ABSTRACT
@#Proteases of nematodes play a crucial role in larval molting and, in addition to their active role in egg hatching, proteases are also considered a crucial factor in tissue invasion and connective tissue remodeling. In Toxocara canis, proteases play important roles throughout the complex life cycle. They can degrade components of a model of extracellular matrix, basement membranes and different physiological substrates. In the present study, measurements of the proteolytic activity of the perivitelline fluid (PF) surrounding Toxocara canis embryos at different stages of development, the hatching fluid (HF) surrounding the infective larvae, as well as the excretory secretory (ES) products of the larvae in the culture media were performed. Measurements were made using casein as substrate following the Sigma non-specific protease activity assay. The results showed that enzyme activity increased as the embryo matured. The infective larvae were found to continuously produce proteases in the surrounding HF and ES products after in vitro cultivation indicating that Toxocara canis proteases might be important for the worm in the egg and the host. Optimal enzymatic activity was found at pH 8. Incubation of the antiserum from infected mice with the HF and ES products decreased their proteolytic activities, suggesting that there may be a link between the proteases present in these fluids and the immune response.
ABSTRACT
Perivitelline fluid, extracted from the fertilized eggs of horseshoe crabs, has been reported to play a vital role in supporting embryogenesis as well as cell proliferation. The present study aims to evaluate the effect of PVF on the expression of COL1A1 in human dental pulp stem cells (DPSCs). The cells were grouped into two; untreated (control) and treated with a single dose of PVF (0.019 mg/ml). Gene expression was quantified for COL1A1 on day 1, 3 and 7 using reverse transcriptase PCR. The expression of COL1A1 on day 3 of treated group with PVF was the highest though there was a decline of COL1A1 expression on day 7. Mann Whitney test was utilized to determine the significance of COL1A1 expression between treated and untreated groups. Significant difference in the expression of COL1A1 was observed between the treated and untreated groups on day 3 though there was no significance in the expression on day 7. The present study indicates that PVF may have the potential to increase cell proliferation in human DPSCs.
Subject(s)
Dental Pulp , Horseshoe Crabs , Stem CellsABSTRACT
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 expression of cell cycle regulatory genes from human dental pulp stem cells (DPSCs) between different cell passages viz. 4, 5, 6. The cells were treated with a single dose of PVF (26.89 mg/ml) PVF. Gene expression was quantified for CDKNA2A, PTEN, MDM2 and TP53 genes using reverse transcriptase PCR. CDKN2A and MDM2 expression for treated and untreated DPSCs, expressed a similar pattern of expression. The higher expression of CDKN2A showed that the treatment increased cell proliferation and prevented cell senescence. DPSCs with PVF treatment showed increased expression of MDM2 at passage 4 and drastically declined expression at passage 5 and slightly increased at passage 6. TP53 expression of DPSCs treated group showed a higher expression compared to untreated group. On the other hand, the expression of PTEN in DPSCs treated group started to increase from passage 5 to 6. However, on the whole, the PTEN expression was higher than the untreated group in all the passages studied here. The results showed that PVF could enhance cell cycle regulatory gene expression in DPSCs as indicated by the higher expression of all the genes considered in this study at different cell passages in the treated group compared to the untreated group. Mann Whitney test was utilized to determine the significance of cell cycle regulatory genes expression between treated and untreated group. Significant difference in expression of genes between the treated and untreated groups were found at all passages except for CDKN2A gene whereby, its expression was not significantly different at passage 5 though it did express slightly higher in PVF treated DPSCs.