Inducible indoleamine 2,3-dioxygenase 1 and programmed death ligand 1 expression as the potency marker for mesenchymal stromal cells.

AIM: Establishment of a potency assay in the manufacturing of clinical-grade mesenchymal stromal cells (MSCs) has been a challenge due to issues of relevance to function, timeline and variability of responder cells. In this study, we attempted to develop a potency assay for MSCs. METHODS: Clinical-grade bone marrow-derived MSCs were manufactured. The phenotype and immunosuppressive functions of the MSCs were evaluated based on the International Society for Cellular Therapy guidelines. Resting MSCs licensed by interferon (IFN)-γ exposure overnight were evaluated for changes in immune suppression and immune-relevant proteins. The relationship of immune-relevant protein expression with immunosuppression of MSCs was analyzed. RESULTS: MSC supressed third-party T-lymphocyte proliferation with high inter-donor and inter-test variability. The suppression of T-lymphocyte proliferation by IFN-γ-licensed MSCs correlated with that by resting MSCs. Many cellular proteins were up-regulated after IFN-γ exposure, including indoleamine 2,3-dioxygenase 1 (IDO-1), programmed death ligand 1 (PD-L1), vascular cell adhesion molecule 1 (VCAM-1), intercellular adhesion molecule 1 (ICAM-1) and bone marrow stromal antigen 2 (BST-2). The expression levels of IDO-1 and PD-L1 on licensed MSCs, not VCAM-1, ICAM-1 or BST-2 on licensed MSCs, correlated with MSC suppression of third-party T-cell proliferation. CONCLUSION: A flow cytometry-based assay of MSCs post-IFN-γ exposure measuring expression of intracellular protein IDO-1 and cell surface protein PD-L1 captures two mechanisms of suppression and offers the potential of a relevant, rapid assay for MSC-mediated immune suppression that would fit with the manufacturing process.

Knowledge and awareness about dentistry among medical students, residents, and medical practitioners in Bhairahawa city, Nepal.

INTRODUCTION: Oral health is considered to be a mirror of systemic health and vice-versa. Medical professionals are considered to be the primary caregivers in a given population and as such a vast majority of the population visits them for health-related problems. Therefore, this study aimed to evaluate the level of knowledge and awareness of dentistry among first-year medical students, residents, and medical practitioners. METHODS: A cross-sectional questionnaire survey was carried out to assess the knowledge and awareness of dentistry among MBBS first-year students, medical residents, and medical practitioners of Bhairahawa city, Nepal. Data analysis was done with SPSS version 11. RESULTS: As per the demographic characteristics out of 147 subjects 82 (55.8%) were male and 65 (44.2%) were female. Most of the participants were from the age group of 21-30 years, 66 (44.9%) and 115 (78.2%) of them were married. Among 147 subjects every respondent (100%) knew about the profession of dentistry giving statistically significant results. CONCLUSION: The present study shows that medical professionals had better knowledge of dentistry as compared to medical students and residents. This unnecessary gap between dental and medical professionals can only be fulfilled by including dental education in both undergraduate and postgraduate medical curriculum.

Are serum-free and xeno-free culture conditions ideal for large scale clinical grade expansion of Wharton's jelly derived mesenchymal stem cells? A comparative study.

INTRODUCTION: Mesenchymal stromal/stem cells (MSCs) for clinical use have largely been isolated from the bone marrow, although isolation of these cells from many different adult and fetal tissues has been reported as well. One such source of MSCs is the Whartons Jelly (WJ) of the umbilical cord, as it provides an inexhaustible source of stem cells for potential therapeutic use. Isolation of MSCs from the umbilical cord also presents little, if any, ethical concerns, and the process of obtaining the cord tissue is relatively simple with appropriate consent from the donor. However, a great majority of studies rely on the use of bovine serum containing medium for isolation and expansion of these cells, and porcine derived trypsin for dissociating the cells during passages, which may pose potential risks for using these cells in clinical applications. It is therefore of high priority to develop a robust production process by optimizing culture variables to efficiently and consistently generate MSCs that retain desired regenerative and differentiation properties while minimizing risk of disease transmission. METHODS: We have established a complete xeno-free, serum-free culture condition for isolation, expansion and characterization of WJ-MSCs, to eliminate the use of animal components right from initiation of explant culture to clinical scale expansion and cryopreservation. Growth kinetics, in vitro differentiation capacities, immunosuppressive potential and immunophenotypic characterization of the cells expanded in serum-free media have been compared against those cultured under standard fetal bovine serum (FBS) containing medium. We have also compared the colony-forming frequency and genomic stability of the large scale expanded cells. Secretome analysis was performed to compare the angiogenic cytokines and functional angiogenic potency was proved by Matrigel assays. RESULTS: Results presented in this report identify one such serum-free, xeno-free medium for WJ expansion. Cells cultured in serum-free, xeno-free medium exhibit superior growth kinetics and functional angiogenesis, alongside other MSC characteristics. CONCLUSIONS: We report here that WJ-MSCs cultured and expanded in Mesencult XF, SF Medium retain all necessary characteristics attributed to MSC for potential therapeutic use.