Individual umbilical cord mesenchymal stem cells (hUCMSCs) are inexhaustible and will be harvested at an inexpensive lacking any invasive procedure. thickness, brand-new bone tissue vessel and quantity density. To conclude, hUCMSCs seeded on CPC had been proven to match the bone tissue regeneration efficiency of hBMSCs for the very first time. Both hUCMSC-CPC and hBMSC-CPC constructs produced a lot more brand-new bone tissue and blood vessels than CPC without cells. Macroporous RGD-grafted CPC with stem cell seeding is usually encouraging for craniofacial and orthopedic repairs. and were not tumorigenic . These advantages make hUCMSCs a highly attractive alternative to hBMSCs for bone regeneration. Although a few reports used hUCMSCs for bone tissue engineering research [18,22-25], there is still a lack of studies comparing the bone regenerative efficacy of hUCMSCs with hBMSCs. A scaffold serves as a template for cell attachment, proliferation, differentiation and bone growth [37,38]. However, a literature search revealed no statement on comparison of hUCMSCs with hBMSCs seeded on CPC for bone regeneration in animals. Therefore, the objectives of this study were to investigate the behavior of stem cell-seeded CPC scaffolds in an animal model, and compare the bone regeneration efficacy of hUCMSCs with hBMSCs for the first time. RGD was grafted in chitosan Rabbit Polyclonal to TRAPPC6A which was then incorporated into CPC. A gas-foaming method was Vidaza manufacturer used to produce macropores in CPC. A critical sized cranial defect model Vidaza manufacturer in athymic rats was utilized to judge and evaluate the bone tissue regeneration efficacy of hUCMSCs and hBMSCs. Three hypotheses were tested: (1) hUCMSCs and hBMSCs will have similarly good attachment and osteogenic differentiation on macroporous CPC-RGD scaffold; (2) hUCMSCs seeded on CPC will match the bone regeneration efficacy of hBMSCs which require an invasive process to harvest; (3) Both hUCMSCs and hBMSCs seeded with CPC scaffolds will generate significantly more new bone than CPC control without stem cells. 2. Materials and methods 2.1 Fabrication of RGD-grafted macroporous CPC CPC powder consisted of an equimolar mixture of TTCP (Ca4[PO4]2O) and DCPA (CaHPO4). TTCP was synthesized from a solid-state reaction between equimolar amounts of DCPA and CaCO3 (J. T. Baker, Phillipsburg, NJ), which were mixed and heated at 1500 C for 6 h in a furnace (Model 51333, Lindberg, Watertown, WI). The heated combination was quenched to room temperature, ground in a ball mill (Retsch PM4, Brinkman, NY) and sieved to obtain TTCP particles with sizes of approximately 1-80 m, with a median of 17 m. DCPA was ground for 24 h to obtain particle sizes of 0.4-3.0 m, with a median of 1 1.0 m. TTCP and DCPA powders were mixed in a blender at a molar ratio of 1 1:1 to form the CPC powder. The CPC liquid consisted of RGD-grafted chitosan mixed with distilled water at a chitosan/(chitosan + Vidaza manufacturer water) mass portion of 7.5%. RGD grafting was performed by coupling G4RGDSP (Thermo Fisher) with chitosan malate (Vanson, Redmond, WA). This was achieved by forming amide bonds between carboxyl groups in peptide and residual amine groups in chitosan using 1-Ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride (EDC, Thermo Fisher) and sulfo-N-hydroxysuccinimide (Sulfo-NHS, Thermo Fisher) as coupling brokers [37,39,40]. After dissolving G4RGDSP peptide (24.8 mg, 32.64 10?6 mol) in 0.1 mol/L of 2-(N-Morpholino) ethanesulfonic acid (MES) buffer (4 mL) (Thermo Fisher), EDC (7.52 mg, 39.2 10?6 Vidaza manufacturer mol) and Sulfo-NHS (4.14 mg, 19.52 10?6 mol) were added to the peptide solution (molar ratio of G4RGDSP:EDC:NHS = 1:1.2:0.6). The solution was incubated at room heat for 30 min to activate the terminal carboxyl group of proline. Then, this answer was added to a chitosan answer dissolved in 0.1 mol/L of MES buffer (100 mL, 1 wt%). The coupling reaction was performed for 24 h at room temperature. The products were dialyzed against distilled water using a Dialysis Cassettes (MWCO = 3.5 kDa) (Thermo Fisher) for 3 d to remove uncoupled peptides by changing water 3 times daily. Finally, the products were freeze-dried to obtain the RGD-grafted chitosan [37,39,40]. A gas-foaming method was used to fabricate.