Comprehensive Analyses and Experiments Confirmed IGFBP5 as a Prognostic Predictor Based on an Aging-genomic Landscape Analysis of Ovarian Cancer.

BACKGROUND: Ovarian cancer (OC) is one of the malignant diseases of the reproductive system in elderly women. Aging-related genes (ARGs) were involved in tumor malignancy and cellular senescence, but the specifics of these mechanisms in OC remain unknown. METHODS: ARGs expression and survival data of OC patients were collected from TCGA and CPTAC databases. Subtype classification was used to identify the roles of hub ARGs in OC progression, including function enrichment, immune infiltration, and drug sensitivity. LASSO regression was utilized to confirm the prognosis significance for these hub ARGs. MTT, EdU, Transwell, and wounding healing analysis confirmed the effect of IGFBP5 on the proliferation and migration ability of OC cells. RESULTS: ARGs were ectopically expressed in OC tissues compared to normal ovary tissues. Three molecular subtypes were divided by ARGs for OC patients. There were significant differences in ferroptosis, m6A methylation, prognosis, immune infiltration, angiogenesis, differentiation level, and drug sensitivity among the three groups. LASSO regression indicated that 4 signatures, FOXO4, IGFBP5, OGG1 and TYMS, had important prognosis significance. Moreover, IGFBP5 was significantly correlated with immune infiltration. The hub ARG, IGFBP5, expression was significantly decreased in OC patients compared to normal women. IGFBP5 could also reduce the migration and proliferation ability of OC cells compared to vector and NC groups. CONCLUSION: IGFBP5 was correlated with OC prognosis and associated with OC migration and proliferation. This gene may serve as potential prognostic biomarkers and therapeutic targets for OC patients.

Regulation of the integrin αVß3- actin filaments axis in early osteogenic differentiation of human mesenchymal stem cells under cyclic tensile stress.

BACKGROUND: Integrins are closely related to mechanical conduction and play a crucial role in the osteogenesis of human mesenchymal stem cells. Here we wondered whether tensile stress could influence cell differentiation through integrin αVß3. METHODS: We inhibited the function of integrin αVß3 of human mesenchymal stem cells by treating with c(RGDyk). Using cytochalasin D and verteporfin to inhibit polymerization of microfilament and function of nuclear Yes-associated protein (YAP), respectively. For each application, mesenchymal stem cells were loaded by cyclic tensile stress of 10% at 0.5 Hz for 2 h daily. Mesenchymal stem cells were harvested on day 7 post-treatment. Western blotting and quantitative RT-PCR were used to detect the expression of alkaline phosphatase (ALP), RUNX2, ß-actin, integrin αVß3, talin-1, vinculin, FAK, and nuclear YAP. Immunofluorescence staining detected vinculin, actin filaments, and YAP nuclear localization. RESULTS: Cyclic tensile stress could increase the expression of ALP and RUNX2. Inhibition of integrin αVß3 activation led to rearrangement of actin filaments and downregulated the expression of ALP, RUNX2 and promoted YAP nuclear localization. When microfilament polymerization was inhibited, ALP, RUNX2, and nuclear YAP nuclear localization decreased. Inhibition of YAP nuclear localization could reduce the expression of ALP and RUNX2. CONCLUSIONS: Cyclic tensile stress promotes early osteogenesis of human mesenchymal stem cells via the integrin αVß3-actin filaments axis. YAP nuclear localization participates in this process of human mesenchymal stem cells. Video Abstract.

Seasonal changes driving shifts in microbial community assembly and species coexistence in an urban river.

Microbial communities play a vital role in urban river biogeochemical cycles. However, the seasonal variations in microbial community characteristics, particularly phylogenetic group-based community assembly and species coexistence, have not been extensively investigated. Here, we systematically explored the microbiome characteristics and assembly mechanisms of urban rivers in different seasons using 16S rRNA gene sequencing and multivariate statistical methods. The results indicated that the microbial community presented significant temporal heterogeneity in different seasons, and the diversity decreased from spring to winter. The phylogenetic group-based microbial community assembly was governed by dispersal limitation and drift in spring, summer, and autumn but was structured by homogeneous selection in winter. Moreover, the main functions of nitrification, denitrification, and methanol oxidation were susceptible to dispersal limitation and drift processes, whereas sulfate respiration and aromatic compound degradation were controlled by dispersal limitation and homogeneous selection. Network analyses indicated that network complexity decreased and then increased with seasonal changes, while network stability showed the opposite trend, suggesting that higher complexity and diversity reduced community stability. Temperature was determined to be the primary driver of microbial community structure and assembly processes in different seasons based on canonical correspondence analysis and linear regression analysis. In conclusion, seasonal variation drives the dynamics of microbial community assembly and species coexistence patterns in urban rivers. This study provides new insights into the generation and maintenance of microbial community diversity in urban rivers under seasonal change conditions.

miR-21 Targets ASPP2 to Inhibit Apoptosis via CHOP-Mediated Signaling in Helicobacter pylori-Infected Gastric Cancer Cells.

Helicobacter pylori (H. pylori) infection affects cell survival pathways, including apoptosis and proliferation in host cells, and disruption of this balance is the key event in the development of H. pylori-induced gastric cancer (HPGC). H. pylori infection induces alterations in microRNAs expression that may be involved in GC development. Bioinformatic analysis showed that microRNA-21 (miR-21) is significantly upregulated in HPGC. Furthermore, quantitative proteomics and in silico prediction were employed to identify potential targets of miR-21. Following functional enrichment and clustered interaction network analyses, five candidates of miR-21 targets, PDCD4, ASPP2, DAXX, PIK3R1, and MAP3K1, were found across three functional clusters in association with cell death and survival, cellular movement, and cellular growth and proliferation. ASPP2 is inhibited by H. pylori-induced miR-21 overexpression. Moreover, ASPP2 levels are inversely correlated with miR-21 levels in HPGC tumor tissues. Thus, ASPP2 was identified as a miR-21 target in HPGC. Here, we observed that H. pylori-induced ASPP2 suppression enhances resistance to apoptosis in GC cells using apoptosis assays. Using protein interaction network and coimmunoprecipitation assay, we identified CHOP as a direct mediator of the ASPP2 proapoptotic activity in H. pylori-infected GC cells. Mechanistically, ASPP2 suppression promotes p300-mediated CHOP degradation, in turn inhibiting CHOP-mediated transcription of Noxa, Bak, and suppression of Bcl-2 to enact antiapoptosis in the GC cells after H. pylori infection. Clinicopathological analysis revealed correlations between decreased ASPP2 expression and higher HPGC risk and poor prognosis. In summary, the discovery of H. pylori-induced antiapoptosis via miR-21-mediated suppression of ASPP2/CHOP-mediated signaling provides a novel perspective for developing HPGC management and treatment.

Long non-coding RNAs in hepatocellular carcinoma.

Long noncoding RNAs (lncRNAs) refer to a class of RNAs greater than 200 nucleotides in length, most of which are considered unable to encode proteins, thus deemed to be junk genes formerly. But with emerging studies about lncRNAs coming out in recent years, it is much more clearly depicted that they can regulate gene expression at different levels, with various mechanisms, thus participating in diverse biological or pathological processes, including complicated tumor-associated pathways. Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer, the third leading cause of cancer-related mortality worldwide, which has been found to tightly associate with aberrant expression of a variety of lncRNAs regulating tumor proliferation, invasion, drug resistance, and so on, making it a potential novel tumor marker and therapeutic target. In this review, we highlight a few lncRNAs that are closely related to the occurrence and progression of HCC and try to cover their multifarious roles from different layers.

Seasonal succession of microbial community co-occurrence patterns and community assembly mechanism in coal mining subsidence lakes.

Coal mining subsidence lakes are classic hydrologic characteristics created by underground coal mining and represent severe anthropogenic disturbances and environmental challenges. However, the assembly mechanisms and diversity of microbial communities shaped by such environments are poorly understood yet. In this study, we explored aquatic bacterial community diversity and ecological assembly processes in subsidence lakes during winter and summer using 16S rRNA gene sequencing. We observed that clear bacterial community structure was driven by seasonality more than by habitat, and the α-diversity and functional diversity of the bacterial community in summer were significantly higher than in winter (p < 0.001). Canonical correspondence analysis indicated that temperature and chlorophyll-a were the most crucial contributing factors influencing the community season variations in subsidence lakes. Specifically, temperature and chlorophyll-a explained 18.26 and 14.69% of the community season variation, respectively. The bacterial community variation was driven by deterministic processes in winter but dominated by stochastic processes in summer. Compared to winter, the network of bacterial communities in summer exhibited a higher average degree, modularity, and keystone taxa (hubs and connectors in a network), thereby forming a highly complex and stable community structure. These results illustrate the clear season heterogeneity of bacterial communities in subsidence lakes and provide new insights into revealing the effects of seasonal succession on microbial assembly processes in coal mining subsidence lake ecosystems.

Compositional changes and ecological characteristics of earthworm mucus under different electrical stimuli.

Earthworm mucus is rich in nutrients that can initiate the mineralization and humification of organic matter and is of great importance for contaminated soil remediation and sludge reutilization. In this study, six voltage and current combinations were utilized to promote earthworm mucus production (5 V and 6 V at 10, 20 and 30 mA, respectively), to explore the compositional changes of the mucus produced under different electrical stimuli, and to propose the best electrical stimulation group and mucus fraction applicable to soil heavy metal pollution remediation and sludge reutilization. The results showed that the mucus produced by the six electrical stimuli was mainly composed of proteins, amino acids, carbohydrates, fatty acids, and polysaccharides, with small amounts of alcohol, phenol, and ester organic substances. Under different electrical stimuli, each component changed significantly (P < 0.05). pH and conductivity were higher at 6 V 20 mA, total nitrogen and phosphorus contents reached their maximum at 5 V 30 mA, and total potassium at 6 V 10 mA. Protein, amino acids, and carbohydrates were most abundant in the mucus produced at 5 V 10 mA, while trace metal elements reached their lowest values at 5 V 10 mA. Finally, based on principal component analysis and combined with previous studies, it was concluded that the mucus produced at 5 V 10 mA was weakly alkaline, high in amino acids and nutrients and low in trace metal elements, and most suitable for sludge and straw composting experiments, soil remediation and amendment experiments.

Phospholipase C delta 1 inhibits WNT/ß-catenin and EGFR-FAK-ERK signaling and is disrupted by promoter CpG methylation in renal cell carcinoma.

BACKGROUND: PLCD1, located at 3p22, encodes an enzyme that mediates cellular metabolism and homeostasis, intracellular signal transduction and movement. PLCD1 plays a pivotal role in tumor suppression of several types of cancers; however, its expression and underlying molecular mechanisms in renal cell carcinoma (RCC) pathogenesis remain elusive. METHODS: RT-PCR and Western blot were used to detect PLCD1 expression in RCC cell lines and normal tissues. Bisulfite treatment, MSP and BGS were utilized to explore the CpG methylation status of PLCD1 promoter. Online databases were analyzed for the association between PLCD1 expression/methylation and patient survival. In vitro experiments including CCK8, colony formation, wound-healing, transwell migration and invasion, immunofluorescence and flow cytometry assays were performed to evaluate tumor cell behavior. Luciferase assay and Western blot were used to examine effect of PLCD1 on WNT/ß-catenin and EGFR-FAK-ERK signaling. RESULTS: We found that PLCD1 was widely expressed in multiple adult normal tissues including kidney, but frequently downregulated or silenced in RCC due to its promoter CpG methylation. Restoration of PLCD1 expression inhibited the viability, migration and induced G2/M cell cycle arrest and apoptosis in RCC cells. PLCD1 restoration led to the inhibition of signaling activation of WNT/ß-catenin and EGFR-FAK-ERK pathways, and the EMT program of RCC cells. CONCLUSIONS: Our results demonstrate that PLCD1 is a potent tumor suppressor frequently inactivated by promoter methylation in RCC and exerts its tumor suppressive functions via suppressing WNT/ß-catenin and EGFR-FAK-ERK signaling. These findings establish PLCD1 as a promising prognostic biomarker and treatment target for RCC.

Roles of oncogenes in esophageal squamous cell carcinoma and their therapeutic potentials.

Esophageal squamous cell carcinoma (ESCC) is the most common type of esophageal cancer (EC) in Asia. It is a malignant digestive tract tumor with abundant gene mutations. Due to the lack of specific diagnostic markers and early cancer screening markers, most patients are diagnosed at an advanced stage. Genetic and epigenetic changes are closely related to the occurrence and development of ESCC. Here, We review the activation of proto-oncogenes into oncogenes through gene mutation and gene amplification in ESCC from a genetic and epigenetic genome perspective, We also discuss the specific regulatory mechanisms through which these oncogenes mainly affect the biological function and occurrence and development of ESCC through specific regulatory mechanisms. In addition, we summarize the clinical application value of these oncogenes is summarized, and it provides a feasible direction for clinical use as potential therapeutic and diagnostic markers.

Ecological Risk Assessment and Source Apportionment of Heavy Metals in the Soil of an Opencast Mine in Xinjiang.

To study the influence of open-pit coal mining on the surrounding soil environment and human health, this study selected the Hongshaquan coal mine in Xinjiang as the research area and took 31 soil samples from the dump and artificial forest of the mining area. The contents of seven heavy metals (As, Cd, Cr, Cu, Ni, Pb and Zn) in the soil were analyzed. The pollution index method, geoaccumulation index method (Igeo), potential ecological risk index method, health ecological risk assessment model and principal component analysis (PCA) were used to evaluate and analyze the heavy metal pollution, potential ecological risk and health ecological risk of the soil. The results showed that compared with the background value of soil in Xinjiang, except for Pb, other heavy metal elements were essentially pollution-free and belonged to the low ecological risk area. The health risk assessment model showed that Pb and As were the main pollution factors of noncarcinogenic risk, and that exposure to Ni, Pb and As had a lower carcinogenic risk. The PCA showed that Cu, Cr, Ni, Pb, As and Zn in the dump were from transportation and industrial activities, Cd was from natural resources, and Cr, Zn, Ni, Cd and Pb were from transportation in the artificial forest. Cu came from industrial sources and As from soil parent material. The dump was more seriously disturbed by human factors than by artificial forests. Our research provides a reference for heavy metal pollution and source analysis caused by mining.

Application of Rice Husk Biochar and Earthworm on Concentration and Speciation of Heavy Metals in Industrial Sludge Treatment.

The aim of this study was to assess the total concentration and speciation variation of heavy metals (Pb, Cd, Cu and Zn) during composting and vermicomposting of industrial sludge with different addition rations of rice husk biochar. Results indicated that pH, EC, total phosphorus (TP) and total potassium (TK) were increased and total organic carbon (TOC) and total nitrogen (TN) were decreased during the composting of industrial sludge with biochar compared with the control (sludge without biochar). The addition of earthworm to the biochar-amended sludge further decreased pH and TOC but highly enhanced the EC, TN, TP and TK. Comparatively lower concentrations of total and DTPA-extractable heavy metals were observed in biochar-amended sludge treatments mixed with earthworm in comparison with the biochar-amended sludge treatments without earthworm or the control. Sequential extraction methods demonstrated that vermicomposting of sludge with biochar converted more metals bound with exchangeable, carbonate and organic matter into the residual fraction in comparison with those composting treatments of sludge with biochar. As a result, the combination of rice husk biochar and earthworm accelerated the passivation of heavy metals in industrial sludge during vermicomposting. Rice husk biochar and earthworm can play a positive role in sequestering the metals during the treatment of industrial sludge. This research proposed a potential method to dispose the heavy metals in industrial sludge to transform waste into resource utilization.

Identification of bromodomain-containing proteins prognostic value and expression significance based on a genomic landscape analysis of ovarian serous cystadenocarcinoma.

Background: Ovarian serous cystadenocarcinoma (OSC), a common gynecologic tumor, is characterized by high mortality worldwide. Bromodomain (BRD)-containing proteins are a series of evolutionarily conserved proteins that bind to acetylated Lys residues of histones to regulate the transcription of multiple genes. The ectopic expression of BRDs is often observed in multiple cancer types, but the role of BRDs in OSC is still unclear. Methods: We performed the differential expression, GO enrichment, GSEA, immune infiltration, risk model, subtype classification, stemness feature, DNA alteration, and epigenetic modification analysis for these BRDs based on multiple public databases. Results: Most BRDs were dysregulated in OSC tissues compared to normal ovary tissues. These BRDs were positively correlated with each other in OSC patients. Gene alteration and epigenetic modification were significant for the dysregulation of BRDs in OSC patients. GO enrichment suggested that BRDs played key roles in histone acetylation, viral carcinogenesis, and transcription coactivator activity. Two molecular subtypes were classified by BRDs for OSC, which were significantly correlated with stemness features, m6A methylation, ferroptosis, drug sensitivity, and immune infiltration. The risk model constructed by LASSO regression with BRDs performed moderately well in prognostic predictions for OSC patients. Moreover, BRPF1 plays a significant role in these BRDs for the development and progression of OSC patients. Conclusion: BRDs are potential targets and biomarkers for OSC patients, especially BRPF1.

The role of serum amyloid A1 in the adipogenic differentiation of human adipose-derived stem cells basing on single-cell RNA sequencing analysis.

BACKGROUND: Adipose-derived stem cells (ASCs) are obtained from a variety of sources in vivo where they present in large quantities. These cells are suitable for use in autologous transplantation and the construction of tissue-engineered adipose tissue. Studies have shown that ASCs differentiation is in a high degree of heterogeneity, yet the molecular basis including key regulators of differentiation remains to clarify. METHODS: We performed single-cell RNA sequencing and bioinformatics analysis on both undifferentiated (ASC-GM group) and adipogenically differentiated human ASCs (ASC-AD group, ASCs were cultured in adipogenic inducing medium for 1 week). And then, we verified the results of serum amyloid A1 (SAA1) with western blotting, immunofluorescence staining, oil red O staining. After these experiments, we down-regulated the expression of serum amyloid A1 (SAA1) gene to verify the adipogenic differentiation ability of ASCs. RESULTS: In single-cell RNA sequence analyzing, we obtained 4415 cells in the ASC-GM group and 4634 cells in the ASC-AD group. The integrated sample cells could be divided into 11 subgroups (0-10 cluster). The cells in cluster 0, 2, 5 were came from ASC-GM group and the cells in cluster 1, 3, 7 came from ASC-AD group. The cells of cluster 4 and 6 came from both ASC-GM and ASC-AD groups. Fatty acid binding protein 4, fatty acid binding protein 5, complement factor D, fatty acid desaturase 1, and insulin like growth factor binding protein 5 were high expressed in category 1 and 7. Regulation of inflammatory response is the rank 1 biological processes. And cellular responses to external stimuli, negative regulation of defense response and acute inflammatory response are included in top 20 biological processes. Based on the MCODE results, we found that SAA1, C-C Motif Chemokine Ligand 5 (CCL5), and Annexin A1 (ANXA1) significantly highly expressed during adipogenic differentiation. Western blot and immunofluorescent staining results showed that SAA1 increased during adipogenesis. And the area of ORO positive staining in siSAA1 cells was significantly lower than in the siControl (negative control) cells. CONCLUSIONS: Our results also indicated that our adipogenic induction was successful, and there was great heterogeneity in the adipogenic differentiation of ASCs. SAA1 with the regulation of inflammatory response were involved in adipogenesis of ASCs based on single-cell RNA sequencing analysis. The data obtained will help to elucidate the intrinsic mechanism of heterogeneity in the differentiation process of stem cells, thus, guiding the regulation of self-renewal and differentiation of adult stem cells.

Phytoplankton-Zooplankton Community Structure in Coal Mining Subsidence Lake.

Land subsidence from coal mining has shaped new artificial aquatic ecosystems, these subsidence lakes are known for their restricted ecological system, water pollution, and extreme habitat conditions. However, knowledge concerning the community structure of plankton in these types of water bodies is still limited. Therefore, both phytoplankton and zooplankton communities' abundance, distribution, and diversity, as well as relations of these communities to physicochemical water quality variables were analyzed, alongside the interaction between phytoplankton and zooplankton groups. The results indicate zooplankton abundance was 842.375 to 186,355.0 ind./L. Biomass ranged from 0.3408 to 10.0842 mg/L. Phytoplankton abundance varied between 0.541 × 106 cell/L and 52.340 × 106 cell/L while phytoplankton wet biomass ranged from 0.5123 to 5.6532 mg/L. Pearson correlation analysis revealed that both the zooplankton and phytoplankton total densities were significantly correlated with nutrients (TN, TP, PO43-) and CODcr; zooplankton abundance was significantly correlated with phytoplankton abundance. According to the biodiversity index of Shannon-Wiener, both phytoplankton and zooplankton revealed less biodiversity in the subsidence water region than in the Huihe river system and Xiangshun canal, with values ranging from 0.20 to 2.60 for phytoplankton and 1.18 to 2.45 for zooplankton; however, the phytoplankton community showed lower biodiversity index values compared to the zooplankton community. Overall, the knowledge gleaned from the study of plankton community structure and diversity represents a valuable approach for the evaluation of the ecological conditions within the subsidence lakes, which has significant repercussions for the management and protection of aquatic environments in mining areas.

Lamin A/C-Dependent Translocation of Megakaryoblastic Leukemia-1 and ß-Catenin in Cyclic Strain-Induced Osteogenesis.

Lamins are intermediate filaments that play a crucial role in sensing mechanical strain in the nucleus of cells. ß-catenin and megakaryoblastic leukemia-1 (MKL1) are critical signaling molecules that need to be translocated to the nucleus for their transcription in response to mechanical strain that induces osteogenesis. However, the exact molecular mechanism behind the translocation of these molecules has not been fully investigated. This study used 10% cyclic strain to induce osteogenesis in the murine osteoblast precursor cell line (MC3T3). The translocation of ß-catenin and MKL1 was studied by performing knockdown and overexpression of lamin A/C (LMNA). Cyclic strain increased the expression of osteogenic markers such as alkaline phosphatase (ALP), runt-related transcription factor 2 (RUNX2), and enhanced ALP staining after seven days of incubation. Resultantly, MKL1 and ß-catenin were translocated in the nucleus from the cytoplasm during the stress-induced osteogenic process. Knockdown of LMNA decreased the accumulation of MKL1 and ß-catenin in the nucleus, whereas overexpression of LMNA increased the translocation of these molecules. In conclusion, our study indicates that both MKL1 and ß-catenin molecules are dependent on the expression of LMNA during strain-induced osteogenesis.

Regulation of the integrin αVß3- actin filaments axis in early osteogenesis of human fibroblasts under cyclic tensile stress.

BACKGROUND: Integrins play a prominent role in osteogenic differentiation by transmitting both mechanical and chemical signals. Integrin expression is closely associated with tensile stress, which has a positive effect on osteogenic differentiation. We investigated the relationship between integrin αVß3 and tensile stress. METHODS: Human fibroblasts were treated with c (RGDyk) and lentivirus transduction to inhibit function of integrin αVß3. Y-15, cytochalasin D and verteporfin were used to inhibit phosphorylation of FAK, polymerization of microfilament and function of nuclear YAP, respectively. Fibroblasts were exposed to a cyclic tensile stress of 10% at 0.5 Hz, once a day for 2 h each application. Fibroblasts were harvested on day 4 and 7 post-treatment. The expression of ALP, RUNX2, integrin αVß3, ß-actin, talin-1, FAK, vinculin, and nuclear YAP was detected by Western blot or qRT-PCR. The expression and distribution of integrin αVß3, vinculin, microfilament and nuclear YAP. RESULTS: Cyclic tensile stress was found to promote expression of ALP and RUNX2. Inhibition of integrin αVß3 activation downregulated the rearrangement of microfilament and the expression of ALP, RUNX2 and nuclear YAP. When the polymerization of microfilament was inhibited the expression of ALP, RUNX2 and nuclear YAP were decreased. The phosphorylation of FAK induced by cyclic tensile stress reduced by the inhibition of integrin αVß3. The expression of ALP and RUNX2 was decreased by inhibition of phosphorylation of FAK and inhibition of nuclear YAP. CONCLUSIONS: Cyclic tensile stress promotes osteogenesis of human fibroblasts via integrin αVß3-microfilament axis. Phosphorylation of FAK and nuclear YAP participates in this process.

Single-cell transcriptomic sequencing analyses of cell heterogeneity during osteogenesis of human adipose-derived mesenchymal stem cells.

Mesenchymal stem cells (MSCs) are known for their multilineage differentiation potential with immune-modulatory properties. The molecular underpinnings of differentiation remain largely undefined. In this study, we investigated the cellular and molecular features of chemically induced osteogenesis from MSC isolated from human adipose tissue (human adipose MSCs, hAMSCs) using single-cell RNA-sequencing (scRNA-seq). We found that a near complete differentiation of osteogenic clusters from hAMSCs under a directional induction. Both groups of cells are heterogeneous, and some of the hAMSCs cells are intrinsically prepared for osteogenesis, while variant OS clusters seems in cooperation with a due division of the general function. We identified a set of genes related to cell stress response highly expressed during the differentiation. We also characterized a series of transitional transcriptional waves throughout the process from hAMSCs to osteoblast and specified the unique gene networks and epigenetic status as key markers of osteogenesis.

Mechanical Sensing Element PDLIM5 Promotes Osteogenesis of Human Fibroblasts by Affecting the Activity of Microfilaments.

Human skin fibroblasts (HSFs) approximate the multidirectional differentiation potential of mesenchymal stem cells, so they are often used in differentiation, cell cultures, and injury repair. They are an important seed source in the field of bone tissue engineering. However, there are a few studies describing the mechanism of osteogenic differentiation of HSFs. Here, osteogenic induction medium was used to induce fibroblasts to differentiate into osteoblasts, and the role of the mechanical sensitive element PDLIM5 in microfilament-mediated osteogenic differentiation of human fibroblasts was evaluated. The depolymerization of microfilaments inhibited the expression of osteogenesis-related proteins and alkaline phosphatase activity of HSFs, while the polymerization of microfilaments enhanced the osteogenic differentiation of HSFs. The evaluation of potential protein molecules affecting changes in microfilaments showed that during the osteogenic differentiation of HSFs, the expression of PDLIM5 increased with increasing induction time, and decreased under the state of microfilament depolymerization. Lentivirus-mediated PDLIM5 knockdown by shRNA weakened the osteogenic differentiation ability of HSFs and inhibited the expression and morphological changes of microfilament protein. The inhibitory effect of knocking down PDLIM5 on HSF osteogenic differentiation was reversed by a microfilament stabilizer. Taken together, these data suggest that PDLIM5 can mediate the osteogenic differentiation of fibroblasts by affecting the formation and polymerization of microfilaments.

Actin polymerization state regulates osteogenic differentiation in human adipose-derived stem cells.

BACKGROUND: Actin is an essential cellular protein that assembles into microfilaments and regulates numerous processes such as cell migration, maintenance of cell shape, and material transport. METHODS: In this study, we explored the effect of actin polymerization state on the osteogenic differentiation of human adipose-derived stem cells (hASCs). The hASCs were treated for 7 days with different concentrations (0, 1, 5, 10, 20, and 50 nM) of jasplakinolide (JAS), a reagent that directly polymerizes F-actin. The effects of the actin polymerization state on cell proliferation, apoptosis, migration, and the maturity of focal adhesion-related proteins were assessed. In addition, western blotting and alizarin red staining assays were performed to assess osteogenic differentiation. RESULTS: Cell proliferation and migration in the JAS (0, 1, 5, 10, and 20 nM) groups were higher than in the control group and the JAS (50 nM) group. The FAK, vinculin, paxillin, and talin protein expression levels were highest in the JAS (20 nM) group, while zyxin expression was highest in the JAS (50 nM) group. Western blotting showed that osteogenic differentiation in the JAS (0, 1, 5, 10, 20, and 50 nM) group was enhanced compared with that in the control group, and was strongest in the JAS (50 nM) group. CONCLUSIONS: In summary, our data suggest that the actin polymerization state may promote the osteogenic differentiation of hASCs by regulating the protein expression of focal adhesion-associated proteins in a concentration-dependent manner. Our findings provide valuable information for exploring the mechanism of osteogenic differentiation in hASCs.

Spatial organization and crosstalk of vimentin and actin stress fibers regulate the osteogenic differentiation of human adipose-derived stem cells.

Human adipose-derived stem cells (hASCs) are ideal seed cells for tissue engineering due to their multidirectional differentiation potential. Microfilaments, microtubules, and intermediate filaments are responsible for supporting the intracellular space. Vimentin, a type III intermediate filament protein that is specifically expressed in cells of mesenchymal origin, can function as a scaffold and endow cells with tension and shear stress resistance. Actin stress fibers (ASF) act as an important physical device in stress signal transduction, providing stiffness for cells, and promoting osteogenesis. Through direct physical contact, cross-linkers, and spatial interactions, vimentin and actin networks exist as intersecting entities. Spatial interactions occur in the overlapping area of cytoskeleton subsystems, which could affect cell morphology, cell mechanics, and cell fate. However, how does the spatial organization between the cytoskeletal subsystems changed during osteogenesis, especially between vimentin and ASF, is still not understood, and its mechanism effect on cell fate remains unclear. In our study, WB experiment was used to detect the expression changes in Vimentin, ASF, and other proteins. Cells were reconstructed by three-dimensional scanning with fluorescence microscope, and the spatial thickness of vimentin and ASF cytoskeletons and the thickness of the overlapping area between them were calculated, respectively, so as to observe the spatial reorganization of vimentin and ASF in cells. Cytochalasin D (an inhibitor of actin polymerization) and vimentin upregulated/downregulated cells were used to verify the change in the spatial organization between vimentin and ASF and its influence on osteogenesis. Then, heat shock protein 27 (HSP27) was downregulated to illuminate the regulatory mechanisms of spatial organization between vimentin and ASF during osteogenesis. The amounts and the spatial positions of vimentin and actin stress fiber exhibited opposite trends during osteogenesis. Through controlling the anchor sites on the nucleus, intermediate filaments vimentin can reduce the spatial proportion of actin stress fibers, which can be regulated by HSP27. In addition, depolymerization of actin stress fibers lead to lower osteogenic differentiation ability, resulting in osteogenesis and lipogenesis existed simultaneously, that can be resisted by vimentin. Our data indicate that the spatial reorganization of vimentin and actin stress fibers is a key factor in the regulation of the differentiation state of hASCs. And their spatial overlapping area is detrimental to hASCs osteogenesis, providing a new perspective for further exploring the mechanism underlying hASCs osteogenesis.