The functional barrier for exchange between the mother and fetus in the placenta is created by the fusion of cytotrophoblasts with one another to form a continuous, multinuclear syncytiotrophoblast, which is maintained by the incorporation of underlying proliferative cytotrophoblasts. Disruption to this process has been hypothesised to be involved in the aetiology of preeclampsia. Recently we investigated caspase-14 in the context of trophoblast differentiation as it is crucial to epidermal differentiation and keratin stabilisation, revealing disparate expression of caspase-14 in the differentiating BeWo cell line. Consequently, further examination as to its functional role in trophoblast differentiation was conducted using RNA Interference (RNAi), with the hypothesis that differentiation would be suppressed following caspase-14 silencing. 100nM siRNA were delivered into the BeWo cell line for 16 h before the addition of 20µM Forskolin. Cultures were incubated for a further 24, 48 or 72 h before the extraction of RNA and protein. Transcription of the trophoblast hormone β-hCG, the endothelial mediator of eNOS, and cytokeratin 18 were found to be increased after both 24 and 48 h of differentiation following silencing, implicating caspase-14 in the regulation of these pathways. As both β-hCG and eNOS are significantly increased with trophoblast differentiation, this indicates that caspase-14 functionally suppresses BeWo differentiation. Furthermore, the differential expression of cytokeratin 18 indicates a conserved role for caspase-14 in keratin homeostasis in barrier formation. In conclusion, the suppression of caspase-14 in the BeWo cell line resulted in augmented differentiation, a trait often observed in preeclampsia. Further investigation of caspase-14 activity would provide important insight into mechanisms of trophoblast differentiation, particularly in relation to preeclampsia.