Lower right quadrant represents viable cells, lower left quadrant represents depolarized/live cells, upper right quadrant is depolarized/dead cells, and the upper right quadrant is dead cells
Lower right quadrant represents viable cells, lower left quadrant represents depolarized/live cells, upper right quadrant is depolarized/dead cells, and the upper right quadrant is dead cells. derived from hESCs. 200 M hydrogen peroxide (H2O2) increases ROS following a six-hour acute treatment. NIHMS1512545-supplement-Supp3.jpg (85K) GUID:?994D1D97-6594-4726-985E-1FEB72EFD9D2 Supplemental Fig. S4: PFOS, PFOA, and PFNA decrease ROS generation in spermatogenic cells derived from hESCs. Flow cytometry based analysis of DHE labeling reporting percent ROS? and percent ROS+ cells for the low to middle concentrations of PFOS, PFOA, and PFNA assessed. Blue indicates ROS?. Red indicates ROS+. NIHMS1512545-supplement-Supp4.jpg (353K) GUID:?B064BE29-89F3-489B-9477-6870F56E9FD0 Supplemental Fig. S5: PFOS, PFOA, and PFNA do not impact mitochondrial membrane potential in spermatogenic cells derived from hESCs. Flow cytometry analyses for indicating PR-171 (Carfilzomib) percent live cells, percent a depolarized/live cells, percent depolarized/dead cells, and percent dead cells for the low to middle concentrations of PFOS, PFOA, and PFNA assessed. Lower right quadrant represents viable cells, lower left quadrant represents depolarized/live cells, upper right quadrant is depolarized/dead cells, and the upper right quadrant is dead cells. NIHMS1512545-supplement-Supp5.jpg (682K) GUID:?CF2C626D-9A9E-4909-8298-4B0225A81EC1 Supplemental Fig. S6: PFOS, PFOA, and PFNA PR-171 (Carfilzomib) do not affect the cell cycle or haploid cell viability in spermatogenic cells derived from hESCs. Flow cytometry analyses of cell cycle profiles for spermatognic cells treated in the low to middle concentrations of PFOS, PFOA, and PFNA assessed. Green, blue, purple, and beige populations on flow cytometry correspond to haploid, G0/G1, S, and G2 phases, respectively. NIHMS1512545-supplement-Supp6.jpg (394K) GUID:?DDD0AACB-D937-40A1-B365-899EBE626C93 Supplemental Fig. S7: PFOS, PFOA, and PFNA impact PLZF area and intensity in spermatogonia derived under spermatogenic conditions. Representative 5X images obtained by the Cellomics ArrayScan VT1 of PLZF + (green) and DAPI (blue)-stained colonies treated with the low to middle concentrations of PFOS, PFOA, and PFNA assessed. All images are taken under the same imaging conditions and parameters. NIHMS1512545-supplement-Supp7.jpg (258K) GUID:?0B99E4C9-FF7D-4B3B-AE5A-8EDFFBDD30F4 Supplemental Fig. S8: PFOS, PFOA, and PFNA influence HILI area and intensity in primary spermatocytes derived under spermatogenic conditions. Representative 5X PR-171 (Carfilzomib) images obtained by the Cellomics ArrayScan VT1 of HILI + (green) and DAPI (blue)-stained colonies treated with with the low to middle concentrations of PFOS, PFOA, and PFNA assessed. All images are taken under the same imaging conditions and parameters. NIHMS1512545-supplement-Supp8.jpg (207K) GUID:?0B92D741-7D0F-4009-8539-B64042B33683 Abstract Per- and polyfluoroalkyl substances (PFASs) represent a highly ubiquitous group of synthetic chemicals used in products ranging from water and oil repellents and Rabbit polyclonal to ADAM17 lubricants to firefighting foam. These substances can enter and accumulate in multiple tissue matrices in up to 100% of people assessed. Though animal models strongly identify these compounds as male reproductive toxicants, with exposed rodents experiencing declines in sperm count, alterations in hormones, and DNA damage in spermatids, among other adverse outcomes, human studies report conflicting conclusions as to the reproductive toxicity of these chemicals. Using an innovative, human stem cell based PR-171 (Carfilzomib) model of spermatogenesis, we assessed the effects of the per- and polyfluoroalkyl substances perfluorooctanesulfonic acid (PFOS), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), and a mixture of PFOS, PFOA, and PFNA for their impacts on human spermatogenesis under conditions relevant to the general and occupationally exposed populations. Here we show that PFOS, PFOA, PFNA, and a mixture of PFOS, PFOA, and PFNA do not decrease germ cell viability under the conditions examined. However, PFOS, PFOA, and PFNA exposure do reduce expression of markers for spermatogonia and primary spermatocytes. While not having direct effects on germ cell viability, these effects suggest the potential for long-term impacts on male fertility through the exhaustion of the spermatogonial stem cell pool and abnormalities in primary spermatocytes. spermatogenesis, male reproductive toxicity INTRODUCTION Per- and polyfluoroalkyl substances (PFASs), previously called perfluorinated compounds (PFCs), are a group of synthetic chemicals that have been used in products ranging from water and oil repellents, lubricants, detergent products, coatings for furniture and food packages, waxes, firefighting foam, and other products since the 1940s (Arvaniti and Stasinakis 2015; Lei et al. 2015; Louis GM et al. 2015; Hu XC et al. 2016). Perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) are the two most widely produced and used PFASs in the United States, along with perfluorohexane sulfonic acid (PFHxS) and perfluorononanoic acid (PFNA) (Lei et al. 2015; Louis GM et al. 2015). PFASs enter the human body through ingestion, inhalation, and contact with commonly used consumer products, where they bind albumin in the blood stream and readily bioaccumulate within the bodys tissues.