Transplantation of SDF-1α-loaded liver extracellular matrix repopulated with autologous cells attenuated liver fibrosis in a rat model.

Cell-based therapy and tissue engineering are promising substitutes for liver transplantation to cure end-stage liver disorders. However, the limited sources for healthy and functional cells and poor engraftment rate are main challenges to the cell-based therapy approach. On the other hand, feasibility of production and size of bioengineered tissues are primary bottlenecks in tissue engineering. Here, we induce regeneration in a rat fibrotic liver model by transplanting a natural bioengineered scaffold with a native microenvironment repopulated with autologous stem/progenitor cells. In the main experimental group, a 1 mm3 stromal derived factor-1α (SDF-1α; S) loaded scaffold from decellularized liver extracellular matrix (LEM) was transplanted (Tx) into a fibrotic liver and the endogenous stem/progenitor cells were mobilized via granulocyte colony stimulating factor (G-CSF; G) therapy. Four weeks after transplantation, changes in liver fibrosis and necrosis, efficacy of cell engraftment and differentiation, vasculogenesis, and liver function recovery were assessed in this (LEM-TxSG) group and compared to the other groups. We found significant reduction in liver fibrosis stage in the LEM-TxSG, LEM-TxS and LEM-TxG groups compared to the control (fibrotic) group. Liver necrosis grade, and alanine transaminase (ALT) and aspartate transaminase (AST) levels dramatically reduced in all experimental groups compared to the control group. However, the number of engrafted cells into the transplanted scaffold and ratio of albumin (Alb) positive cells per total incorporated cells were considerably higher in the LEM-TxSG group compared to the LEM-Tx, LEM-TxS and LEM-TxG groups. Serum Alb levels increased in the LEM-Tx, LEM-TxS, and LEM-TxG groups, and was highest in the LEM-TxSG group, which was significantly more than the fibrotic group. Small vessel formation in the LEM-TxSG group was significantly higher than the LEM-Tx and LEM-TxS groups. Totally, these findings support application of the in vivo tissue engineering approach as a possible novel therapeutic strategy for liver fibrosis.

Combination of SB431542, Chir9901, and Bpv as a novel supplement in the culture of umbilical cord blood hematopoietic stem cells.

BACKGROUND: Small molecule compounds have been well recognized for their promising power in the generation, expansion, and maintenance of embryonic or adult stem cells. The aim of this study was to identify a novel combination of small molecules in order to optimize the ex vivo expansion of umbilical cord blood-derived CD34+ cells. METHODS: Considering the most important signaling pathways involved in the self-renewal of hematopoietic stem cells, CB-CD34+ cells were expanded with cytokines in the presence of seven small molecules including SB, PD, Chir, Bpv, Pur, Pµ, and NAM. The eliminativism approach was used to find the best combination of selected small molecules for effective ex vivo expansion of CD34+ cell. In each step, proliferation, self-renewal, and clonogenic potential of the expanded cells as well as expression of some hematopoietic stem cell-related genes were studied. Finally, the engraftment potential of expanded cells was also examined by the mouse intra-uterine transplantation model. RESULTS: Our data shows that the simultaneous use of SB431542 (TGF-ß inhibitor), Chir9901 (GSK3 inhibitor), and Bpv (PTEN inhibitor) resulted in a 50-fold increase in the number of CD34+CD38- cells. This was further reflected in approximately 3 times the increase in the clonogenic potential of the small molecule cocktail-expanded cells. These cells, also, showed a 1.5-fold higher engraftment potential in the peripheral blood of the NMRI model of in utero transplantation. These results are in total conformity with the upregulation of HOXB4, GATA2, and CD34 marker gene as well as the CXCR4 homing gene. CONCLUSION: Taken together, our findings introduce a novel combination of small molecules to improve the yield of existing protocols used in the expansion of hematopoietic stem cells.

Assessment of Short-Term Engraftment Potential of Ex Vivo Expanded Hematopoietic Stem Cells Using Normal Fetal Mouse in Utero Transplantation Model.

OBJECTIVE: Ex vivo expansion is a promising strategy to overcome the low number of human umbilical cord blood hematopoietic stem cells (hUCB-HSCs). Although based on the obtained results in unnatural physiological condition of irradiated genetically immune-deficient mouse models, there has always been concern that the expanded cells have less engraftment potential. The purpose of this study was to investigate effect of common ex vivo expansion method on engraftment potential of hUCB-mononuclear cells (MNCs), using normal fetal mouse, as a model with more similarity to human physiological conditions. MATERIALS AND METHODS: In this experimental study, briefly, isolated hUCB-MNCs were cultured in common expansion medium containing stem cell factor, Flt3 ligand and thrombopoietin. The unexpanded and expanded cells were transplanted to the fetal mice on gestational days of 11.5-13.5. After administration of human hematopoiesis growth factors (hHGFs), presence of human CD45+ cells, in the peripheral blood of recipients, was assessed at various time points after transplantation. RESULTS: The expanded MNCs showed 32-fold increase in the expression of CD34+38- phenotype and about 3-fold higher clonogenic potential as compared to the uncultured cells. Four weeks after transplantation, 73% (19/26) of expanded-cell recipients and 35% (7/20) of unexpanded-cell recipients were found to be successfully engrafted with human CD45+ cells. The engraftment level of expanded MNCs was significantly (1.8-fold) higher than unexpanded cells. After hHGFs administration, the level was increased to 3.2, 3.8 and 2.6-fold at respectively 8, 12, and 16 weeks of post transplantation. The increased expression of CXCR4 protein in expanded MNCs is a likely explanation for the present findings. CONCLUSION: The presented data showed that expanded MNCs compared to unexpended cells are capable of more rapid and higher short-term engraftment in normal fetal mouse. It could also be suggested that in utero transplantation (IUT) of normal fetal mice could be an appropriate substitute for NOD/SCID mice in xenotransplantation studies.