Cell-free mtDNA level and its biomarker potency for ART outcome are different in follicular fluid of PCOS and non-PCOS women

•Cf-mtDNA in follicular fluid of PCOS patients was significantly lower than non-PCOS women.•We found that cf-mtDNA in follicular fluid of women with different reproductive status has not an equal efficiency regarding biomarker of ART outcome.•Cf-mtDNA is not an appropriate biomarker for ART outcome in PCOS patients.•Our data suggests that elevation of cf-mtDNA in follicular fluid might result from impairment of mtDNA packaging by TFAM, and the consequence of this event will be triggered by induction of inflammatory cytokines. Lack of reliable biomarkers for estimating the outcome is one of the current gaps in ART. In this study, we assessed whether cell-free mitochondrial DNA within the follicular fluid (FF cf-mtDNA) of PCOS patients has biomarker applicability or not. Furthermore, probable involved mechanisms in the FF cf-mtDNA pathway were evaluated. The level of FF cf-mtDNA was compared between 50 PCOS patients and 50 women without any certain reproductive disorder, and analyzed for correlations with ART outcome. The associations between levels of FF cf-mtDNA and TFAM, POLG, and RNase H1 genes expression in mural granulosa cells (MGCs), as well as IL-6, and TNFα in follicular fluid (FF) were assessed. We identified that FF cf-mtDNA level was significantly lower in PCOS women and was accompanied by a reduction in the expression of mtDNA biogenesis genes in MGCs of the patients. Although a significant association between FF cf-mtDNA level and ART outcome was observed in the control group, no correlation could be proved in the PCOS group. Moreover, the expression level of TFAM was negatively associated, while amounts of IL-6 and TNFα were positively correlated with FF cf-mtDNA level in both groups. PCOS patients present a lower FF cf-mtDNA level in comparison with non-PCOS women. FF cf-mtDNA has biomarker applicability for ART outcome in women without any certain reproductive disorder, but not for those with PCOS. It seems that mtDNA packaging dysfunction results in elevated FF cf-mtDNA, and subsequent effects are triggered by increasing the inflammatory cytokines.