# Effects of Perivitelline Fluid Obtained from Horseshoe Crab on The Proliferation and Genotoxicity of Dental Pulp Stem Cells

## TL;DR

This study investigates whether perivitelline fluid from horseshoe crabs affects dental pulp stem cells' growth and genetic safety.

## Contribution

The study provides new evidence on the non-genotoxic and limited proliferative effects of horseshoe crab perivitelline fluid on stem cells.

## Key findings

- PVF from horseshoe crabs showed no significant proliferative activity on dental pulp stem cells.
- PVF was found to be non-genotoxic based on chromosome aberration and Ames tests.
- Cell viability and proliferation levels were comparable to the control group.

## Abstract

Perivitelline fluid (PVF) of the horseshoe crab embryo has been reported to
possess an important role during embryogenesis by promoting cell proliferation. This
study aims to evaluate the effect of PVF on the proliferation, chromosome aberration (CA)
and mutagenicity of the dental pulp stem cells (DPSCs).

This is an in vitro experimental study. PVF samples were
collected from horseshoe crabs from beaches in Malaysia and the crude extract was
prepared. DPSCs were treated with different concentrations of PVF crude extract in
an 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay (cytotoxicity test). We choose two inhibitory concentrations (IC50 and IC25) and two PVF
concentrations which produced more cell viability compared to a negative control
(100%) for further tests. Quantitative analysis of the proliferation activity of PVF was
studied using the AlamarBlue®assay for 10 days. Population doubling times (PDTs)
of the treatment groups were calculated from this assay. Genotoxicity was evaluated
based on the CA and Ames tests. Statistical analysis was carried out using independent t test to calculate significant differences in the PDT and mitotic indices in the CA
test between the treatment and negative control groups. Significant differences in the
data were P<0.05.

A total of four PVF concentrations retrieved from the MTT assay were
26.887 mg/ml (IC50), 14.093 mg/ml (IC25), 0.278 mg/ml (102% cell viability) and 0.019
mg/ml (102.5% cell viability). According to the AlamarBlue®assay, these PVF groups
produced comparable proliferation activities compared to the negative (untreated)
control. PDTs between PVF groups and the negative control were insignificantly different (P>0.05). No significant aberrations in chromosomes were observed in the
PVF groups and the Ames test on the PVF showed the absence of significant positive
results.

PVF from horseshoe crabs produced insignificant proliferative activity on
treated DPSCs. The PVF was non-genotoxic based on the CA and Ames tests.