isolating-sequencing-analyzing-extracellular-micrornas-from-human

Isolating, Sequencing and Analyzing Extracellular MicroRNAs from Human Mesenchymal Stem Cells

Extracellular and circulating RNAs (exRNA) are produced by many cell types of the body and exist in numerous bodily fluids such as saliva, plasma, serum, ...

Aarhus University

Bone remodeling and regeneration are highly regulated multistep processes involving posttranscriptional regulation by microRNAs (miRNAs). Here, we performed a global profiling of differentially expressed miRNAs in bone-marrow-derived skeletal cells (BMSCs; also known as stromal or mesenchymal stem cells) during in vitro osteoblast differentiation. We functionally validated the regulatory effects of several miRNAs on osteoblast differentiation and identified 15 miRNAs, most significantly miR-222 and miR-423, as regulators of osteoblastogenesis. In addition, we tested the possible targeting of miRNAs for enhancing bone tissue regeneration. Scaffolds functionalized with miRNA nano-carriers enhanced osteoblastogenesis in 3D culture and retained this ability at least 2 weeks after storage. Additionally, anti-miR-222 enhanced in vivo ectopic bone formation through targeting the cell-cycle inhibitor CDKN1B (cyclin-dependent kinase inhibitor 1B). A number of additional miRNAs exerted additive osteoinductive effects on BMSC differentiation, suggesting that pools of miRNAs delivered locally from an implanted scaffold can provide a promising approach for enhanced bone regeneration.

Global MicroRNA Profiling in Human Bone Marrow Skeletal-Stromal or Mesenchymal-Stem Cells Identified Candidates for Bone Regeneration.

 Non-invasive tracking of transplanted cells is critical in evaluating delivery, migration and prognosis of cell therapies.  We formulated a nano-contrast agent consisting of a perfluorooctylbromide (PFOB) core within a shell of poly (lactic-co-glycolic acid) (PLGA) followed by a coat of polystyrene sulfonate (PSS) for F MRI. The nano-contrast agent (PSS-NP) was characterised by DLS and the uptake efficiency of the nano-contrast agent (PSS-NP) was tested using flow cytometry,  MRI and confocal microscopy.  and  assays of labelled cells were tested for their ability to provide an MRI signal while retaining their osteoblastic differentiation capabilities.  PSS-NPs were internalised via caveolae-mediated endocytosis in mesenchymal stromal/stem cells without affecting cell proliferation and differentiation in osteoblasts, both  and . Furthermore, labelled cells were monitored by F MRI for up to 2 months after transplantation in mice. In particular, PSS-NP-labelled cells can be used to monitor the enhanced immune rejection of grafted human cells in normal BALB/c mice compared to immune-compromised NOD/SCID mice. One week after transplantation, 40% of the F MRI signal was lost in normal mice, whereas only 10% was lost in immune-compromised mice.  Overall, these results show that PSS-NPs can label MSCs effectively, and be employed  as a novel nano-contrast agent for non-invasive cell tracking using clinically relevant F MRI techniques.

Caveolae-mediated mesenchymal stem cell labelling by PSS-coated PLGA PFOB nano-contrast agent for MRI.

The prospective study is to investigate the associations between serum testosterone levels and LUTS among middle-aged men ≥40 years receiving health check-up. Lower urinary tract symptoms were evaluated by the self-administered International Prostate Symptom Score questionnaire. Serum prostate specific antigen and total testosterone level were checked in all subjects. A total of 1752 men were enrolled into the study. The mean age was 55.6 ± 9.7 years. All study subjects were stratified into low, medium and high testosterone levels by two cut-off, 3.0 and 4.11 ng/mL. We found that testosterone levels were significantly associated with metabolic syndrome and body fat components. Compared to those with low testosterone levels, subjects with high and medium testosterone had a significantly higher IPSS (5.84 ± 5.55 vs 6.71 ± 5.68 and 6.34 ± 5.66, p = 0.032) and storage score (2.76 ± 2.29 vs 3.20 ± 2.49 and 2.90 ± 2.49; p = 0.009), and a more moderate/severe LUTS (IPSS ≧ 8) (26.5% vs 35.7% and 29.9%; p = 0.002). Multivariate analyses showed that high vs low testosterone levels (OR, 1.76; 95% CI, 1.26-2.45) and prostate volume ≧25 vs <25 mL (OR, 1.38; 95% CI, 1.04-1.82) significantly associated with the presence of moderate/severe LUTS. Pearson correlation analyses showed significantly positive correlations between testosterone level and IPSS in whole study sample (Pearson correlation coefficient, 0.066; p < 0.01) and in the subgroup of moderate/severe LUTS (Pearson correlation coefficient, 0.038; p < 0.05). In conclusion, high testosterone and prostate volume adversely impacted LUTS in our target population.

Article	Year
Global MicroRNA Profiling in Human Bone Marrow Skeletal-Stromal or Mesenchymal-Stem Cells Identified Candidates for Bone Regeneration. Molecular therapy : the journal of the American Society of Gene Therapy\| PubMed ID: 29331291	2018
Caveolae-mediated mesenchymal stem cell labelling by PSS-coated PLGA PFOB nano-contrast agent for MRI. Theranostics\| PubMed ID: 29774066	2018
Factors Associated with Bothersome Lower Urinary Tract Symptoms in Middle-Aged Men Receiving Health Checkup. Scientific reports\| PubMed ID: 30696907	2019

Chi-Chih Chang

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