Culture of rat mesenchymal stem cells on PHBV-PCL scaffolds: analysis of conditioned culture medium by FT-Raman spectroscopy

Mesenchymal stem cells (MSCs) have great potential for application in cell therapy and tissue engineering procedures because of their plasticity and capacity to differentiate into different cell types. Given the widespread use of MSCs, it is necessary to better understand some properties related to osteogenic differentiation, particularly those linked to biomaterials used in tissue engineering. The aim of this study was to develop an analysis method using FT-Raman spectroscopy for the identification and quantification of biochemical components present in conditioned culture media derived from MSCs with or without induction of osteogenic differentiation. All experiments were performed between passages 3 and 5. For this analysis, MSCs were cultured on scaffolds composed of bioresorbable poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) and poly(ε-caprolactone) (PCL) polymers. MSCs (GIBCO®) were inoculated onto the pure polymers and 75:25 PHBV/PCL blend (dense and porous samples). The plate itself was used as control. The cells were maintained in DMEM (with low glucose) containing GlutaMAX® and 10% FBS at 37oC with 5% CO2 for 21 days. The conditioned culture media were collected and analyzed to probe for functional groups, as well as possible molecular variations associated with cell differentiation and metabolism. The method permitted to identify functional groups of specific molecules in the conditioned medium such as cholesterol, phosphatidylinositol, triglycerides, beta-subunit polypeptides, amide regions and hydrogen bonds of proteins, in addition to DNA expression. In the present study, FT-Raman spectroscopy exhibited limited resolution since different molecules can express similar or even the same stretching vibrations, a fact that makes analysis difficult. There were no variations in the readings between the samples studied. In conclusion, FT-Raman spectroscopy did not meet expectations under the conditions studied.

Study on Protein Nanomarker Combined with Vascular Endothelial Growth Factor to Improve Vascularization of Rabbit Urethral Defect Tissue Engineering

To construct the tissue engineering urethral material that is closest to the normal urethral structure in the true sense in vitro. Abdominal ADSC from a 2-month-old New Zealand white rabbit was extracted and directly compounded with non-woven polyglycolic acid (PGA) (control group) to induce the differentiation of myoblasts and epithelial-like cells in vitro and shaped into urethral structure lumen Observation group); After Gd chelating protein nano-labeling and VEGF-loaded sustained release, the rabbit model of a long urethral defect was replanted and cultured for 4 weeks, 8 weeks and 12 weeks, respectively. There was no difference in urinary tract patency rate, urinary tract infection, and renal dysfunction rate between the two groups (P > 0.05). The urine flow rate in the observation group was significantly higher than that in the control group, and the residual volume decreased (P < 0.05). The blood vessel density and CD31 percentage in the observation group increased (P < 0.05). Compared with the conventional ADSC directly in contact with the composite material to construct the urethra, in vitro induction of ADSC to myoblasts and epithelial-like cells respectively, and then use the cell membrane technology to build a tissue engineering urethral material that is closest to the normal urethral structure in the true sense, and loaded with VEGF Loop release technology can significantly improve urodynamic functions, optimize tissue engineering urethral structure and vascularization, and is expected to become a new technology for constructing new tissue engineering urethral materials.