Whole transcriptome scanning and validation of negatively related genes in UC-MSCs.

Background: Human umbilical cord mesenchymal stem cells (UC-MSCs) are one of the most extensively researched stem cell types due to their potential for multi-lineage differentiation, secretion of regenerative factors, modulations of immunological activities, and the release of regenerative substances and influence immunological processes. Since UC-MSCs must be cultivated on a large scale for clinical use, selecting the appropriate storing passage, such as the usage-based passage of UC-MSCs, is critical for long-term autologous or allogeneic usage. Long-term cultivation of stem cells, on the other hand, causes them to lose their pluripotent differentiation capacity. As a result, distinguishing between high and low passages of UC-MSCs and identifying the particular variations associated with stem cells and their modes of action is essential for regenerative medicine. Therefore, we investigated the biological features and transcriptional changes of UC-MSCs over passages. Methods: UC-MSCs were isolated from the tissues of the human umbilical cord, and UC-MSCs from five passages (P1, P3, P5, P10 and P15) with three repetitions were compared and identified based on morphology, cell markers, differentiation capacity, and aging-related characteristics. It was previously assumed that the phenotype of cells before the P10 passage was stable, defined as early passage, and that culture could be continued until the 15th passage, defined as late passage. Next, the five passages of UC-MSCs were sequenced using high-throughput complete transcriptome sequencing. Fuzzy C-Means Clustering (FCM) and Weighted Gene Co-expression Network Analysis (WGCNA) were used to find hub genes, and gene silencing was performed to investigate the impact of missing genes on the stemness of UC-MSC cells. Results: UC-MSCs of different passages displayed similar surface markers, including CD73, CD105, CD90, CD34, CD45 and HLA-DR. However, the proliferation time of late-phase UC-MSCs was longer than that of early-phase UC-MSCs, and the expression of the senescence-associated (SA)-ß-gal staining marker was higher. At the same time, pluripotency markers (NANOG, OCT4, SOX2 and KIF4A) were down-regulated, and the multi-differentiation potential was reduced. Meanwhile, KIFC1 and UBE2C were down-regulated in late-phase UC-MSCs, which were involved in the maintenance of stemness. Conclusions: KIFC1 and UBE2C were highly expressed in early-UC-MSCs and showed a downward gradient trend with cell expansion in vitro. They regulated UC-MSC proliferation, colony sphere formation, multiple differentiation, stemness maintenance, and other biological manifestations. Therefore, they are anticipated to be new biomarkers for UC-MSCs quality identification in regenerative medicine applications.

Prognostic and immune infiltrative biomarkers of CENPO in pan-cancer and its relationship with lung adenocarcinoma cell proliferation and metastasis.

BACKGROUND: The centromere protein O (CENPO) is an important member of the centromere protein family. However, the role of CENPO in pan-cancer and immune infiltration has not been reported. Here, we investigated the role of CENPO in pan-cancer and further validated its role in lung adenocarcinoma (LUAD) by in vitro experiments. METHOD: The UCSC Xena database and The Cancer Genome Atlas (TCGA)-LUAD data were used to assess the expression levels of CENPO. The potential value of CENPO as a diagnostic and prognostic biomarker for pan-cancer was evaluated using TCGA data and the GEPIA database. The -expression profiles of LUAD patients and the corresponding clinical data were downloaded for correlation analysis. The role of CENPO in immune infiltration was investigated using the UCSC Xena database. Subsequently, qRT-PCR was performed to detect the expression of CENPO. Cell proliferation, migration, and invasion were determined using CCK-8, wound-healing assay, and transwell assay, respectively. RESULTS: CENPO is highly expressed in most cancers, and the upregulation of CENPO is associated with poor prognosis in many cancers. CENPO expression correlates with age, TNM stage, N stage, T stage, and receipt of radiotherapy in LUAD patients, and LUAD patients with high CENPO expression have poorer overall survival (OS) and disease-free survival (DFS). In addition, CENPO expression is associated with immune cell infiltration and immune checkpoint inhibitors. Moreover, the expression of CENPO was closely related to the expression of tumor mutational load and microsatellite instability. In vitro experiments showed that CENPO expression was increased in LUAD cell lines and that knockdown of CENPO significantly inhibited the proliferation, cell invasion, and migration ability of LUAD cells. CONCLUSION: CENPO may be a potential pan-cancer biomarker and oncogene, especially in LUAD. In addition, CENPO is associated with immune cell infiltration and may serve as a new molecular therapeutic target and effective prognostic marker for LUAD.

Preparation, Identification and Application of ß-Lactoglobulin Hydrolysates with Oral Immune Tolerance.

To reveal, for the first time, the mechanism of T cell epitope release from ß-lactoglobulin that induces oral immune tolerance, a strategy for the prediction, preparation, identification and application of ß-lactoglobulin hydrolysate with oral immune tolerance was established using the bioinformatics method, hydrolysis, mass spectrometry, T cell proliferation assays and animal experiments. Some T cell epitope peptides of ß-lactoglobulin were identified for the first time. The hydrolysates of trypsin, protamex and papain showed oral tolerance, among which the hydrolysates of protamex and papain have been reported for the first time. Although the neutral protease hydrolysate contained T cell epitopes, it still had allergenicity. The mechanism behind oral immune tolerance induction by T cell epitopes needs to be further revealed. In addition, the trypsin hydrolysate with abundant T cell epitopes was added to whey protein to prepare the product for oral immune tolerance. Overall, this study provides insights into the development of new anti-allergic milk-based products and their application in the clinical treatment of milk allergies.

CXCL8 Up-Regulated LSECtin through AKT Signal and Correlates with the Immune Microenvironment Modulation in Colon Cancer.

BACKGROUND: The role of CXCL8 and LSECtin in colon cancer liver metastasis and immune checkpoint inhibitors (ICIs) treatment effect were widely recognized. However, the regulatory role of CXCL8 on LSECtin is still unclear. METHODS: The expression of CXCL8 or LSECtin was analyzed by TCGA database, and verified by GES110225 and clinical samples. The relationship between the expression of CXCL8 or LSECtin and immune cells infiltration, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, Gene Ontology (GO) items, stromal score, Estimation of STromal and Immune cells in MAlignant Tumours (ESTIMAT) immune score, tumor mutation burden (TMB), mismatch repair gene and immune checkpoints expression were analyzed by Spearman. The effects of CXCL8 on LSECtin expression, proliferation, and invasion ability were clarified by recombinant CXCL8 or CXCL8 interfering RNA. RESULTS: In colon cancer, the expression of CXCL8 was higher, but LSECtin was lower than that in normal mucosa. The expression of CXCL8 or LSECtin was significantly positively correlated with immune cells infiltration, stromal score, ESTIMATE immune score, TMB, and immune checkpoints expression. The expression of LSECtin was closely related to the cytokine-cytokine receptor interaction pathway and response of chemokine function, such as CXCL8/CXCR1/2 pathway. There was a significant positive correlation between the expression of CXCL8 and LSECtin in colon cancer. CXCL8 up-regulated LSECtin through AKT signal and promoted the proliferation and invasion ability of colon cancer. CONCLUSIONS: CXCL8 up-regulated LSECtin by activating AKT signal and correlated with the immune microenvironment modulation in colon cancer.