Gut microbiota limits heavy metals burden caused by chronic oral exposure
Introduction
Inorganic cadmium (Cd) and lead (Pb) ions are the most representative toxic non-essential elements which can contaminate food, water or air. In industrial areas, occupationally exposed workers as well as environmentally exposed populations can experience moderate to severe health perturbations due to the chronic ingestion of heavy metals. It has been shown that a significant fraction of inhaled Cd (60%) ends up in the gastrointestinal tract, as a result of mucocilliary clearance and subsequent ingestion (Satarug et al., 2003). Following oral entry of Cd and Pb, the body burden of these metals has been clearly linked to various sorts of diseases based on various mechanisms, including those involved in oxidative stress and extended toxicity such as genotoxicity and carcinogenesis.
The impact of gut ecology on the absorption, distribution, metabolism and excretion of xenobiotics has received little attention (Nicholson et al., 2005). However, the gut microbiota is likely an important mediator of the bioavailability and toxicity of environmental pollutants including heavy metals. On the one hand, the microbiota by itself may interact with metals inside the gut, either by active uptake or by passive ad- or absorbtion (Morozzi et al., 1986, Halttunen et al., 2008). On the other hand, intestinal barrier integrity, as the first line to control the entry of ingested toxic metals, also depends on microbial-host interactions that involve epithelial junctions and physical impediments of the mucous layer. Finally, the gut microbiota and its metabolites will also impact on environmental parameters such as pH, oxidative balance, detoxification enzymes, and xenobiotic-metabolizing and transporting host proteins (Claus et al., 2011), all of which may highly influence the bioavailability of chemicals in the gut lumen. The microbial status furthermore affects hepatic and renal metabolism.
While Louis Pasteur considered that germ-free life was impossible (Pasteur, 1885), today we know that germ-free mice are available and can be grown as far as proper technical requirements associated with rearing are achieved; the concept of the gut microbiota as a “forgotten organ”, essential for gut homeostasis, is also well accepted. Therefore, axenic mice, so called germ-free mice provide useful tools to study the interplay between the host and the gut inhabitants (Smith et al., 2007, Yi and Li, 2012). Germ-free animals are, however, physiologically different from conventional specific pathogen free (SPF) individuals and thus may show differences in the in vivo handling of xenobiotics (Ilett et al., 1990).
Recently, we reported on the dissemination to the intestine and to primary organs of continuous ingestion of low- and moderate environmentally relevant concentrations of Cd and Pb in mice (Breton et al., 2013). The aim of the current study was to address the role of the gut microbiota in the bioaccumulation and retention of Cd and Pb in primary organs following oral exposure. To examine these events, we used germ-free and SPF mice subjected to a chronic ingestion of various environmentally relevant doses of Cd and Pb. We also determined changes in gene expression of several intestinal markers, addressing aspects of transport- and oxidative stress functionality of the proximal (duodenum) and distal (colon) mucosa.
Section snippets
Animals
Forty germ-free (GF) female mice, aged 6–7 weeks and with a C57BL/6J genetic background were bred in the animal facility at the Transgenose Institute (TAAM, National Center for Scientific Research, Orléans, France) while 40 age-matched C57BL/6J female specific pathogen free (SPF) mice were obtained from Charles River (Saint-Germain-sur-l’Arbresle, France). Both groups (GF and SPF, respectively) were kept in separate isolators in a controlled sterile environment (22 °C temperature, 12 h daylight
General comments
While germ-free mice had a slightly higher (non-significant) food intake to compensate the lower metabolic rate of these animals, both groups showed similar daily drinking consumption, both for control untreated and metal-contaminated water. No significant difference was found for body weights at the end of the experiment. An oral gavage with FITC-dextran of 4 kDa was used as a marker of paracellular permeability for macromolecules. No differences in blood fluorescence were measured between SPF (
Discussion
Our results clearly show that mice lacking an intestinal microbiota are more susceptible to the accumulation of heavy metals in their blood and target organs, and this in a dose dependent way. Several facts, including the particular physiological and morphological properties of germ-free animals as well as a distinct basal expression and regulation of key genes involved in uptake and transport of metals, can explain this observation. Another intervening factor could be the direct interaction
Conclusion
Little is known about the impact of chronic ingestion of heavy metals on the digestive system itself, especially in the small intestine and the colon. The part of the ingested xenobiotics that are efficiently absorbed in healthy subjects is highly variable, depending on factors such as age, fasting and/or meal composition as well as speciation form and salt conjugates. For instance, Cd accumulation in tissue is associated with a basal metabolic rate in mice (Maciak et al., 2011). The role of
Conflict of interest
The authors declare no conflict of interest for any aspect of this research.
Acknowledgements
The present study was performed in the context of a French multidisciplinary project called “Mélodie-Reve” (Métaux lourds, désordres immunitaires et écotoxicologie intestinale – (bio)-Remédiation in vivo, standing for: Heavy metals, immune disorders and intestinal Ecotoxicology – in vivo (bio)-remediation), and supported by the National Research Agency (ANR-09-CES-016).
The authors thank the Dr Fabienne Jean for her support in the project. We are also very grateful to Dr Bernard Ryffel for his
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