RAD: a web application to identify region associated differentially expressed genes.

SUMMARY: With the advance of genomic sequencing techniques, chromatin accessible regions, transcription factor binding sites and epigenetic modifications can be identified at genome-wide scale. Conventional analyses focus on the gene regulation at proximal regions; however, distal regions are usually less focused, largely due to the lack of reliable tools to link these regions to coding genes. In this study, we introduce RAD (Region Associated Differentially expressed genes), a user-friendly web tool to identify both proximal and distal region associated differentially expressed genes (DEGs). With DEGs and genomic regions of interest (gROI) as input, RAD maps the up- and down-regulated genes associated with any gROI and helps researchers to infer the regulatory function of these regions based on the distance of gROI to differentially expressed genes. RAD includes visualization of the results and statistical inference for significance. AVAILABILITY AND IMPLEMENTATION: RAD is implemented with Python 3.7 and run on a Nginx server. RAD is freely available at https://labw.org/rad as online web service. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Comprehensive assessment of mRNA isoform detection methods for long-read sequencing data.

The advancement of Long-Read Sequencing (LRS) techniques has significantly increased the length of sequencing to several kilobases, thereby facilitating the identification of alternative splicing events and isoform expressions. Recently, numerous computational tools for isoform detection using long-read sequencing data have been developed. Nevertheless, there remains a deficiency in comparative studies that systemically evaluate the performance of these tools, which are implemented with different algorithms, under various simulations that encompass potential influencing factors. In this study, we conducted a benchmark analysis of thirteen methods implemented in nine tools capable of identifying isoform structures from long-read RNA-seq data. We evaluated their performances using simulated data, which represented diverse sequencing platforms generated by an in-house simulator, RNA sequins (sequencing spike-ins) data, as well as experimental data. Our findings demonstrate IsoQuant as a highly effective tool for isoform detection with LRS, with Bambu and StringTie2 also exhibiting strong performance. These results offer valuable guidance for future research on alternative splicing analysis and the ongoing improvement of tools for isoform detection using LRS data.

Exosomes from miR-29a-modified adipose-derived mesenchymal stem cells reduce excessive scar formation by inhibiting TGF-ß2/Smad3 signaling.

Pathological scars mainly refer to hypertrophic scars and keloids, and have a high incidence. Moreover, these scars seriously affect the patient's appearance and are associated with significant pain. The present study aimed to investigate the inhibitory effect of microRNA (miR)­29a from human adipose­derived mesenchymal stem cells (hADSCs) exosomes on scar formation. Firstly, the expression of miR­29a in thermal skin tissues of mice and human hypertrophic scar fibroblasts (HSFBs) was detected via reverse transcription­quantitative PCR. Exosomes derived from miR­29a­modified hADSCs were extracted and the influence of miR­29a­modified hADSCs­exo on the proliferation and function of HSFBs was determined. Lastly, the effect of miR­29a­modified hADSCs­exo on scar formation was determined using a thermal mouse model. The results demonstrated that miR­29a was downregulated in scar tissues after scalding and in HSFBs. After treating HSFBs with miR­29a­modified hADSC exosomes, miR­29a­overexpressing hADSC exosomes inhibited the proliferation and migration of HSFBs. Moreover, it was found that TGF­ß2 was the target of miR­29a, and that hADSC exosome­derived miR­29a inhibited the fibrosis of HSFBs and scar hyperplasia after scalding in mice by targeting the TGF­ß2/Smad3 signaling pathway. In summary, the current data indicated that miR­29a­modified hADSC exosome therapy can decrease scar formation by inhibiting the TGF­ß2/Smad3 signaling pathway via its derived exogenous miR­29a, and this may be useful for the future treatment of pathological scars by providing a potential molecular basis.

Alterations of 63 hub genes during lingual carcinogenesis in C57BL/6J mice.

To identify potential biomarkers of lingual cancer, 75 female C57BL/6J mice were subjected to 16-week oral delivery of 4-nitroquinoline-1-oxide (4NQO; 50 mg/L), with 10 mice used as controls. Lingual mucosa samples representative of normal tissue (week 0) and early (week 12) and advanced (week 28) tumorigenesis were harvested for microarray and methylated DNA immunoprecipitation sequencing (MeDIP-Seq). Combined analysis with Short Time-series Expression Miner (STEM), the Cytoscape plugin cytoHubba, and screening of differentially expressed genes enabled identification of 63 hub genes predominantly altered in the early stage rather than the advanced stage. Validation of microarray results was carried out using qRT-PCR. Of 63 human orthologous genes, 35 correlated with human oral squamous cell carcinoma. KEGG analysis showed "pathways in cancer", involving 13 hub genes, as the leading KEGG term. Significant alterations in promoter methylation were confirmed at Tbp, Smad1, Smad4, Pdpk1, Camk2, Atxn3, and Cdh2. HDAC2, TBP, and EP300 scored ≥10 on Maximal Clique Centrality (MCC) in STEM profile 11 and were overexpressed in human tongue cancer samples. However, expression did not correlate with smoking status, tumor differentiation, or overall survival. These results highlight potentially useful candidate biomarkers for lingual cancer prevention, diagnosis, and treatment.

[Peripheral blood endothelial progenitor cells enhance the expressions of SDF-1 and MCP-1 of bone marrow stromal cells and promote their homing ability].

OBJECTIVE: To investigate the effects of peripheral blood endothelial progenitor cells (PB-EPCs) on the homing ability of bone marrow stromal cells (BMSCs) as well as the potential mechanism. METHODS: BMSCs were injected intravenously with lentiviral expression vector expressing enhanced green fluorescent protein (EGFP) for tracing. Biological bone graft was made to repair rabbit radial defect. In the experimental group, PB-EPCs and BMSCs mixed at a ratio of 1:1 were combined with partially deproteinized bone (PDPB) for implantation to repair rabbit models with radial bone defect. BMSCs alone were combined with PDPB in the control group. The models in the blank group were not repaired. Protein and mRNA levels of endogenous stromal-derived factor-1 (SDF-1) and monocyte chemotactic protein-1 (MCP-1) were evaluated by ELISA and real-time quantitative PCR at 2, 4, 8 weeks after the operation. At the same time points, immunohistochemical staining was performed to detect EGFP expression in the defect sites. RESULTS: The mRNA and protein levels of SDF-1 and MCP-1 in the experimental group were higher than those in the other two groups. Immunohistochemistry showed that the number of EGFP-positive cells was larger in the experimental group than in the control or the blank group. CONCLUSION: PB-EPCs can increase the expressions of SDF-1 and MCP-1 and promote the migration of EGFP-positive BMSCs to bone defect site.

Effects of PB-EPCs on Homing Ability of Rabbit BMSCs via Endogenous SDF-1 and MCP-1.

UNLABELLED: Traumas, infections, tumors, and some congenital malformations can lead to bone defects or even bone loss. The goal of the present study was to investigate whether inclusion of endothelial progenitor cells derived from peripheral blood (PB-EPCs) in cell-seeded partially deproteinized bone (PDPB) implants would stimulate recruitment of systemically injected bone marrow stromal cells (BMSCs) to the implant. METHODS: BMSCs were injected intravenously with lentiviral expression vector expressing enhanced green fluorescent protein (eGFP) for tracing. Recruitment of eGFP-positive BMSCs was tested for the following implant configurations: 1) seeded with both BMSC and PB-EPC, 2) BMSC alone, 3) PB-EPC alone, and 4) unseeded PDPB. Protein and mRNA levels of endogenous stromal-derived factor-1 (SDF-1) and its receptor CXCR4, as well as monocyte chemotactic protein-1 (MCP-1) and its receptor CCR2, were evaluated on the 8th week. Immunohistochemical staining was performed to determine eGFP-positive areas at the defective sites. Masson's trichrome staining was conducted to observe the distribution of collagen deposition and evaluate the extent of osteogenesis. RESULTS: The mRNA and protein levels of SDF-1 and CXCR4 in the co-culture group were higher than those in other groups (p < 0.05) 8 weeks after the surgery. MCP-1 mRNA level in the co-culture group was also higher than that in the other groups (p < 0.05). Immunohistochemical assays revealed that the area covered by eGFP-positive cells was larger in the co-culture group than in the other groups (p < 0.05) after 4 weeks. Masson's trichrome staining revealed better osteogenic potential of the co-culture group compared to the other groups (p < 0.05). CONCLUSION: These experiments demonstrate an association between PB-EPC and BMSC recruitment mediated by the SDF-1/CXCR4 axis that can enhance repair of bone defects.

The effect of ultraviolet radiation on the transforming growth factor beta 1/Smads pathway and p53 in actinic keratosis and normal skin.

Ultraviolet (UV) radiation is considered to be essential for the progression of actinic keratosis (AK) to squamous cell carcinoma (SCC); however, the mechanisms have not been fully elucidated. To understand this process, the effects of UV radiation on the transforming growth factor beta 1 (TGFß1)/Smads pathway and p53 in normal skin and AK were studied. Normal human skin and AK tissues were cultured and divided into the following four groups according to the UV radiation dose: 0 (control group), 5, 10, and 20 J/cm2. The tissues were radiated for four consecutive days; 24 h after radiation, the tissues were collected for investigation. Compared with the control group, greater proliferative inhibition and apoptosis were induced by UV radiation in normal skin than AK. The expression of TGFß1, Smad7, and p53 was increased in AK and normal skin, while the level of TßRII was decreased. Smad2 was reduced in AK only. The expressions of TßRI, Smad3, and Smad4 were not significantly changed. The results demonstrated that although p53 was induced, suppression of the TGFß1/Smads pathway by UV radiation might contribute to the progression of AK to SCC.