Project description:Exosomes and exosomal miRNAs from the plasma of volunteers were isolated from thyroid nodules and papillary tyriod cancer patients . Profiling of exosomal miRNA was performed using next-generation sequencing(NGS) to identify miRNA candidates for diagnosis.
Project description:To explore the role of the mitochondrial Ca2+ uniporter (MCU) in exosomal microRNA(miRNA) sorting in MDA-MB-231 cells, we isolated miRNA from the exosomes derived from MCU-downregulated and control MDA-MB-231 cells. We then performed Next Generation Sequencing (NGS) and real-time quantitative polymerase chain reaction (qRT-PCR) to find and validate differetial expressed exosomal miRNAs that regulated by MCU.
Project description:We report the application of miRNA next generation sequencing (NGS) for the analysis of impact of processing on miRNA in human breast milk, donated by 3 volunteers. MiRNA content of total and exosomal fraction was compared between unprocessed milk and sample subjected to either Holder (thermal) pasteurization (HoP) or elevated pressure processing (HPP). NGS reads were mapped to miRBase in order to obtain miRNA counts. Then, we analyzed differences in the miRNA abundance and function between raw and processed material. It was observed that both processing methods reduce number of miRNA reads and HoP is significantly more detrimental to miRNA than HPP.
Project description:Primary ovarian insufficiency (POI) is an early decline in ovarian function that leads to infertility. Conventional treatments for chemotherapy-induced POI are unsatisfactory. Mesenchymal stem cells (MSCs) have emerged as a therapeutic option, but the impact of estrogen niche-respond MSC secretome on ovarian regeneration and circadian rhythm remains unknown. This study revealed that the secretome of ER+pcMSCs (conditioned medium [CM] and E2-CM, respectively) significantly reduced the CTX-induced defects in ovarian folliculogenesis and circadian rhythm. The CM/E2-CM also reduced granulosa cell apoptosis and rescued angiogenesis in POI ovarian tissues. E2-CM presented a better effect than the CM. Cytokine array analysis showed a significant increase in cytokine/growth factors associated with immunomodulation and angiogenesis (including angiogenin). Neutralizing angiogenin in the CM/E2-CM significantly decreased its ability to promote HUVEC tube formation in vitro. Importantly, the ER+pcMSC secretome restored the CTX-induced circadian rhythm defects, including the expression of both the genes and proteins associated with ovarian circadian clock (such as Rora, E4bp4, Rev-erbα, and Dbp) and the locomotor activity. Further exosomal miRNA analysis showed the involved miRNAs in targeting the genes associated with POI rescue (Pten and Pdcd4), Caspase-3, estrogen synthesis (Cyp19a1), and importantly the ovarian clock regulation (E4bp4, Rev-erbα and Rev-erbβ).
Project description:Background/Aim: We investigated alterations in the expression of serum exosomal miRNAs with the progression of liver fibrosis and evaluated their clinical applicability as biomarkers. Methods: This study prospectively enrolled 71 patients who underwent liver biopsy at an academic hospital in Korea. Exosomes were extracted from serum samples, followed by next-generation sequencing (NGS) of miRNAs and targeted real-time quantitative polymerase chain reaction. A model was derived to discriminate advanced fibrosis based on miRNA levels and the performance of this model was evaluated. Validation of the effect of miRNA on liver fibrosis in vitro was followed. Methods: This study prospectively enrolled 71 patients who underwent liver biopsy at an academic hospital in Korea. Exosomes were extracted from serum samples, followed by next-generation sequencing (NGS) of miRNAs and targeted real-time quantitative polymerase chain reaction. A model was derived to discriminate advanced fibrosis based on miRNA levels and the performance of this model was evaluated. Validation of the effect of miRNA on liver fibrosis in vitro was followed. Results: NGS data revealed that exosomal miR-660-5p, miR-125a-5p, and miR-122 expression were changed significantly with the progression of liver fibrosis, of which miR-122 exhibited high read counts enough to be used as a biomarker. The level of exosomal miR-122 decreased as the pathologic fibrosis grade progressed and patients with biopsy-proven advanced fibrosis had significantly lower levels of exosomal miR-122 (P < 0.001) than those without advanced fibrosis. Exosomal miR-122 exhibited a fair performance in discriminating advanced fibrosis especially in combination with fibrosis-4 score and transient elastography. In a subgroup of patients with a non-viral etiology of liver disease, the performance of exosomal miR-122 as a biomarker was greatly improved. Inhibition of miR-122 expression increased the proliferation of the human hepatic stellate cell line, LX-2, and upregulated the expression of various fibrosis related proteins. Conclusion: Exosomal miR-122 may serve as a useful non-invasive biomarker for liver fibrosis, especially in patients with non-viral etiologies of chronic liver disease.
Project description:The subpopulations of mesenchymal stem cells isolated from breast adipose tissue which display migrated towards conditioned media from MDA-MB231 cell line were expanded for miRNA analysis. The tumor microenvironment (TM) is known to promote tumor growth and progression. Ubiquitously distributed tissue resident stem cells (MSCs) elicit regenerative properties. In addition, they are capable of homing to sites of inflammation, injury, and tumor. Considering the tumor tropic property of MSCs, the interaction between the breast cancer (BC) microenvironment and breast resident adipose tissue derived MSCs (ASCs) merits further investigations. Initial data indicate that a subset of ASCs derived from breast adipose tissue (B-ASCs) display a high affinity towards the conditioned media (CM) from two breast cancer cell lines, MDA-MB231 (MDA-CM) and MCF7 (MCF-CM). Profiling secreted cytokines of these CMs identified significant expression of angiogenin, GM-CSF, and IL-6. While the expression of GRO-α, MCP-1, RANTES, and IL-1α is more pronounced in MDA-CM, MCF-CM contains higher amounts of IGFBP2, TRAIL, and ErbB3. Gene expression profiling suggests that despite the distinct differences, both migratory subset of B-ASCs towards MDA-CM and MCF-CM display perturbed expression of genes like KISS1, TNSF1, IL18 and MMP2, which could be associated with a defensive role of B-ASCs. In addition, the BC microenvironment alters microRNA (miRNA) expressions in B-ASCs. in this study the migratory cells were evaluated for miRNA expression versus non-migratory counterparts. as controls unexposed parental cell lines (2) were used on the same hybridization chip in which one labeled Hy3 and other labeled Hy5. The ratio of the control parental cells was used as base nanalysis of miRNA expression in MSCs. we also include another control in this study, the migratory subpopulations of MSCs which display migratory potentials against protein gradient (M5) were analyzed for miRNA expression versus non-migratory counterparts (NM-5). using this control facilitate identification of those miRNA responsive to tumor CM. the data analysis confirm that altered gene and miRNA profiles resulted from exposure of MCF-CM and MDA-CM on B-ASCs are similar to those observed in B-ASCs isolated from breast adipose tissue of BC patients. Analysis the signaling between the B-ASCs and TM may help in understanding the possible role of B-ASCs in stasis, progression, or regression of the BC.
Project description:The subpopulations of mesenchymal stem cells isolated from breast adipose tissue which display migrated towards conditioned media from MDA-MB231 cell line were expanded for miRNA analysis. The tumor microenvironment (TM) is known to promote tumor growth and progression. Ubiquitously distributed tissue resident stem cells (MSCs) elicit regenerative properties. In addition, they are capable of homing to sites of inflammation, injury, and tumor. Considering the tumor tropic property of MSCs, the interaction between the breast cancer (BC) microenvironment and breast resident adipose tissue derived MSCs (ASCs) merits further investigations. Initial data indicate that a subset of ASCs derived from breast adipose tissue (B-ASCs) display a high affinity towards the conditioned media (CM) from two breast cancer cell lines, MDA-MB231 (MDA-CM) and MCF7 (MCF-CM). Profiling secreted cytokines of these CMs identified significant expression of angiogenin, GM-CSF, and IL-6. While the expression of GRO-M-NM-1, MCP-1, RANTES, and IL-1M-NM-1 is more pronounced in MDA-CM, MCF-CM contains higher amounts of IGFBP2, TRAIL, and ErbB3. Gene expression profiling suggests that despite the distinct differences, both migratory subset of B-ASCs towards MDA-CM and MCF-CM display perturbed expression of genes like KISS1, TNSF1, IL18 and MMP2, which could be associated with a defensive role of B-ASCs. In addition, the BC microenvironment alters microRNA (miRNA) expressions in B-ASCs. in this study the migratory cells were evaluated for miRNA expression versus non-migratory counterparts. as controls unexposed parental cell lines (2) were used on the same hybridization chip in which one labeled Hy3 and other labeled Hy5. The ratio of the control parental cells was used as base nanalysis of miRNA expression in MSCs. we also include another control in this study, the migratory subpopulations of MSCs which display migratory potentials against protein gradient (M5) were analyzed for miRNA expression versus non-migratory counterparts (NM-5). using this control facilitate identification of those miRNA responsive to tumor CM. the data analysis confirm that altered gene and miRNA profiles resulted from exposure of MCF-CM and MDA-CM on B-ASCs are similar to those observed in B-ASCs isolated from breast adipose tissue of BC patients. Analysis the signaling between the B-ASCs and TM may help in understanding the possible role of B-ASCs in stasis, progression, or regression of the BC. The microRNA expression of migratory subpopulations were compared to parental populations of MSCs.
Project description:Human induced pluripotent stem cells provide an unlimited, scalable source of youthful tissue progenitors and secretome for regenerative therapies. The aim of our study was to assess the potential of conditioned medium (CM) derived from hiPSC-mesenchymal progenitors (hiPSC-MPs) to stimulate osteogenic differentiation of adult and aged human bone marrow-mesenchymal stromal cells (MSCs). In addition, we evaluated whether extended cultivation or osteogenic pre-differentiation of hiPSC-MPs could enhance the CM stimulatory activity.
Project description:The disturbance of adipose tissue in obesity highly correlates with cancer progression. To investigate the impacts of obesity on triple-negative breast cancer (TNBC), we performed an in vitro system culturing TNBC cells with human white adipocytes-conditioned medium (hAd-CM). The gene expression profiling between each condition was then assessed by RNA-seq.
Project description:Age-related bone degeneration is driven by structural deterioration, chronic inflammation, and impaired regenerative capacity, leading to an increased incidence of osteoarticular disorders. Mesenchymal stromal/stem cells (MSCs) have emerged as promising therapeutic candidates due to their immunomodulatory and regenerative properties, mediated largely through their secretome and extracellular vesicles (EVs). However, the therapeutic efficacy of MSC-derived products depends on both the cell source and the conditioning stimuli to which MSCs are exposed. In this study, we performed a comparative proteomic and exosomal microRNA (miRNA) profiling of secretomes derived from placenta-derived MSCs (PSCs) and adipose-derived MSCs (ASCs), and assessed the impact of priming strategies, including hypoxia, interferon-gamma (IFNγ), and interleukin-1 beta (IL1β), on paracrine properties of these cells. Proteomic analysis identified over 7,000 proteins, with PSC secretome enriched in pathways related to osteogenesis, chondrogenesis, extracellular matrix organization, angiogenesis, and immune regulation, whereas ASC secretome displayed limited enrichment in these processes. Functional scoring highlighted IL1β priming as the most effective strategy to enhance osteochondral and immunomodulatory protein signatures in PSCs. Differently, IFNγ priming selectively expanded the repertoire of exosomal miRNAs, with enrichment in signaling networks including Wnt/β-catenin, TGF-β, NF-κB, and T cell receptor pathways, underscoring its role in fine-tuning immune and regenerative functions. Together, these findings reveal that PSCs secrete a broader and more functionally relevant spectrum of bioactive molecules for osteoarticular applications compared with ASCs. Moreover, distinct priming strategies can differentially modulate their paracrine outputs. IL1β primarily enhances protein-driven regenerative and immunomodulatory activity, whereas IFNγ promotes a functionally enriched exosomal miRNA cargo. These results support PSC-derived secretomes as versatile candidates for next-generation, cell-free therapies in osteoarticular disease.