Effect of pregnancy-associated plasma protein-A (PAPP-A) single-nucleotide polymorphisms on the level and activity of PAPP-A and the hormone profile in.

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Presentation transcript:

Effect of pregnancy-associated plasma protein-A (PAPP-A) single-nucleotide polymorphisms on the level and activity of PAPP-A and the hormone profile in fluid from normal human small antral follicles Jane Alrø Bøtkjær, M.Sc., Tanni Borgbo, Ph.D., Søren Kløverpris, Ph.D., Pernille Rimmer Noer, M.Sc., Claus Oxvig, Ph.D., Claus Yding Andersen, D.M.Sc. Fertility and Sterility Volume 106, Issue 7, Pages 1778-1786.e8 (December 2016) DOI: 10.1016/j.fertnstert.2016.09.008 Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

Figure 1 Melting curves from competitive amplification of differentially melting amplicons (CADMA). Melting curves achieved from CADMA of 327 (A–D) and 1224 (E–H) single-nucleotide polymorphisms (SNPs) in duplicates from DNA extracted from human ovary tissue derived from women having one ovary removed for fertility preservation by cryopreservation before chemotherapy. Sanger sequencing was performed on DNA from five women to test the reliability of the melting curves derived from CADMA. These are shown in A and E and were used as the controls, where blue melting curves show the T/T or A/A genotype, purple shows the T/C or A/C genotype, and red shows the C/C genotype. The y-axis shows the fluorescence signal, and the x-axis shows the melting temperature (C°). Fertility and Sterility 2016 106, 1778-1786.e8DOI: (10.1016/j.fertnstert.2016.09.008) Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

Figure 2 Pregnancy-associated plasma protein-A (PAPP-A) activity toward radiolabeled IGFBP-4 versus molar ratio of PAPP-A/STC2. A strong positive correlation is observed between proteolytic activity toward radiolabeled IGFBP-4 in follicular fluid and the molar ratio of PAPP-A/STC2. Fertility and Sterility 2016 106, 1778-1786.e8DOI: (10.1016/j.fertnstert.2016.09.008) Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

Supplemental Figure 1 Intrafollicular pregnancy-associated plasma protein-A (PAPP-A) levels grouped according to the PAPP-A SNPs. Statistical significance: P=.0058 (overall analysis of variance). Multiple comparisons of means: *A/C versus C/C, P=.0014 and **A/A versus C/C P=.0040. Fertility and Sterility 2016 106, 1778-1786.e8DOI: (10.1016/j.fertnstert.2016.09.008) Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

Supplemental Figure 2 Alternative cleavage pattern of IGFBP-4. Proteolytic activity toward radiolabeled IGFBP-4 in follicular fluid (FF) in the presence or absence of 1 μM of an inhibitory monoclonal antibody specific for pregnancy-associated plasma protein-A (PAPP-A) (1/41) or a control IgG (Ctrl IgG). As controls, IGFBP-4 was incubated with (rPAPP-A) or without (Blank) 35 pM recombinant PAPP-A. Cleavage reactions were stopped after 8 hours. FF1 represents the FF sample from a woman in the study group showing the typical PAPP-A cleavage pattern of IGFBP-4 giving rise to two comigrating bands at approximately 15 kDa. FF(677) represents the FF sample from one of the two women who showed the alternative cleavage pattern of IGFBP-4 giving rise to three fragments and almost no visible intact radiolabeled IGFBP-4. Fertility and Sterility 2016 106, 1778-1786.e8DOI: (10.1016/j.fertnstert.2016.09.008) Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

Supplemental Figure 3 Intrafollicular hormone levels grouped according to the pregnancy-associated plasma protein-A (PAPP-A) 1224 single-nucleotide polymorphism (SNP). Intrafollicular levels of antimüllerian hormone (AMH), estradiol, progesterone, testosterone, and androstenedione according to PAPP-A 1224 and 327 SNP genotype. The data are the mean of the intrafollicular hormone levels, and the error bars are ± standard error of the mean values. Statistical significance: None. Fertility and Sterility 2016 106, 1778-1786.e8DOI: (10.1016/j.fertnstert.2016.09.008) Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions

Supplemental Figure 4 Intrafollicular hormone levels grouped according to the pregnancy-associated plasma protein-A (PAPP-A) 327 single-nucleotide polymorphism (SNP). Intrafollicular levels of antimüllerian hormone (AMH), estradiol, progesterone, testosterone, and androstenedione according to PAPP-A 1224 and 327 SNP genotype. The data are the mean of the intrafollicular hormone levels, and the error bars are ± standard error of the mean values. Statistical significance: None. Fertility and Sterility 2016 106, 1778-1786.e8DOI: (10.1016/j.fertnstert.2016.09.008) Copyright © 2016 American Society for Reproductive Medicine Terms and Conditions