Mesenchymal Stromal Cells from the Maternal Segment of Human Umbilical Cord is Ideal for Bone Regeneration in Allogenic Setting

Abstract

Umbilical cord (UC) is a discarded product from the operating theatre and a ready source of mesenchymal stromal cells (MSCs). MSCs from UC express both embryonic and adult mesenchymal stem cell markers and are known to be hypoimmunogenic and non-tumorigenic and thus suitable for allogeneic cell transplantation. Our study aimed to determine the degree of immunotolerance and bone-forming capacity of osteodifferentiated human Wharton's jelly-derived mesenchymal stromal cells (hWJ-MSCs) from different segments of UC in an allogenic setting. UCs were obtained from healthy donors delivering a full-term infant by elective Caesarean section. hWJ-MSCs were isolated from 3 cm length segment from the maternal and foetal ends of UCs. Three-dimensional fibrin constructs were formed and implanted intramuscularly into immunocompetent mice. The mice were implanted with 1) fibrin construct with maternal hWJ-MSCs, 2) fibrin construct with foetal hWJ-MSCs, or 3) fibrin without cells; the control group received sham surgery. After 1 month, the lymphoid organs were analysed to determine the degree of immune rejection and bone constructs were analysed to determine the amount of bone formed. A pronounced immune reaction was noted in the fibrin group. The maternal segment constructs demonstrated greater osteogenesis than the foetal segment constructs. Both maternal and foetal segment constructs caused minimal immune reaction and thus appear to be safe for allogeneic bone transplant. The suppression of inflammation may be a result of increased anti-inflammatory cytokine production mediated by the hWJ-MSC. In summary, this study demonstrates the feasibility of using bone constructs derived from hWJ-MSCs in an allogenic setting.

Keywords: Allogeneic; Bone; Mesenchymal stromal cells; Tissue engineering; Wharton’s jelly.

Fig. 1

Representative lymphoid organs isolated from the A Control group, B Maternal segment group, C Foetal segment group and D Fibrin group. Lymph nodes, spleen and thymus are indicated by the bold, dashed and normal arrows, respectively

Fig. 2

Change in the size of bone construct over the 30-day implantation period. Gross morphology of bone construct before and after implantation. (S1-sample 1, S2-sample 2, S3-sample 3)

Fig. 3

Alizarin red staining of calcium deposition in A Maternal bone construct, B Foetal bone construct and C Fibrin construct. The apparent stained area was an artefact due to tissue folding. D Surface area of bone nodules formed on the maternal and foetal segment constructs. White area is empty spaces. This is most likely due to spreading of tissue section during histological preparation. Arrows indicate calcium deposition. Values are presented as mean ± SEM, n = 3. *Represents statistical significance at p = 0.04

Fig. 4

H&E staining of longitudinal sections of the spleen. A Control specimen with follicle size < 1.22 mm. B Maternal segment construct specimen showing mild lymphocytic proliferation. C Foetal segment construct specimen showing moderate lymphocytic proliferation. D Fibrin construct specimen showing severe lymphocyte proliferation. E Higher magnification of a fibrin construct specimen with arrows indicating multinucleated giant cells. F Higher magnification of a fibrin construct specimen with arrows showing apoptotic bodies in the white pulp of the spleen

Fig. 5

H&E staining of longitudinal sections of lymph nodes. A Control specimen without lymphoblasts. B Maternal segment construct specimen with mild lymphocyte proliferation. C Foetal segment construct specimen with moderate lymphocyte proliferation. D Fibrin construct specimen with severe lymphocyte proliferation. In addition, fibrin construct specimens also showed E Vacuolar degeneration and F Sinusoids

Fig. 6

H&E staining of longitudinal sections of the thymus. A Control specimen with minimal histiocytosis. B Maternal segment construct specimen showing minimal lymphocytic proliferation. C Foetal segment construct specimen showing moderate lymphocyte proliferation. D Fibrin construct specimen showing severe lymphocyte proliferation. E Higher magnification of a fibrin segment with an arrow indicating lymphocytic apoptosis

Fig. 7

Measured cytokines normalised against control group levels. A Changes in pro-inflammatory and B Anti-inflammatory cytokines detected in the blood are presented relative to control group levels. *Represents a difference (p < 0.05) from the control group, ^#represents a difference (p < 0.05) relative to the fibrin group, ^$represents a difference (p < 0.05) relative to the maternal segment group. (MT maternal segment group, FT foetal segment group)