CHARACTERIZATION OF PLACENTAL AUTOPHAGY AND BIOENERGETICS, AND MATERNAL MICRORNAS PROFILING IN OBESE PREGNANCIES AND GESTATIONAL DIABETES MELLITUS

BACKGROUND Maternal obesity (MO) is expanding worldwide, contributing to Gestational Diabetes Mellitus (GDM) prevalence. The altered placental function and intrauterine environment of MO and GDM are associated with adverse maternal and fetal outcomes, with short- and long-term complications. Growing evidence suggests that both MO and GDM are characterized by placental autophagy disturbances, mitochondrial (mt) dysfunctions and epigenetic alterations, which could contribute to the pathogenesis of pregnancy-related metabolic diseases. For these reasons the aim of this work was to characterize placental autophagy and mitochondrial bioenergetics, and maternal plasmatic microRNAs profile in the context of MO and GDM. METHODS 60 women with spontaneous singleton pregnancies delivering by elective Caesarean section were enrolled: 25 normal-weight (NW), 22 obese without comorbidities (OB/GDM(-)), 13 obese with GDM (OB/GDM(+)). The expression of antioxidant metabolism markers and autophagy-related genes expression was evaluated in placental villous tissue by Real-time PCR with SYBR green chemistry. In placenta, we also quantified ATP by CellTiter-Glo Luminescent Cell Viability Assay adapted to placental villous tissue, and mt Complexes I-V protein content by Western Blot experiments. Moreover, in a subgroup of 7 NW, 6 OB/GDM(-) and 6 OB/GDM(+), microRNA (miRNA) profiling in maternal plasma at delivery was performed with miRCURY LNA Serum/Plasma miRNA Focus PCR Panel (Qiagen). Statistical analysis and bioinformatics predictive tools: SPSS v.27 (IBM); GeneGlobe (Qiagen); miTALOS v.2; miRPath v.3. RESULTS AND CONCLUSIONS Placental antioxidant genes expression (CAT, GPX1, GSS, GSR) tended to decrease, the pro-autophagic ULK1 increased and the chaperone-mediated autophagy (CMA) regulator PHLPP1 decreased in OB/GDM(-) vs NW. On the other hand, PHLPP1 expression increased in OB/GDM(+) vs OB/GDM(-). These preliminary findings showed placental alterations of autophagy in MO and GDM, paving the way for further analyses aimed at elucidating the role of placental autophagy in metabolic disorders during pregnancy. Moreover, placentas of OB/GDM(-) and OB/GDM(+) women had reduced ATP level vs NW. All OB showed decreased Complex I vs NW. Furthermore, lower expression of Complexes III and V was found in OB/GDM(+) group (vs NW; vs NW and OB/GDM(-), respectively), along with a reduced fetal/placental weight ratio (i.e. placental efficiency) vs NW. These results might suggest impaired placental mt function in obese subjects, especially with GDM. Sexual dimorphism in terms of the above-mentioned placental molecular markers was also investigated, with intriguing results suggesting that females and males might respond differently to insults and/or have differential, sex-dependent compensatory mechanisms in order to deal with the perturbed environment. MiRNA profiling in maternal plasma highlighted patterns of significantly differentially expressed miRNAs between the three groups. In particular, OB/GDM(-) vs NW differed in the expression of 4 miRNAs, OB/GDM(+) vs NW of 1, and OB/GDM(+) vs OB/GDM(-) of 14. Pathway enrichment analysis by miRPath and miTALOS revealed many pathways associated with the set of miRNAs of each comparison, among which the most interesting and relevant in our context were related to nutrients and hormones metabolism, inflammation and oxidative stress. Indeed, miRNAs could be promising biomarkers of metabolic alterations in MO and GDM. Nevertheless, future investigations are needed to further deepen the pregnancy epigenetic landscape.