Changes in Migratory Speed Rate of Human Dental Pulp Stromal Cells Cultured in Advanced Platelet-Rich Fibrin.

OBJECTIVE: Migratory speed rate evaluation of human dental pulp stromal cells (hDP-SCs) is one of the important steps in dental pulp regeneration. Therefore, the aim of the study is to analyze various concentrations of advanced platelet-rich fibrin (A-PRF) culture media toward hDP-SCs' migratory speed rate evaluations. MATERIALS AND METHODS: The hDP-SCs were divided into four groups: control: hDP-SCs in Dulbecco's modified Eagle medium + 10% fetal bovine serum group; hDP-SCs in 1% A-PRF group; hDP-SCs in 5% A-PRF group; and hDP-SCs in 10% A-PRF group, which were planted in 24-well (5 × 104 cell/well). The migratory speed rate of all groups was measured by using cell migration assay (scratch wound assay) after 24 hours. Cell characteristics were evaluated under microscope (Inverted microscope, Zeiss, Observer Z1, UK) that can be read through image-J interpretation. This image J represented the measurement of migratory speed rate (nm/h) data. Statistical analysis was conducted using one-way analysis of variance and post hoc Tamhane's test (p < 0.05) (IBM SPSS Statistics Software, version 22.0). RESULTS: There was a statistically significant difference in the migratory speed rates of hDP-SCs among various concentration groups of A-PRF (1, 5, and 10%) compared with the control group. CONCLUSION: The increase in the migratory speed rate of hDP-SCs was highest in 10% A-PRF group.

Proteomic analysis of hypoxia and non-hypoxia secretome mesenchymal stem-like cells from human breastmilk.

INTRODUCTION: Breastmilk contains proteins and cells which have stem cell properties. The human breastmilk stem cell mimick mesenchymal stem cells and expresses pluripotency genes. The protein level of breastmilk is high in colostrum and gradually subsides in the first year of lactation. The mesenchymal stem cells from breastmilk can be an alternative source of stem cells that can potentially affect cardiovascular therapy. This study aimed to identify the proteomic analysis of secretome mesenchymal stem-like cells under hypoxia compared to non-hypoxia from human breastmilk stem cells. MATERIAL AND METHODS: The human breastmilk was collected from six healthy breastfeeding women and transported to the laboratory under aseptic conditions. The breastmilk cells were isolated then cultured. After 72 h, the human breastmilk stem cells reached confluence then cleaned up and isolated in serum-free media (spheroid) to allow serial passaging every 48 h. The acquisition stem cell was made with flow cytometry. The cells were divided into hBSC secretomes under hypoxia (A) and non-hypoxia (B) and analyzed for LC-MS to identify the peptide structure. RESULTS: The human breastmilk cells contained several mesenchymal stem-like cells in density 2.4 × 106 cell/mL for hypoxia and 2 × 106 cell/mL for non-hypoxia conditions. The human breastmilk stem cell surface markers derived from the third cell passage process were 93.77% for CD44, 98.69% for CD73, 88.45% for CD90, and 96.30% for CD105. The protein level of secretome mesenchymal stem -like cells under hypoxia was measured at 5.56 µg/mL and 4.28 µg/mL for non-hypoxia. The liquid chromatography-mass spectrometry analysis identified 130 and 59 peptides from hypoxia and non-hypoxia of the human breastmilk stem cell secretome sequentially. Some important proteomics structures were found in the hypoxic human breastmilk stem cell secretome, such as transforming growth factor-ß, VE-cadherin, and caspase. CONCLUSION: The human breastmilk cells contain mesenchymal stem-like cells and a high concentration of CD44, CD73, CD90, and CD105 as surface markers at third passage culture. The hypoxic hBSC secretome produces a higher protein level compare to non-hypoxia. The transforming growth factor -ß was found in the hypoxic hBSC secretome as a modulator of VEGF-mediated angiogenesis.

Comparison of Advanced Platelet Rich Fibrin (A-PRF) and Culture Media Conditioned Warton's Jelly (CMCWJ) on Fibroblast Cells Proliferation

Objective: To compare the potency of fibroblast cells proliferation in 12.5% and 25% Culture Media Conditioned Warton's Jelly (CMCWJ) and Advanced Platelet Rich Fibrin (A-PRF) cultured medium. Material and Methods: Fibroblast cells were divided into five groups: Group I (Control Group): serum-starved fibroblast without any treatment as a negative control; Group II: fibrolast that supplemented in 12.5% CMCWJ medium; Group III: fibrolast that supplemented 12.5% A-PRF medium; Group IV: fibrolast that supplemented 25% CMCWJ medium, and Group V: fibrolast that supplemented 25% A-PRF medium. The fibroblasts proliferation was counted by an automated cell counter. Statistical analysis was performed using One-way ANOVA and Post hoc Tamhane test was conducted to analyze the potential fibroblast proliferation differences in different concentration of CMCWJ and A-PRF group. Results: There were no significant differences in the fibroblast cell proliferation between GI and GIV, GII and GIV, GII and GIII, GII and GV, also GIV and GV. There were significant differences between GI and GII, GI and GIII, GI and GV, also GIII and GIV. Conclusion: The 12.5% CMCWJ group, 12.5% A-PRF group and 25% A-PRF group has excellent potential ability of fibroblast cells proliferation, meanwhile 25% CMCWJ group has the lowest mean potency of fibroblast cells proliferation compared to other groups. The 12.5% A-PRF Group has the highest mean of fibroblast cell proliferation amongst other groups.