The study objectives include, enhancing the proliferations of aged bone marrow stem cells (BMSCs) and adipose stem cells (ADSCs); and evaluating the shelf lives of clinical grade chondrogenically induced cells from both samples. ADSCs and BMSCs from 56 patients (76 ± 8 yrs) were proliferated using basal medium (FD) and at (5, 10, 15, 20 and 25) ng/ml of basal fibroblast growth factor (bFGF). They were induced to chondrogenic lineage and stored for more than 120 hrs in FD, serum, Dulbecco's phosphate buffered saline (DPBS) and saline at 4 °C. In FD, cells stagnated and BMSCs' population doubling time (PDT) was 137 ± 30 hrs, while ADSCs' was 129.7 ± 40 hrs. bFGF caused PDT's decrease to 24.5 ± 5.8 hrs in BMSCs and 22.0 ± 6.5 hrs in ADSCs (p = 0.0001). Both cells were positive to stem cell markers before inductions and thereafter, expressed significantly high chondrogenic genes (p = 0.0001). On shelf life, both cells maintained viabilities and counts above 70% in FD and serum after 120 hrs. BMSCs' viabilities in DPBS fell below 70% after 96 hrs and saline after 72 hrs. ADSCs' viability fell below 70% in DPBS after 24 hrs and saline within 24 hrs. Concentrations between 20 ng/ml bFGF is ideal for aged adult cells' proliferation and delivery time of induced BMSCs and ADSCs can be 120 hrs in 4 °C serum.
Hematopoietic stem cells- (HSCs-) based therapy requires ex vivo expansion of HSCs prior to therapeutic use. However, ex vivo culture was reported to promote excessive production of reactive oxygen species (ROS), exposing HSCs to oxidative damage. Efforts to overcome this limitation include the use of antioxidants. In this study, the role of Hibiscus sabdariffa L. (Roselle) in maintenance of cultured murine bone marrow-derived HSCs was investigated. Aqueous extract of Roselle was added at varying concentrations (0-1000 ng/mL) for 24 hours to the freshly isolated murine bone marrow cells (BMCs) cultures. Effects of Roselle on cell viability, reactive oxygen species (ROS) production, glutathione (GSH) level, superoxide dismutase (SOD) activity, and DNA damage were investigated. Roselle enhanced the survival (P < 0.05) of BMCs at 500 and 1000 ng/mL, increased survival of Sca-1(+) cells (HSCs) at 500 ng/mL, and maintained HSCs phenotype as shown from nonremarkable changes of surface marker antigen (Sca-1) expression in all experimental groups. Roselle increased (P < 0.05) the GSH level and SOD activity but the level of reactive oxygen species (ROS) was unaffected. Moreover, Roselle showed significant cellular genoprotective potency against H2O2-induced DNA damage. Conclusively, Roselle shows novel property as potential supplement and genoprotectant against oxidative damage to cultured HSCs.
The aim of this study was to produce cloned caprine embryos using either caprine bone marrow-derived mesenchymal stem cells (MSCs) or ear fibroblast cells (EFCs) as donor karyoplasts. Caprine MSCs were isolated from male Boer goats of an average age of 1.5 years. To determine the pluripotency of MSCs, the cells were induced to differentiate into osteocytes, chondrocytes and adipocytes. Subsequently, MSCs were characterized through cell surface antigen profiles using specific markers, prior to their use as donor karyoplasts for nuclear transfer. No significant difference (p > 0.05) in fusion rates was observed between MSCs (87.7%) and EFCs (91.3%) used as donor karyoplasts. The cleavage rate of cloned embryos derived with MSCs (87.0%) was similar (p > 0.05) to those cloned using EFCs (84.4%). However, the in vitro development of MSCs-derived cloned embryos (25.3%) to the blastocyst stage was significantly higher (p < 0.05) than those derived with EFCs (20.6%). In conclusion, MSCs could be reprogrammed by caprine oocytes, and production of cloned caprine embryos with MSCs improved their in vitro developmental competence, but not in their fusion and cleavage rate as compared to cloning using somatic cells such as EFCs.
The use of mesenchymal stem cells (MSCs) for cartilage repair has generated much interest owing to their multipotentiality. However, their significant presence in peripheral blood (PB) has been a matter of much debate. The objectives of this study are to isolate and characterize MSCs derived from PB and, compare their chondrogenic potential to MSC derived from bone marrow (BM). PB and BM derived MSCs from 20 patients were isolated and characterized. From 2 ml of PB and BM, 5.4 ± 0.6 million and 10.5 ± 0.8 million adherent cells, respectively, were obtained by cell cultures at passage 2. Both PB and BM derived MSCs were able to undergo tri-lineage differentiation and showed negative expression of CD34 and CD45, but positively expressed CD105, CD166, and CD29. Qualitative and quantitative examinations on the chondrogenic potential of PB and BM derived MSCs expressed similar cartilage specific gene (COMP) and proteoglycan levels, respectively. Furthermore, the s-GAG levels expressed by chondrogenic MSCs in cultures were similar to that of native chondrocytes. In conclusion, this study demonstrates that MSCs from PB maintain similar characteristics and have similar chondrogenic differentiation potential to those derived from BM, while producing comparable s-GAG expressions to chondrocytes.