Human Wharton's Jelly stromal cells as an allogenic cell source for bone tissue engineering for pre-clinical proof of concept

Abstract

Human Wharton’s Jelly has gained popularity amongst tissue engineers as a source of stromal cell due to its minimal associated ethical issues, easy availability and often a discarded product of conception. Human Wharton’s Jelly derived stromal cells (hWJSCs) expressed stromal cell markers of embryonic and adult origin and known to be hypoimmunogenic and non-tumourogenic. Hence, a perfect candidate for allogenic cell transplant. The objective of this study is to determine differences amongst hWJ MSCs derived from three anatomical segments (maternal, middle and fetal segment) of the same human umbilical cord (UC). Three cm-length of each segment were cultured in vitro. Growth kinetic, immunophenotyping surface expression, immunocytochemical detection of pluripotency markers and level of Human leucocyte antigen G (HLA-G) were analyzed. Quantitative PCR was used to determine embryonic and osteogenic gene expression. Chondrogenic, adipogenic and osteogenic differentiation of hWJSCs was determined. Immunophenotyping of all three segments of hWJSCs showed presence of MSC markers CD105, CD73, CD90, CD44, CD13, CD29 and HLA–ABC MHC Class I while absence for haematopoeitic markers. Maternal and fetal segments expressed higher level of HLA-G (18.38±7.1% and 19.83±11.6%). All three segments expressed Nanog and Oct 3 and 4. Higher potency of osteogenic potential was recorded for maternal and fetal segments with higher number of bone nodules of 13.0 ± 2.08 for maternal segment and fetals segment 12.76±2.07. This study has proven that the maternal and fetal segment is an ideal alternative cell source for bone tissue engineering in terms of growth rate, cell viability, expression of pluripotent embryonic markers and osteogenic markers and also differentiation potential. This was followed by invivo study to determine the safety and efficacy of using osteocytes - differentiated hWJSCs (maternal and fetal segment) in a form of 3D construct with human fibrin as a scaffold. Immunomodulation was evaluated and lymphoid organ changes were determined after one month of implatation. This study demonstrated the feasibility of using only 3cm of human umbilical cord to form a 3D bone construct. Osteo-hWJ-MSCs from the maternal segment were able to suppress the proliferation of stimulated lymphocytes in vitro, suggesting that they can tolerate inflammatory setting. The maternal segment construct had greater osteogenic capacity compared to fetal segment construct in vivo. Allogeneic fetal and maternal segment constructs might have the ability to induce anti-inflammatory cytokine production to suppress acute reaction in vivo. Presence of hWJ-MSCs from maternal and fetal segments suppressed the immune reaction evoked by fibrin. Marked immune reaction was noticed in the fibrin group. Osteodifferentiated hWJSCs from maternal segment appeared to be safer for allogenic bone transplant due minimal immune reaction from host.