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Cellular and Molecular Bioengineering

, Volume 12, Issue 6, pp 599–613 | Cite as

Human Amniotic Membrane as a Matrix for Endothelial Differentiation of VEGF-Treated Dental Stem Cells

  • Siti Nurnasihah Md Hashim
  • Muhammad Fuad Hilmi Yusof
  • Wafa’ Zahari
  • Hamshawagini Chandra
  • Khairul Bariah Ahmad Amin Noordin
  • Thirumulu Ponnuraj Kannan
  • Suzina Sheikh Abdul Hamid
  • Khairani Idah Mokhtar
  • Ahmad AzlinaEmail author
Article

Abstract

Introduction

Endothelial cells cover the surface of the capillary wall and literature review has cemented its angiogenic roles in wound healing and tissue regeneration. However, the angiogenic in vitro models available are inadequate to understand the endothelial differentiation process.

Methods

A construct was made using human amniotic membrane (HAM) as a matrix to assist the dental stem cells to differentiate into endothelial-like cells. This study aimed to assess the biological interaction between stem cells from human exfoliated deciduous teeth (SHED) and the stromal side (SS) of the glycerol-preserved HAM in angiogenic-induced environment media using VEGF. The changes were evaluated through cell morphology, migration, as well as gene expression level.

Results

There were morphological changes observed in SHED in angiogenic-induced media. SHED appeared to be differentiated from fibroblast-like cells to a new structure, mimicking endothelial-like structure through microscopy analysis. Besides, the cross-section of the construct revealed that the cells seeded on the matrix were able to maintain its monolayer structure at day 1, 7 and 14 but infiltrated into the HAM at day 21, suggesting cell migration. The cells were also able to maintain its stemness (Nestin, Nanog and CD29) and at the same time express the angiogenic markers (IL-8, VEGF and MMP-2).

Conclusion

HAM promotes SHED proliferation, migration and has the potential as a differentiating matrix for endothelial-like cells.

Keywords

Angiogenic Endothelial differentiation Extracellular matrix Regenerative medicine Stem cells from human exfoliated deciduous teeth Vascular endothelial growth factor 

Abbreviations

APC

Allophycocyanin

B-actin

Beta-actin

BS

Basement side

CD

Cluster of differentiation

COX-2

Cyclooxygenase-2

ECM

Extracellular matrix

ECs

Epithelial cells

FITC

Fluorescein isothiocyanate

H&E

Haematoxylin and eosin

HAM

Human amniotic membrane

HMDS

Hexamethyldisilazane

HUVEC

Human umbilical vein endothelial cells

IL-8

Interleukin-8

MMP-2

Matrix metalloproteinase-2

NaClO

Sodium hypochlorite

PBS

Phosphate buffer saline

PE

Phycoerythrin

PerCP

Peridium‐chlorophyll protein complex

PVDF

Polyvinylidene fluoride

RT

Room temperature

RT-PCR

Reverse-transcriptase polymerase chain reaction

S

SHED only

SA

SHED cultured on HAM

SAV

SHED cultured on HAM treated with VEGF

SDS

Sodium dodecyl sulfate

SEM

Scanning electron microscope

SHED

Stem cells from human exfoliated deciduous teeth

SS

Stromal side

SV

SHED treated with VEGF

T-PBS

PBS containing 0.1% Tween-20

VEGF

Vascular endothelial growth factor

Notes

Acknowledgments

The authors would like to acknowledge the help rendered by the staff of Craniofacial Sciences Laboratory, School of Dental Sciences, Universiti Sains Malaysia. Ms. Siti Nurnasihah Md Hashim acknowledges the MyMaster scholarship from Malaysia Higher Education Ministry for her post-graduate programme.

Funding

The work was supported by Universiti Sains Malaysia Research University Grant (1001/PPSG/813075).

Author Contributions

SNMH performed the research and analysed the data. SNMH and AA wrote the manuscript. MFHY, WZ and HC involved in the scaffold material preparation and contributed to the technical part. The final draft of the manuscript was corrected by MFHY, TPK and AA. KBAAN, TPK, SSAH, KIM and AA designed the research study. All authors read and approved the final manuscript.

Conflict of Interest

The authors declare that they have no conflict of interest.

Ethical Statement

This study was approved by Human Research Ethics Committee of USM (USM/JEPeM/14110477), for the usage of HAM from the Tissue Bank Unit, School of Medical Sciences, Universiti Sains Malaysia. No animal studies were carried out by the authors for this article.

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Copyright information

© Biomedical Engineering Society 2019

Authors and Affiliations

  • Siti Nurnasihah Md Hashim
    • 1
  • Muhammad Fuad Hilmi Yusof
    • 1
  • Wafa’ Zahari
    • 1
  • Hamshawagini Chandra
    • 1
  • Khairul Bariah Ahmad Amin Noordin
    • 1
  • Thirumulu Ponnuraj Kannan
    • 1
    • 2
  • Suzina Sheikh Abdul Hamid
    • 3
  • Khairani Idah Mokhtar
    • 4
  • Ahmad Azlina
    • 1
    • 2
    Email author
  1. 1.School of Dental SciencesUniversiti Sains MalaysiaKubang KerianMalaysia
  2. 2.Human Genome Centre, School of Medical SciencesUniversiti Sains MalaysiaKubang KerianMalaysia
  3. 3.Tissue Bank, School of Medical SciencesUniversiti Sains MalaysiaKubang KerianMalaysia
  4. 4.Kulliyyah of DentistryInternational Islamic University MalaysiaKuantanMalaysia

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