Article Figures One through Five; PNG Files

Figures detailing the relationship between FRAC and NPB Polarization.

Fig. 1 Schematic of zygote centriole dynamics: (a) The spermatozoon neck contains two centrioles: the canonical proximal and the atypical distal centrioles. (b) The fertilized egg inherits a paternal pronucleus (blue with dashed line) and two centrioles (green dots). The centrioles form a centrosome and send out a microtubule aster (green lines) to pull the maternal pronucleus (blue) towards the paternal pronucleus; (c) The two pronuclei now sit in between the inherited centrosome and the newly formed centrosome with Nucleolus Precursor Bodies (orange) appearing in the two pronuclei; (d) The two centrosomes interact with the pronuclei’s nuclear pores and polarize the DNA and Nucleolus Precursor Bodies to prepare for the first cleavage; (e) The two centrosomes associate with the dual spindles poles, helping to organize and ensure correct cell cleavage [32]

Fig. 2 The studied population had seven couples with negative FRAC values and eight with positive FRAC values. Graph depicting the average FRAC ratio from each male (± 95% confidence intervals) for the nine parameters (for example, acetylated tubulin PC is a parameter). Each parameter shows male patients 1 to 15 from left to right. Black bold bars at the center of each group of confidence intervals represent the reference average plus/minus two standard deviations. Arrows point at 95% confidence intervals up to one standard deviation (blue arrows) or more than two standard deviations (orange arrows) outside the reference range

Fig. 3 Three scenarios of NPB polarization in the pronuclei of zygotes. (a-c) Represented model (top row) and annotated image (bottom row). Each pronucleus was divided into two halves, separated into either an “A” zone, the half that was furthest away from the junction, or a “J” zone, the half that was towards the junction; (a) A scenario of full NPB polarization in the two non-overlapping pronuclei. There were six NPBs (orange) in total, all in either J1 or J2 zones; (b) A scenario of full NPB polarization in the two overlapping pronuclei. There were nine NPBs in total, with five being in either J1 or J2 zones and four NPBs (green) being in the “O” zone, the zone created by the overlap of the two pronuclei; (c) A scenario showing partial NPB polarization in the two pronuclei with overlap. There were nine NPBs in total, with three NPBS being in the “J1” or “J2” zones, two NPBs in the “O” zone, and four NPBs (blue) being in the “A2” zone

Fig. 4 Half of the abnormal FRAC couples had high rates of reduced polarization pattern. (a) Individual breakdown for polarization patterns for each couple. Couples 1, 2, 4, 7, 8, 9, and 11 had normal FRAC scores. Most of their embryos are in either the full (green) or eventual (orange) polarization pattern. Couples 3, 5, 6, 10, 12, 13, 14, and 15 had abnormal FRAC values. Couples 3, 5, 6, and 10 had similar pattern distributions as those in the normal FRAC category. In contrast, couples 12, 13, 14, and 15 had a majority (couples 12 and 14) or all (couples 13 and 15) of their embryos classified as reduced (red) polarization. A dotted red box is placed around the four abnormal FRAC couples with the lowest patterned NPB polarization rates (or the highest rates of the reduced polarization pattern); (b) Most outlier values are concentrated in the acetylated tubulin biomarker. PC: proximal centriole, DC: distal centriole, Ax: axoneme

Fig. 5 Normal FRAC embryos had fewer reduced polarization pattern rates when compared to abnormal FRAC embryos. (a) Patterned NPB polarization of couples with normal FRAC and abnormal FRAC. Each column represents an average ± 1 SD. Each dot represents a couple (seven per column in normal FRAC and eight per column in abnormal FRAC). Each couple is color-coded to match those in Figure 2. A dotted red box is placed around the four abnormal FRAC couples with the lowest patterned polarization rate (or the highest rates of the reduced polarization pattern). (b) Patterned NPB polarization of embryos belonging to normal FRAC couples (n=62 embryos) and abnormal FRAC couples (n=78 embryos). Each column represents the overall percentage observed in our study. A Chi-Square Test of Independence was used for embryo population analysis. FP: Full Polarization, EP: Eventual Polarization, RP: Reduced Polarization