Environmental and human relevant PFOS and PFOA doses alter human mesenchymal stem cell self-renewal, adipogenesis and osteogenesis

https://doi.org/10.1016/j.ecoenv.2018.11.064Get rights and content

Highlights

  • PFOS and PFOA impaired hMSC self-renewal and differentiation abilities.

  • Low-dose PFOS and PFOA stimulated adipogenic differentiation via PPARγ.

  • PFOA might induce hMSC osteogenic differentiation at human relevant concentrations.

  • PFOS and PFOA might exert developmental toxicity towards fat and bone tissues.

Abstract

PFOS and PFOA are two of the most abundant perfluorinated compounds (PFCs) in the environment. Previous studies have reported they have a long half-life (up to five years) once they enter into the human body. Moreover, they can potentially promote the adipogenic process by activating PPARγ. However, little is known about PFOS and PFOA chronic health impacts on humans. In this study, we employed primary human mesenchymal stem cells (hMSCs) and demonstrated that PFOS and PFOA exerted acute cytotoxicity and affected adipogenesis and osteogenesis at environmental and human relevant doses. In fact, PFOS and PFOA impaired the proper expression of CD90 (a surface antigen highly enriched in undifferentiated hMSCs) and promoted adipogenesis, presumably via their interaction with PPARγ. Moreover, PFOA partly disturbed osteogenesis. Thus, our findings not only validated the health risks of PFOS and PFOA, but also revealed new potential long-term PFOS/PFOA impacts on humans.

Introduction

Perfluorinated compounds (PFCs) are artificial compounds with widespread utilization in industrial and commercial products. Since PFCs are not easily degraded, they accumulate in the environment and animals. Therefore, people have raised many concerns about the potential adverse impact of PFCs on our health (Bjorklund et al., 2009, Paul et al., 2009).

Perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) are two abundant PFCs, with half-lives lasting several years (Butenhoff et al., 2006, Olsen et al., 2012). One epidemiological study showed that PFOS and PFOA could be as high as 880 ng/mL and 7320 ng/mL in blood samples of 3 M employees, respectively (Ehresman et al., 2007). Recent studies reported exceptionally high levels of serum PFOS due to either occupational exposure (up to 36,625 ng/mL), or fish-based diet (up to 10,400 ng/mL) (Gao et al., 2015, Zhou et al., 2014). Moreover, PFOS and PFOA have also been detected in human umbilical cord blood and breast milk.

Prenatal exposure to PFOS and PFOA correlated with adiposity in humans (Lauritzen et al., 2018, Mora et al., 2017; Halldorsson et al., 2012). Similarly, CD-1 mice receiving prenatal PFOA exposure became overweight as adults (Hines et al., 2009). Mechanistically, it had been shown that the activation of Peroxisome proliferator-activated receptor gamma (PPARγ) by PFOS and PFOA resulted in excessive adipogenesis and formation of lipids in zebrafish and mice (Ma et al., 2018, Mahapatra et al., 2017, Watkins et al., 2015, Yamamoto et al., 2015). However, this has not been proved in humans yet.

Studies also showed an influence of PFCs on bone health (Khalil et al., 2018, Lin et al., 2014) and their accumulation in bones and bone marrow (Bogdanska et al., 2011, Bogdanska et al., 2014, Celik et al., 2013). For instance, both PFOS and PFOA were shown to associate with changes in several bone health indicators, such as bone mineral density in femur and lumbar spine (Khalil et al., 2016). In addition, Koskela and colleagues reported that maternal exposure to PFOA gave rise to its accumulation in bones and reduced bone mineral density. In addition, PFOA could also induce osteoblast formation in primary bone marrow stem cells (Koskela et al., 2016). Another group found that PFOA treatment in MC3T3-E1 osteoblasts resulted in a dose-dependent decrease in cell viability, generation of reactive oxygen species (ROS) and reduction in anti-oxidant abilities (Choi et al., 2017).

Human mesenchymal stem cells (hMSCs) are multipotent stem cells found in multiple organs and tissues, and are able to self-renew and generate several lineages, such as adipocytes and osteoblasts (Jiang et al., 2002, Pittenger et al., 1999). CD44, CD73, CD90 and CD105 are commonly used surface markers for hMSC characterization. In fact: CD44 is involved in hMSC dependent survival, proliferation and migration (Yu et al., 2016); CD73 regulates osteogenesis (Hau et al., 2017); CD90 blocks the early commitment of hMSCs to differentiation (Moraes et al., 2016); and CD105 cooperates with transforming growth factor beta (TGFβ) superfamily to control early chondrogenesis (Izgi et al., 2017). PPARγ is the master transcription factor regulating adipogenesis of hMSCs. After binding with its ligands, PPARγ heterodimerizes with retinoid X receptor (RXR) and translocates into the nucleus where it binds to the PPAR responsive element (PPRE) and activates transcription of its target genes, such as Lpl (lipoprotein lipase) and Ap2 (adipocyte protein 2) (Chandra et al., 2008). Conversely, PPARγ signaling is inhibited during osteogenesis (Takada et al., 2009).

In this study, we treated hMSCs with human relevant doses of PFOS/PFOA in order to answer two major questions: (1) Is hMSC self-renewal affected by PFOS and PFOA? (2) Do PFOS and PFOA perturb hMSC adipogenic and osteogenic differentiation processes? We demonstrated that both chemicals altered hMSC self-renewal and differentiation potentials. Furthermore, while PFOA stimulated adipogenesis and maybe osteogenesis, PFOS enhanced adipogenesis only. Thus, our findings point out previously underappreciated potential toxic effects of PFOS/PFOA.

Section snippets

Chemicals

PFOS and PFOA were purchased from Sigma-Aldrich (PFOS: 77282; PFOA: 171468). Stock solutions were prepared by dissolving PFOS and PFOA in dimethylsulfoxide (DMSO) to achieve a final concentration of 100 mM. DMSO concentration in cell culture media was 0.01% for both chemical-treated and control groups, unless otherwise indicated.

Maintenance and differentiation of hMSCs

Three different hMSC lines were used in this study. One was purchased from the Cell Bank/Stem Cell Core Facility, SIBCB, CAS, and two, in-house derived, were kindly

PFOS and PFOA affected hMSC viability partially via ROS formation

Due to limited information on acute cytotoxicity in human cells and to select non-lethal concentrations for further experiments, we first tested PFOS and PFOA effects on hMSC viability after different doses’ treatment for 24, 48, 72 and 96 h (Fig. 1a lines for 24 h, and data not shown). We then calculated the IC50 values and found that PFOS exhibited slightly more cytotoxicity than PFOA under each measured time point (Table 1).

Since the generation of reactive oxygen species (ROS) is speculated

Discussion

Although volunteer phase-out of PFOS and PFOA production and utilization has been suggested, toxicity evaluations for those two chemicals are still urgent due to the accumulation of PFOS and PFOA in the human body (Karrman et al., 2010, Wu et al., 2015). It has been shown that PFOS and PFOA induce adipogenesis in murine pre-adipocytes via activation of PPARγ (Xu et al., 2016, Yamamoto et al., 2015), but so far little evidence is found in human models. Since PFOS- and PFOA-induced toxicity

Acknowledgments

This work was supported by the National Natural Science Foundation of China [grant numbers 21577166, 21461142001]; the Chinese Academy of Sciences [grant numbers XDB14040301, 29[2015]30, QYZDJ-SSW-DQC017]; and the K. C. Wong Education Foundation, Hong Kong, China. We would like to thank Drs. Xiaolei Liang and Kanghua Li from Peking Union Medical College, for providing in-house derived hMSCs, and Dr. Xinglei Yao for providing hMSC culture medium.

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