SERUM 24,25-DIHYDROXYVITAMIN D3 IS AN ACCURATE INDICATOR OF FOLLICULAR VITAMIN D STATUS AND OOCYTE AGING IN IVF PATIENTS

Abstract

There is increasing evidence supporting the role of vitamin D in telomere biology and thus the potential need for normal vitamin D status to develop a competent oocyte. The unreliable results obtained with immunoassay-based methods have supported the use of liquid chromatography-tandem mass spectrometry (LC-MS/MS) and other vitamin D metabolites measurement. We previously reported that 24,25-dihydroxyvitamin D3 (24,25(OH)2D3), the main catabolite of 25-hydroxyvitamin D3 (25OHD3), is a more accurate and stable indicator of systemic and ovarian vitamin D status in young egg donors. Consequently, we aimed to explore if 24,25(OH)2D3 is a suitable indicator of systemic and follicular vitamin D status in a population of women undergoing IVF PGT-A cycles. Then, we aimed to evaluate the correlation between telomere length in cumulus cells (CCTL) and vitamin D metabolites concentrations in serum (S) and follicular fluid (FF) in this IVF patient population. 27 women between 37 and 40-years old undergoing IVF PGT-A cycle were included in this prospective proof of concept study. On the pick-up day, S and pooled FF from mature follicles were evaluated for 24,25(OH)2D3 and 25OHD3 levels via LC-MS/MS. Cumulus cells (CC) were obtained after oocyte stripping for CCTL measurement. Genomic DNA from CC was isolated and average delta cycle threshold (ΔCt) was determined using a SYBR green quantitative real-time PCR protocol with Taqman probes for relative telomere length (TL) normalized by multicopy Alu. Difference in means between S and FF vitamin D metabolites and correlation between 24,25(OH)2D3 and ΔCtCC, were evaluated by t-test and Pearson correlation test. 25OHD3 and 24,25(OH)2D3 mean concentrations were, respectively, 43.15 ± 10.28 ng/ml and 14.30 ± 4.95 in S, and 33.67 ± 6.87 ng/ml and 11.67 ± 3.53 ng/ml in FF. Both metabolites highly correlated between them in S (r=0.78, p=1.5e-06) and in FF (r=0.73, p=1.8e-05). When analyzing each metabolite separately, 24,25(OH)2D3 showed the strongest association between S and FF levels (r=0.96, p=1.9e-15) followed by 25OHD3 (r=0.92, p=2.2e-11). ΔCtCC mean value was 7.54 ± 1.04, and we found a significant negative correlation with 24,25(OH)2D3 levels in S (r=-0.51, p=0.0068) and FF (r=-0.42, p=0.0307). Since ΔCt values are inversely proportional to the amount of nucleic acids amplified, a higher ΔCt value indicates a shorter telomere. Our results show serum 24,25(OH)2D3 is a better indicator of ovarian vitamin D status than serum 25OHD3. Further, the observed significant inverse correlation between this metabolite and CCTL, indicates that, the higher the vitamin D concentrations in S and FF, the longer the telomeres in the cumulus cells in this homogenous population of women undergoing IVF PGT-A cycle.