TITLE:
RNA Interference Mediated Silencing of Tissue Factor in Human Umbilical Vein Endothelial Cells
AUTHORS:
Xiaojuan Yin, Zhu Chen, Weiqin Zhou, Wen Tang, Zhichun Feng
KEYWORDS:
Neonate, Tissue Factor, Placental Abruption
JOURNAL NAME:
Open Journal of Obstetrics and Gynecology,
Vol.8 No.5,
May
10,
2018
ABSTRACT: Objective: To construct to silence tissue factor (TF) expression in Human Umbilical
Vein Endothelial Cells (HUVECs) for providing evidence of the gene therapy and
prevention of coagulation dysfunction in placental abruption (PA) neonates. Methods: Cultured HUVECs in vitro were divided
into control group and PA group, each including non-prevention, scramble siRNA
and TF-siRNA subgroups. pENTRTM/U6-shRNA/TF vector expressing TF were
constructed and transfected into HUVECs. The mRNA expression of TF was tested
with RT-PCR, and TF protein expression was detected with immunofluorescence
staining. Results: Monolayer HUVECs with short-rod and short spindle
shaped were adherent to the bottom, forming paving stone arrangement. The TF mRNA
expression was significantly different between control group and PA group (P 0.01) and among different
subgroups (P 0.01).
In non-prevention subgroups, significant difference was observed in TF mRNA
expression between control group and PA group (P 0.05). Significant
difference in TF mRNA expression was found in false-prevention subgroups (P 0.05). The TF mRNA
expression was markedly different among different subgroups in control (P 0.01), and the similar
result among different PA subgroups (P 0.01). In both control and
PA groups, the TF mRNA expression was the lowest after TF silencing.
Immunofluorescence staining showed high TF expression in HUVECs in
false-prevention subgroups. In scramble siRNA group, the TF protein expression
reduced as compared to non-prevention group and reduced dramatically after TF
silencing as compared to control. Conclusion: pENTRTM/U6-TF-shRNA
is able to significantly inhibit the TF mRNA and protein expression in HUVECs
from healthy neonates and PA neonates.