Alginate-gelatin encapsulation of human endothelial cells promoted angiogenesis in in vivo and in vitro milieu.

Up to present, many advantages have been achieved in the field of cell-based therapies by applying sophisticated methodologies and delivery approaches. Microcapsules are capable to provide safe microenvironment for cells during transplantation in a simulated physiological 3D milieu. Here, we aimed to investigate the effect of alginate-gelatin encapsulation on angiogenic behavior of human endothelial cells over a period of 5 days. Human umbilical vein endothelial cells were encapsulated by alginate-gelatin substrate and incubated for 5 days. MTT and autophagy PCR array analysis were used to monitor cell survival rate. For in vitro angiogenesis analysis, cell distribution of Tie-1, Tie-2, VEGFR-1, and VEGFR-2 were detected by ELISA. In addition to in vitro tubulogenesis assay, we monitored the expression of VE-cadherin by Western blotting. The migration capacity of encapsulated HUVECs was studied by measuring MMP-2 and MMP-9 via gelatin zymography. The in vivo angiogenic potential of encapsulated HUVECs was analyzed in immune-compromised mouse implant model during 7 days post-transplantation. We demonstrated that encapsulation promoted HUVECs cell survival and proliferation. Compared to control, no significant differences were observed in autophagic status of encapsulated cells (p > 0.05). The level of Tie-1, Tie-2, VEGFR-1, and VEGFR-2 were increased, but did not reach to significant levels. Encapsulation decreased MMP-2, -9 activity and increased the VE-cadherin level in enclosed cells (p < 0.05). Moreover, an enhanced in vivo angiogenic response of encapsulated HUVECs was evident as compared to non-capsulated cells (p < 0.05). These observations suggest that alginate-gelatin encapsulation can induce angiogenic response in in vivo and in vitro conditions.

The effects of encapsulation on NK cell differentiation potency of C-kit+ hematopoietic stem cells via identifying cytokine profiles.

Natural killer cells (NK cells) can kill cancerous cells without prior sensitization. This feature makes them appealing candidates for cellular therapy. Due to the degradation rate and controlled release of these matrices, hydrogels hold great promise in cell differentiation. The study aims to investigate the effect of encapsulated alginate-gelatin on the differentiation potential of C-kit+ cells toward NK cells which are mediated by cytokines detection. Under both encapsulated and unencapsulated conditions, C-kit+ cells can differentiate into NK cells. In the following, real-time PCR and western blotting were done to investigate the mRNA and protein expression, respectively. Determine cytokine profiles from the collected culture medium conducted a Cytokine antibody array. The differentiated cells were then co-cultured with Molt-4 cells to examine the expression levels of INF-γ, TNF-α, and IL-10 using real-time-PCR. There was a substantial change in protein expression of the Notch pathway. Also, the encapsulation increased the mRNA expression of INF-γ and TNF-α in Molt-4 cells. Based on these findings, the encapsulation effects on the differentiation of C-kit+ cells toward NK cells could be related to the secreted cytokines such as interleukin-10 and INF-γ and the Notch protein expression.

Alginate/gelatin encapsulation promotes NK cells differentiation potential of bone marrow resident C-kit+ hematopoietic stem cells.

The ability of natural killer (NK) cells to destroy cancerous cells with no prior sensitization has made them attractive candidates for cell therapy. The application of hydrogels must be notified as cell delivery vehicles in cell differentiation. The present study was conducted to investigate the effect of alginate-gelatin encapsulation on NK cell differentiation potential of C-kit+ cells. C-kit+ cells were differentiated to NK cells under both encapsulated and un-encapsulated conditions. Next, the cells were subjected to real-time polymerase chain reaction (PCR) and western blotting for the assessment of their telomere length and protein expressions, respectively. Afterward, culture medium was collected to measure cytokines levels. Thereafter, the differentiated NK cells were co-cultured with Molt-4 cells to investigate the potency of cell apoptosis by Annexin V/PI assay. A significant change was observed in the protein expression of Janus kinase/Signal transducers (JAK/STAT) pathway components. Additionally, the encapsulation caused an increase in the apoptosis of Molt-4 cells and telomere length of NK cells differentiated C-kit+ cells. Therefore, it can be concluded that the effects of encapsulation on NK cell's differentiation of C-kit+ cells could be resulted from the secreted cytokines of interleukin (IL)-2, IL-3, IL-7, and IL-12 as well as the increased telomere length.

The effect of alginate-gelatin encapsulation on the maturation of human myelomonocytic cell line U937.

Today, many attempts have been collected in the field of tissue engineering for reconstitution of injured bone marrow capacity by transplantation of functional cell source. By having a three-dimensional condition, microcapsules are appropriate candidates for cells transplantation to target sites. Here, we examined the effect of alginate-gelatin microcapsules on functional maturation of human myelomonocytic cell line U937 after 7 days in vitro. U937 cells were encapsulated by the mixture of alginate-gelatin and cultured for 7 days. Trypan blue staining was used to show cell survival rate. Morphological changes were determined by haematoxylin and eosin staining. The expression of monocyte (CD14) and leukocyte (CD33) factors were measured by flow cytometry. The functional maturation of encapsulated cells was shown by immunocytochemistry targeting myeloperoxidase (MPO) activity and level of CD68. Transcription level of adhesion molecules CD68L, CD18, CD11b, and CD49d/VLA was detected by real-time polymerase chain reaction. In vivo constitutive capacity of encapsulated U937 was investigated in rabbits via administration to bone marrow. We showed enhanced U937 viability and monocyte and band cell-like appearance 7 days after encapsulation. These changes coincided with increasing CD33 and CD14 levels and a decrease of CD15, confirming cell maturation (p < 0.05). High level of MPO and CD68-positive cells showed the functional maturation of U937 cells into neutrophils and macrophages. Compared with that of nonencapsulated cells, the level of adhesion factor was up-regulated. We found labelled cells in the peripheral blood after cell transplantation to bone marrow. These data suggest that alginate-gelatin encapsulation of U937 cells promotes functional leukopoiesis and monocytopoiesis.