Pro-Inflammatory Cytokines, IFNγ and TNFα, Influence Immune Properties of Human Bone Marrow and Wharton Jelly Mesenchymal Stem Cells Differentially

BACKGROUND: Wharton's jelly derived stem cells (WJMSCs) are gaining attention as a possible clinical alternative to bone marrow derived mesenchymal stem cells (BMMSCs) owing to better accessibility, higher expansion potential and low immunogenicity. Usage of allogenic mesenchymal stem cells (MSC) could be permissible in vivo only if they retain their immune properties in an inflammatory setting. Thus the focus of this study is to understand and compare the immune properties of BMMSCs and WJMSCs primed with key pro-inflammatory cytokines, Interferon-gamma (IFNgamma) and Tumor Necrosis Factor-alpha (TNFalpha). METHODOLOGY/PRINCIPAL FINDINGS: Initially the effect of priming on MSC mediated suppression of alloantigen and mitogen induced lymphoproliferation was evaluated in vitro. Treatment with IFNgamma or TNFalpha, did not ablate the immune-suppression caused by both the MSCs. Extent of immune-suppression was more with WJMSCs than BMMSCs in both the cases. Surprisingly, priming BMMSCs enhanced suppression of mitogen driven lymphoproliferation only; whereas IFNgamma primed WJMSCs were better suppressors of MLRs. Further, kinetic analysis of cytokine profiles in co-cultures of primed/unprimed MSCs and Phytohematoagglutinin (PHA) activated lymphocytes was evaluated. Results indicated a decrease in levels of pro-inflammatory cytokines. Interestingly, a change in kinetics and thresholds of Interleukin-2 (IL-2) secretion was observed only with BMMSCs. Analysis of activation markers on PHA-stimulated lymphocytes indicated different expression patterns in co-cultures of primed/unprimed WJMSCs and BMMSCs. Strikingly, co-culture with WJMSCs resulted in an early activation of a negative co-stimulatory molecule, CTLA4, which was not evident with BMMSCs. A screen for immune suppressive factors in primed/unprimed WJMSCs and BMMSCs indicated inherent differences in IFNgamma inducible Indoleamine 2, 3-dioxygenase (IDO) activity, Hepatocyte growth factor (HGF) and Prostaglandin E-2 (PGE2) levels which could possibly influence the mechanism of immune-modulation. CONCLUSION/SIGNIFICANCE: This study demonstrates that inflammation affects the immune properties of MSCs distinctly. Importantly different tissue derived MSCs could utilize unique mechanisms of immune-modulation

Aseptic Raman spectroscopy can detect changes associated with the culture of human dental pulp stromal cells in osteoinductive culture.

There is an unmet need for the non-invasive characterisation of stem cells to facilitate the translation of cell-based therapies. Raman spectroscopy has proven utility in stem cell characterisation but as yet no method has been reported capable of taking repeated Raman measurements of living cells aseptically over time. The aim of this study was to determine if Raman spectroscopy could be used to monitor changes in a well characterised cell population (human dental pulp stromal cells (DPSCs)) by taking repeated Raman measurements from the same cell populations in osteoinductive culture over time and under aseptic conditions. DPSCs were isolated from extracted premolar teeth from 3 consenting donors. Following in vitro expansion, DPSCs were maintained for 28 days in osteo-inductive medium. Raman spectra were acquired from the cells at days 0, 3, 7, 10, 14 and 28. Principal component analysis (PCA) was carried out to assess if there was any temporal spectral variation. At day 28, osteoinduction was confirmed using alizarin red staining and qRT-PCR for alkaline phosphatase and osteocalcin. Alizarin red staining was positive in all samples at day 28 and significant increases in alkaline phosphatase (p < 0.001) and osteocalcin (p < 0.05) gene expression were also observed compared with day 0. PCA of the Raman data demonstrated trends in PC1 from days 0-10, influenced by protein associated features and PC2 from days 10-28, influenced by DNA/RNA associated features. We conclude that spectroscopy can be used to monitor changes in Raman signature with time associated with the osteoinduction of DPSCs using repeated measurements via an aseptic methodology

Secretome of mesenchymal stem cells and its impact on Chronic Obstructive Pulmonary Disease

Chronic obstructive pulmonary disease (COPD) is characterized by irreversible loss of lung function that stem from two mechanisms, inflammation and senescence. Crosstalk between these two mechanisms accelerate the development of COPD, thus targeting these two pathways may offer benefits in the treatment of COPD. Growing amount of evidence have shown that mesenchymal stem cells as a promising candidate for the treatment of COPD. Over the years, many studies conducted to decipher the therapeutic effect of MSC in COPD and the mechanisms involve, in the hope of utilizing these cells as new therapeutic strategy for COPD. However, the cell-based therapy by using the MSC presented with many obstacles including low engraftment at the site of injury, the risk of microvascular occlusion, unwanted differentiation, and also the risk of malignant transformation. Recently, recently researchers begin to look at the possibility of using MSC derived extracellular vesicles as an alternative to MSC. Here we review the effect of MSC and MSC derived EV in modulating inflammation, and senescence in COPD. We also review current treatment and the side effect in COPD, and senolytic drugs, a new therapeutic strategy targeting the senescent cells

Abrazivní frézování vodním paprskem pro letecké aplikace.

The diploma thesis deals with the statistical study of determining the optimal input process parameters for the implementation of mask-less milling on difficult-to-machine material - Stainless steel AISI 304 by Abrasive Waterjet technology. In this study, the effect of traverse speed, TS = 1000mm/min and 3000mm/min, abrasive flow rate, AFR = 100g/min and 200g/min, stand-off distance, SOD = 4mm and 15mm for the respective step-over distance, SD = 0.6mm and 0.7mm were studied on the output responses of width, depth, surface roughness parameters Ra and Rz. A total of 16 experiments using full factorial design classified under the two step-over distances were performed and the experimental results were analyzed using the Minitab software. The results demonstrated the maximum possible width is achieved by the execution of lower TS, higher AFR and higher SOD; the maximum possible depth is achieved by the execution of lower TS, higher AFR and lower SOD; and the minimum possible Ra and Rz is achieved by the execution of higher TS, higher AFR and lower SOD. By performing ‘desirability approach’ optimization, the optimum parameter configuration with respect to both step-over distance 0.6mm and 0.7mm was found with TS = 1000mm/min, AFR = 200g/min, and SOD = 4mm. The study concludes the utilization of multiple input variables within the experimental domain to achieve more accurate results as per the desired output requirements.Diplomová práce se zabývá statistickou studií stanovení optimálních vstupních parametrů procesu pro realizaci bezmaskového frézování obtížně obrobitelného materiálu - nerezové oceli AISI 304 technologií abrazivního vodního paprsku. V této studii byl zkoumán vliv rychlosti pojezdu, TS = 1000 mm/min a 3000 mm/min, průtoku abraziva, AFR = 100 g/min a 200 g/min, vzdálenost trysky od materiálu, SOD = 4 mm a 15 mm pro příslušnou vzdálenost kroku, SD = 0,6 mm a 0,7 mm na výstupní odezvy šířky, hloubky, parametrů drsnosti povrchu Ra a Rz. Bylo provedeno celkem 16 experimentů s využitím plného faktorového uspořádání klasifikovaných podle dvou vzdáleností odstupňování a výsledky experimentů byly analyzovány pomocí softwaru Minitab. Výsledky ukázaly, že maximální možné šířky je dosaženo při provedení nižšího TS, vyššího AFR a vyššího SOD; maximální možné hloubky je dosaženo při provedení nižšího TS, vyššího AFR a nižšího SOD; a minimálních možných Ra a Rz je dosaženo při provedení vyššího TS, vyššího AFR a nižšího SOD. Provedením optimalizace podle přístupu "desirability approach" byla nalezena optimální konfigurace parametrů s ohledem na vzdálenost kroku 0,6 mm i 0,7 mm při TS = 1000 mm/min, AFR = 200 g/min a SOD = 4 mm. Studie uzavírá využití více vstupních proměnných v rámci experimentální oblasti k dosažení přesnějších výsledků podle požadovaných výstupních požadavků.346 - Katedra obrábění, montáže a strojírenské metrologievýborn

Functional differences in mesenchymal stromal cells from human dental pulp and periodontal ligament

10.1111/jcmm.12192Journal of Cellular and Molecular Medicine182344-35