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3D Decellularized Native Extracellular Matrix Scaffold for In Vitro Culture Expansion of Human Wharton’s Jelly-Derived Mesenchymal Stem Cells (hWJ MSCs)

  • Balasubramanian Sundaram
  • Anne George Cherian
  • Sanjay Kumar
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1577)

Abstract

Mesenchymal stem cells (MSCs) are derived from Wharton’s jelly tissue of the human umbilical cord. Given appropriate culture conditions, these cells can self-renew and differentiate into multiple cell types across the lineages. Among the properties exhibited by these cells, immunomodulation through secretion of trophic factors has been widely exploited in a broad spectrum of preclinical/clinical regenerative applications. Moreover, the extracellular matrix is found to play a major role apart from niche cells in determining stem cell fate including that of MSCs. Therefore, the currently employed technique of two-dimensional culture expansion can alter the inherent properties of naïve MSCs originally residing within the three-dimensional space. This limitation can be overcome to some extent by using native extracellular matrix scaffold culture system which mimics the in situ microenvironment. In this chapter, we have elucidated the protocol for the preparation of a native extracellular matrix scaffold by decellularization of the MSC sheet and thereof culture expansion and characterization of human Wharton’s jelly-derived MSCs.

Keywords

Decellularization Extracellular matrix Mesenchymal stem cells Scaffold Wharton’s jelly Stem cell niche 

Abbreviations

αMEM

Alpha modification of minimum essential medium

APC

Allophycocyanin

CD

Cluster of differentiation

DAPI

4′,6-Diamidino-2-phenylindole

DMSO

Dimethyl sulfoxide

DNase

Deoxyribonuclease

DPX

Distyrene-plasticizer-xylene

ECM

Extracellular matrix

EDTA

Ethylenediaminetetraacetic acid

FACS

Fluorescence-activated cell sorting

FITC

Fluorescein isothiocyanate

hWJ MSCs

Human Wharton’s jelly-derived MSCs

IgG

Immunoglobulin G

ITS

Insulin-transferrin-selenium

MSCs

Mesenchymal stem cells

μl

Microliters

μM

Micromoles

ml

Milliliters

mM

Millimoles

PBS

Phosphate buffered saline

PBST

Phosphate buffered saline with Tween 20

PE

Phycoerythrin

Pen-Strep

Penicillin-Streptomycin

2D

Two dimensional

3D

Three dimensional

Notes

Acknowledgments

We would like to thank the University Grants Commission (UGC) for granting Junior Research Fellowship (JRF)/Senior Research Fellowship (SRF) to Balasubramanian Sundaram. We would like to thank the Department of Biotechnology (DBT), India, for granting Ramalingaswami Fellowship and research support grant to Sanjay Kumar. We would like to thank Merylin Nixon, Department of Community Health, Christian Medical College Vellore Hospital, Vellore, India, for assisting us in obtaining umbilical cord tissue samples. We would like to appreciate the technical assistance provided by core facilities of Centre for Stem Cell Research, A unit of inStem Bengaluru, Christian Medical College, Vellore, Bagayam, Tamil Nadu, 632002, India.

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Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Balasubramanian Sundaram
    • 1
  • Anne George Cherian
    • 2
    • 3
  • Sanjay Kumar
    • 1
  1. 1.Center for Stem Cell Research, A Unit of inStem Bengaluru, Christian Medical CollegeCMC Rehab Campus BagayamVelloreIndia
  2. 2.Department of Obstetrics and GynaecologyChristian Medical College Vellore HospitalVelloreIndia
  3. 3.Department of Community HealthChristian Medical College Vellore HospitalVelloreIndia

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