 polarized macrophages can further promote NP cell viability, proliferation, and anabolism, while inhibit NP cell apoptosis and catabolism.

Stroke has become the most common cause of death among residents in China, among which ischemic stroke accounts for the vast majority reaching 70% to 80%. It is of great importance to actively investigate the protective mechanism of cerebral ischemia injury after IS (ischemic stroke). We constructed cerebral ischemia injury models in vivo MACO rat and in vitro (oxygen-glucose deprivation cell model) and set up different interference groups. RT-PCR (reverse transcription PCR) was conducted to detect the expression of lncRNA in neuronal cells, brain tissue, and plasma of different groups, and ELISA (enzyme-linked immunosorbent assay) and western blot were used to detect the expression of the protein in neuronal cells, brain tissue, and plasma of different groups. Cell activity was detected by the CCK-8 assay, while cell apoptosis was examined by TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) assay. In the rats' neuronal cells and brain tissue, curcumin can inhibit the expression of lncRNA GAS5 (long noncoding RNA growth arrest-specific 5). In oxygen-glucose-deprived neuronal cells in vitro, curcumin and low-expressed lncRNA GAS5 can enhance cell activity and decline cell apoptosis, but the addition of curcumin and overexpressed lncRNA GAS5 can make this phenomenon disappear. In neuronal cells, plasma, and brain tissue, curcumin and the low-expressed lncRNA GAS5 can inhibit the expression of IL-1β (interleukin 1 beta), TNF-α (tumor necrosis factor alpha), IL-6 (interleukin 6), Sox2 (SRY-box transcription factor 2), Nanog, and Oct4 (octamer-binding transcription factor 4). However, overexpressed lncRNA GAS5 and curcumin made the inhibitory effect disappear. In conclusion, this study demonstrated that curcumin could inhibit the expression of lncRNA GAS5, thereby inhibiting the expression of inflammation-related factors IL-1β, TNF-α, and IL-6, and ultimately achieve the purpose of attenuating cerebral ischemic cell damage. However, curcumin and lncRNA GAS5 may not alleviate cerebral ischemic cell damage by affecting stem cell differentiation.

The effects of the regulation of phosphatase and tensin homolog deleted on chromosome ten (PTEN) by microribonucleic acid- (miR-) 455-3p on bone marrow stem cells' (BMSCs') chondrogenic development were examined based on the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signal pathway. The alterations in miR-455-3p and PTEN were identified using osteoarthritis (OA) and healthy chondrocytes. Rats raised on the SD diet had their BMSCs isolated for chondrocyte-induced differentiation (blank group), transfected miR-455-3p mimic (mimic group), and inhibitor (inhibitor group). Besides, cell proliferation, alizarin red mineralization staining, and the activity of alkaline phosphatase (ALP) were detected. Real-time fluorescent quantitation polymerase chain reaction (PCR) and Western blot were utilized to detect Runx2, OPN, OSX, COL2A1 mRNA, and the difference between PI3K and AKT. Dual-luciferase reporter (DLR) genes were selected to analyze the target relationship of miR-455-3p to PTEN. It was demonstrated that miR-455-3p in OA was downregulated, while PTEN was upregulated (P < 0.05) in comparison to healthy chondrocytes (P < 0.05). Versus those in the blank group, alizarin red mineralization staining and the activity of ALP increased; RUNX, OPN, OSX, COL2A1 mRNA, p-PI3K, and p-AKT were elevated in the mimic group (P < 0.05). Versus those in the blank and mimic groups, alizarin red mineralization staining and the activity of ALP reduced; RUNX, OPN, OSX, COL2A1 mRNA, p-PI3K, and p-AKT were downregulated in the inhibitor group (P < 0.05). miR-455-3p could target PTEN to inhibit its expression, thus activating the PI3K/AKT signal pathway and promoting BMSCs chondrocyte-induced differentiation. The research results provided reference for the occurrence of OA and the study on therapeutic target.

The purpose of this work was to explore the molecular mechanisms by which USP21 regulates nasopharyngeal carcinoma tumor growth and cancer cell stemness. In this study, the USP21 transcript data was obtained from TCGA database. Then, qPCR and western blot tests revealed that, in contrast to normal tissue or normal nasopharyngeal epithelial cells, the expression of USP21 was greater in nasopharyngeal carcinoma tissues or cell lines, respectively. CCK-8 and EdU immunofluorescent staining assays revealed that USP21 promoted the proliferation of nasopharyngeal carcinoma cells. Meanwhile, scratch and transwell assays showed that USP21 facilitated migration and invasion of nasopharyngeal carcinoma cells. Sphere formation assay was performed on nasopharyngeal carcinoma cells after knockdown of USP21, which revealed that knockdown of USP21 inhibited the stemness profiles of nasopharyngeal carcinoma cells. Then, the western blot assays indicated that knockdown of USP21 in nasopharyngeal carcinoma cells would inhibit FOXM1 expression, and overexpression of FOXM1 could reverse the cell proliferation ability, cell migration and invasion ability, and cell stemness profiles. Finally, a nasopharyngeal xenograft model suggested that USP21 facilitated tumor growth in mice. These findings proved that USP21 promoted tumor growth and cancer cell stemness in nasopharyngeal carcinoma by regulating FOXM1.

Objective: This study is aimed at discussing the value of RNA interference technology on inhibiting lacrimal gland fibrosis in IgG4-related ocular disease (IgG4-ROD).

Methods: Six patients with IgG4-ROD who came to the hospital for surgical treatment from October 2018 to August 2019 were selected, and their diseased lacrimal glands were taken for primary cell culture and fibroblast identification. High efficiency and specificity small interference RNA (siRNA) plasmid vector was constructed, its inhibitory effect on fibroblast proliferation was determined by CCK-8 assay, and the appropriate concentration was selected as the siRNA concentration for subsequent experiments. RT-PCR and Western blot detected the relative expression levels of Fibulin-5 mRNA and protein in the cells 48 hours after transfection. The apoptosis rate of each group of cells at 24 hours, 48 hours, and 72 hours after transfection was detected by flow cytometry, and the proliferation and apoptosis of cells after silencing Fibulin-5 were analyzed and compared.

Results: 24 hours after transfection, there was no significant difference in the proliferation rate among the four groups (P > 0.05); 48 hours and 72 hours after Fibulin-5 siRNA transfection, the proliferation activity of the transfected cells was significantly decreased compared with the 0 nM group, and the inhibitory effect of 75 nM siRNA was the strongest. The expression of Fibulin-5 mRNA and protein in the siRNA-transfected cells was significantly decreased compared with the blank and empty vector negative siRNA groups, and the difference was statistically significant (P < 0.05). The apoptosis rate of cells in the Fibulin-5 siRNA transfection group was significantly higher than that of cells in the blank and empty vector negative siRNA groups, and the difference was statistically significant (P < 0.05).

Conclusion: Fibulin-5 siRNA recombinant plasmid can significantly downregulate the mRNA and protein expressions of target gene Fibulin-5 and promote apoptosis after transfection into IgG4-ROD lacrimal gland fibroblasts. It is speculated that Fibulin-5 can be used as a target to effectively inhibit the fibrosis of lacrimal gland tissues by RNAi technique.

The nucleus pulposus is an elastic jelly composed of crisscross fibrous reticular structures, namely, chondrocytes and proteoglycan mucoid matrix. Embryo and adult SC can resist the accumulation of genetic damage and repair them through various DNA repair mechanisms, thus preventing them from spreading to daughter cells. Fresh medullary tissue was fixed with 10% formaldehyde solution, embedded in paraffin, and sectioned at 4 m. The nucleus pulposus was stained with HE, and its degeneration was observed under light microscope. The average apoptotic index (AI) of 20 denatured nuclei was 50.230, the percentage of Fas-positive cells was 74.255%, and the percentage of Bcl-2-positive cells was 55.370%. The average apoptotic index (AI) was 28.317. The percentage of Fas-positive cells, Fas protein-positive cells, and Bcl-2 protein-positive cells in six normal nuclei was 41.717%, 41.717%, and 27.167%, respectively. The average AI value, Fas protein expression, and Bcl-2 protein expression in the two groups were significantly different (P < 0.05).

Long noncoding RNAs (lncRNAs) play crucial roles in melanoma initiation and development, serving as potential therapeutic targets and prognostic markers for melanoma. lncRNA survival-associated mitochondrial melanoma-specific oncogenic noncoding RNA (SAMMSON) is upregulated in many types of human cancers. However, the functions of SAMMSON in melanoma have not been fully elucidated. This study is aimed at investigating the expression and functions of SAMMSON in melanoma development. Bioinformatics analysis was performed to determine the expression of SAMMSON and its correlation with the 10-year overall survival (OS) in melanoma patients. Cell proliferation, migration, invasion, and tumorigenesis were detected by MTT, colony formation, Transwell assays, and mouse xenograft model. The expression of cell cycle-related factors, epithelial-to-mesenchymal transition (EMT) makers, and matrix metalloproteinases (MMPs) was assessed by RT-qPCR and western blotting analysis. The results demonstrated that SAMMSON expression was upregulated in melanoma tissues and cells, and lower SAMMSON expression was correlated with longer 10-year OS. SAMMSON knockdown decreased the proliferation, migration, and invasion of melanoma cells by regulating the expression of proliferation-related genes, EMT factors, and MMPs, respectively. Additionally, Forkhead box protein A2 (FOXA2) was confirmed to be a target of SAMMSON, and the biological effects induced by FOXA2 overexpression were similar to those induced by SAMMSON silencing in melanoma cells. Further studies showed that SAMMSON downregulated FOXA2 expression in melanoma cells by modulating the EZH2/H3K27me3 axis. Taken together, our data indicate that SAMMSON plays an important role in melanoma progression and can be a valuable biomarker and therapeutic target in melanoma.