Author(s)

Saeed Azandeh, Mahmoud Orazizadeh, Mahmoud Hashemitabar, Ali Khodadadi, Ali Akbar Shayesteh, Darioush Bijan Nejad, Anneh Mohammad Gharravi, Elham Allahbakhshi

Cellular and Molecular Research Center (CMRC), Department of Anatomical science, Faculty of Medicine, Ahvaz Jundishapour University of Medical Sciences (AJUMS), Ahvaz, Iran.
Cellular and Molecular Research Center (CMRC), Department of Immunology, Ahvaz University of Medical Sciences, Ahvaz, Iran.
Division of Gastroenterology and Hepatology, Department of Internal Medicine, Imam Hospital, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.
School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran.

We report combination of explants and enzymatic protocol as mixed enzymatic-explant procedure to faster extraction of MSCs from WJ. Umbilical cords (UC) were collected from Imam Khomini Hospital. For explant outgrowth, 6 - 9 pieces of WJ were transferred onto tissue culture flask and waited for attachment. For mixed enzymatic-explant, 1 cm³ pieces WJ were placed in enzymatic cocktail comprising 4 mg/ml Collagenase Type I and 1 mg/ml Hyaluronidase and 0.1% trypsin-EDTA. Then isolated cells were analyzed for surface cell markers such as CD73, CD31. Isolated 1.0 × 10⁶ MSCs/ml were encapsulated in alginate hydrogel. Cells with MSCs phenotype were isolated from mixed enzymatic-explant and explant procedures within 24 - 48 hrs and 7 - 10 days, respectively. Both of procedures were shown to form clumps and colonies with dense centers. Phenotypic changes gradually appeared as round cell in UC pieces into homogeneous spindle-shaped and typical fibroblast-like shape cells. By using flow cytometery MSCs showed positive for CD73, and negative for CD31. the morphology of viable MSCs in the beads did not significantly show a different morphology pattern before and after the bead formation process. These findings are indicated that when mixed enzymatic-explant procedure is performed MSCs can be isolated faster and much higher from WJ. These finding is important in comparing with time consuming explants culture for isolation of MSCs.

Mesenchymal Stem Cells; Isolation; Flow Cytometery; Alginate

Azandeh, S. , Orazizadeh, M. , Hashemitabar, M. , Khodadadi, A. , Shayesteh, A. , Nejad, D. , Gharravi, A. and Allahbakhshi, E. (2012) Mixed enzymatic-explant protocol for isolation of mesenchymal stem cells from Wharton’s jelly and encapsulation in 3D culture system. Journal of Biomedical Science and Engineering, 5, 580-586. doi: 10.4236/jbise.2012.510071.