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Potentiated Osteoinductivity via Cotransfection with BMP-2 and VEGF Genes in Microencapsulated C2C12 Cells.

Shen Y, Qiao H, Fan Q, Zhang S, Tang T - Biomed Res Int (2015)

Bottom Line: Unfortunately, the osteoinductivity of microcapsules has been restricted by many factors, among which the deficiency of functional proteins is a significant priority.Various tissue-derived mesenchymal stem cells and cell lines were compared for BMP-2 and VEGF cotransfection.These results demonstrated that the cotransfection of BMP-2 and VEGF into microencapsulated C2C12 cells is of potent utility for the potentiation of bone regeneration, which would provide a promising clinical strategy for cellular therapy in bone defects.

View Article: PubMed Central - PubMed

Affiliation: Shanghai Key Laboratory of Orthopaedic Implants, Department of Orthopedic Surgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, 639 Zhizaoju Road, Shanghai 20011, China.

ABSTRACT
Microcapsules with entrapped cells hold great promise for repairing bone defects. Unfortunately, the osteoinductivity of microcapsules has been restricted by many factors, among which the deficiency of functional proteins is a significant priority. We potentiated the osteoinductivity of microencapsulated cells via cotransfection with BMP-2 and VEGF genes. Various tissue-derived mesenchymal stem cells and cell lines were compared for BMP-2 and VEGF cotransfection. Ethidium bromide (EB)/Calcein AM staining revealed that all of the cell categories could survive for 4 weeks after microencapsulation. An ELISA assay indicated that all microencapsulated BMP-2 or VEGF transfected cells could secrete gene products constitutively for 1 month. Particularly, the recombinant microencapsulated C2C12 cells released the most desirable level of BMP-2 and VEGF. Further experiments demonstrated that microencapsulated BMP-2 and VEGF cotransfected C2C12 cells generated both BMP-2 and VEGF for 4 weeks. Additionally, the cotransfection of BMP-2 and VEGF in microencapsulated C2C12 cells showed a stronger osteogenic induction against BMSCs than individual BMP-2-transfected microencapsulated C2C12 cells. These results demonstrated that the cotransfection of BMP-2 and VEGF into microencapsulated C2C12 cells is of potent utility for the potentiation of bone regeneration, which would provide a promising clinical strategy for cellular therapy in bone defects.

No MeSH data available.


The morphology of different tissue-derived mesenchymal stem cells and cell lines. (a) The morphology of different tissue-derived mesenchymal stem cells with the appearance of a colony-forming unit. (A) indicates the bone marrow-derived mesenchymal stem cells (BMSCs), (B) indicates the adipose-derived stem cells (ADSCs), and (C) indicates the synovium-derived mesenchymal stem cells (SMSCs). (b) The morphology of different cells lines. (A) represents the C3H10T1/2 cells, (B) indicates the C2C12 cells, and (C) indicates the NIH/3T3 cells. Scale: 20 μm.
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fig1: The morphology of different tissue-derived mesenchymal stem cells and cell lines. (a) The morphology of different tissue-derived mesenchymal stem cells with the appearance of a colony-forming unit. (A) indicates the bone marrow-derived mesenchymal stem cells (BMSCs), (B) indicates the adipose-derived stem cells (ADSCs), and (C) indicates the synovium-derived mesenchymal stem cells (SMSCs). (b) The morphology of different cells lines. (A) represents the C3H10T1/2 cells, (B) indicates the C2C12 cells, and (C) indicates the NIH/3T3 cells. Scale: 20 μm.

Mentions: Inverted optical microscopic observations indicated that all of the nontransfected cell types demonstrated a specific morphology (Figure 1). BMSCs exhibited a rod-like or spindle-shaped formation. After 7 days of incubation, the initial cell colony increased to merge with the other clones, bringing along the adherent spindle- and rod-shaped cells (Figure 1(a)-(A)). Regarding the ADSCs, after the first inoculation of 5–7 hr, some cells began to adhere and exhibited elongated pseudopodia, thereby contributing to a spindle-like appearance. 24 hr later, the cells completely adhered to the dishes, exhibiting a fibroblast-like cell growth with a proliferative peak at 3–5 days (Figure 1(a)-(B)). The presence of SMSCs indicated apparent adherent cells with elongated or polygonal shapes. After 72 hr of cultivation, a multitude of spindle-shaped cells with small round cells scattered inside were present. Cell colony formations were detectable at around day 5 (Figure 1(a)-(C)). Optical microscopic observations of C3H10T1/2, C2C12, and NIH/3T3 cell lines (Figure 1(b)) showed an elongated cell appearance with a rapidly proliferative bioactivity.


Potentiated Osteoinductivity via Cotransfection with BMP-2 and VEGF Genes in Microencapsulated C2C12 Cells.

Shen Y, Qiao H, Fan Q, Zhang S, Tang T - Biomed Res Int (2015)

The morphology of different tissue-derived mesenchymal stem cells and cell lines. (a) The morphology of different tissue-derived mesenchymal stem cells with the appearance of a colony-forming unit. (A) indicates the bone marrow-derived mesenchymal stem cells (BMSCs), (B) indicates the adipose-derived stem cells (ADSCs), and (C) indicates the synovium-derived mesenchymal stem cells (SMSCs). (b) The morphology of different cells lines. (A) represents the C3H10T1/2 cells, (B) indicates the C2C12 cells, and (C) indicates the NIH/3T3 cells. Scale: 20 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4588358&req=5

fig1: The morphology of different tissue-derived mesenchymal stem cells and cell lines. (a) The morphology of different tissue-derived mesenchymal stem cells with the appearance of a colony-forming unit. (A) indicates the bone marrow-derived mesenchymal stem cells (BMSCs), (B) indicates the adipose-derived stem cells (ADSCs), and (C) indicates the synovium-derived mesenchymal stem cells (SMSCs). (b) The morphology of different cells lines. (A) represents the C3H10T1/2 cells, (B) indicates the C2C12 cells, and (C) indicates the NIH/3T3 cells. Scale: 20 μm.
Mentions: Inverted optical microscopic observations indicated that all of the nontransfected cell types demonstrated a specific morphology (Figure 1). BMSCs exhibited a rod-like or spindle-shaped formation. After 7 days of incubation, the initial cell colony increased to merge with the other clones, bringing along the adherent spindle- and rod-shaped cells (Figure 1(a)-(A)). Regarding the ADSCs, after the first inoculation of 5–7 hr, some cells began to adhere and exhibited elongated pseudopodia, thereby contributing to a spindle-like appearance. 24 hr later, the cells completely adhered to the dishes, exhibiting a fibroblast-like cell growth with a proliferative peak at 3–5 days (Figure 1(a)-(B)). The presence of SMSCs indicated apparent adherent cells with elongated or polygonal shapes. After 72 hr of cultivation, a multitude of spindle-shaped cells with small round cells scattered inside were present. Cell colony formations were detectable at around day 5 (Figure 1(a)-(C)). Optical microscopic observations of C3H10T1/2, C2C12, and NIH/3T3 cell lines (Figure 1(b)) showed an elongated cell appearance with a rapidly proliferative bioactivity.

Bottom Line: Unfortunately, the osteoinductivity of microcapsules has been restricted by many factors, among which the deficiency of functional proteins is a significant priority.Various tissue-derived mesenchymal stem cells and cell lines were compared for BMP-2 and VEGF cotransfection.These results demonstrated that the cotransfection of BMP-2 and VEGF into microencapsulated C2C12 cells is of potent utility for the potentiation of bone regeneration, which would provide a promising clinical strategy for cellular therapy in bone defects.

View Article: PubMed Central - PubMed

Affiliation: Shanghai Key Laboratory of Orthopaedic Implants, Department of Orthopedic Surgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, 639 Zhizaoju Road, Shanghai 20011, China.

ABSTRACT
Microcapsules with entrapped cells hold great promise for repairing bone defects. Unfortunately, the osteoinductivity of microcapsules has been restricted by many factors, among which the deficiency of functional proteins is a significant priority. We potentiated the osteoinductivity of microencapsulated cells via cotransfection with BMP-2 and VEGF genes. Various tissue-derived mesenchymal stem cells and cell lines were compared for BMP-2 and VEGF cotransfection. Ethidium bromide (EB)/Calcein AM staining revealed that all of the cell categories could survive for 4 weeks after microencapsulation. An ELISA assay indicated that all microencapsulated BMP-2 or VEGF transfected cells could secrete gene products constitutively for 1 month. Particularly, the recombinant microencapsulated C2C12 cells released the most desirable level of BMP-2 and VEGF. Further experiments demonstrated that microencapsulated BMP-2 and VEGF cotransfected C2C12 cells generated both BMP-2 and VEGF for 4 weeks. Additionally, the cotransfection of BMP-2 and VEGF in microencapsulated C2C12 cells showed a stronger osteogenic induction against BMSCs than individual BMP-2-transfected microencapsulated C2C12 cells. These results demonstrated that the cotransfection of BMP-2 and VEGF into microencapsulated C2C12 cells is of potent utility for the potentiation of bone regeneration, which would provide a promising clinical strategy for cellular therapy in bone defects.

No MeSH data available.