Effects of Endocrine disruptors

Endocrine disruptors can interfere with sexual maturation, development of reproductive organs - abnormalities of the gonads or regression penis - or the reproduction and cause cancer in hormone-dependent. They may also disrupt the function thyroid, alter the immune system or cause disturbance behavior.

Many studies ecotoxicological aquatic organisms, especially on shellfish and fish have shown that these molecules could lead to imposex, that is to say, the change of sex of the animal. The fish are often very sensitive to contaminants such: in some estuaries of Great Britain or downstream of sewage treatment of wastewater, we can find fish populations hermaphrodites. In the flounder males living in a contaminated by pollutants hormone agonists, the oocytes appear among the sperm.

For several years, researchers suspect that many chemical compounds to be endocrine disruptors to humans. Meta-analysis published in the years 1990 showed the steady decline of quality of sperm from the man for 50 years, particularly in North America and Europe. The incidence of cancer of the testis increases for several decades in a number of European countries. There was a correlation between the presence of endocrine disruptors and abnormalities of the reproductive tract, for example between the presence of pesticides and cryptorchidism or between type compounds bisphenol A or dioxin and hypospadias. Among the women, there are function abnormalities ovarian, fertility, of fertilization, the pregnancy and implantation uterus and a shift of the age of puberty.

The doses at which effects occur may be low: ingestion by rats of 20 micro grams of bisphenol A, a compound which ethers are used to protect the interior of the cans, followed by estrogenic effects.

Endocrine disruptors may act in utero Seveso, it became a preponderance of female births among the population infected by the dioxins. It has been shown that DES was responsible for cancers of the reproductive system and affect fertility in men and women exposed in utero. Bisphenol A and diethylstilbestrol (DES) cause hypertrophy of the prostate of mice exposed in utero.

The synergy resulting from interactions among xenobiotics, micro-nutrients and drugs can exacerbate the effects: the simultaneous exposure of female immature to low doses of flavonoids and estradiol results in a strong estrogenic effect.

As with any chemical, the effect is dose related, so if the estrogenic activity of nonylphenol is made, the action potential is very low in comparison with natural estrogens: a study of risk assessment on mammals, prepared by the European Commission has clearly indicated that the harmful effects resulting from the disruption of hormonal system involved only for exhibitions much higher than general toxicity. For mammals, the declining population of seals in the Baltic, exposed to organochlorine compounds (DDE and PCD) could at least partly be explained by disruption of the endocrine and immune systems. In birds infected with low doses of DDT and / or PCBs, the eggshell is thin, and abnormalities of the reproductive defects and are more frequent.


Recent Studies

In China, a study showed a correlation between a sub-fertility in men living in Hong Kong who consumed more than four meals of fish per week, which also have mercury levels higher in the hair (also with problems skin and autism more common among children who have the highest mercury levels (measured in blood, hair and urine) 
In the reserve Native of Aamjiwnaang in the heart of the "chemical valley" of Canada, is suspected of products such as mercury, the dioxins, the HCB or PCBs to be responsible for changing the sex ratio observed: a study conducted by Constanze MacKenzie of the University of Ottawa, showed that the ratio at birth rose from a boy to a girl in 1984 to a boy two girls in 1999. In addition, the rate of miscarriage is 39% against 25% generally and 23% of children under 16 years suffer UNDERNOURISHED of ADHD (hyperactivity with attention deficit) instead of 4% normally.

Between the years 1999 and 2001, a study on the effects of a synthetic estrogen on aquatic populations have been made. The experience takes place on a whole lake in the Experimental Lakes Area in northwestern Ontario in Canada.

In 2000, 24 years after the occurrence of a plant herbicide Seveso (Italy), a study showed that men exposed to the cloud of dioxin were twice as many girls as boys.

In 2002, in Ufa (Russia), researchers have shown that workers of a factory of herbicides, contaminated with dioxin, had daughters in two-thirds of cases.


Research Status

Domain Search located at the confluence of biology, of chemistry and medicine, the study of endocrine disruptors is still booming.

Experiments are underway to assess the effect of endocrine disruptors on humans - such as decreased spermatogenesis and increased genital abnormalities - and the impact of the epigenetic on the sensitivity of certain populations. In the years 2000, research was extended to effects on the endocrine system, including the production of enzymes responsible for sexual differentiation.

The animal is used to develop tests for toxicity dose-response to differentiate the disruptive effect of the hormone-agonist in humans. For example, mouse wild reactions to normal and transgenic mice Arko (Aromatase knockout) - which are deficient in estrogen - are used to measure the effects of molecules with estrogenic activity (feminizing) - such as methoxychlor, an insecticide organo-chlorine - or anti-androgenic (anti-masculinizing) - such as vinclozolin, a fungicide. Molecules that have an estrogenic effect in vitro (test E-screen) are studied in vivo in rats or immature females who underwent oophorectomy.

Some field studies are done or in progress, including fish (eg chub or threespine stickleback in which the spiggin is a biomarker of exposure to androgens). The vitellogenin, which indicates exposure to estrogen agonists, is the biomarker most used. Some studies also look at the effects of metabolism of organic pollutants from oxidative stress and neurotoxicity of some disruptive or potential synergies. The activity aromatase should also be measured in France around 2008 (at the chub, sticklebacks ..).

Along with field studies, the mesocosms and microcosms are relevant models to study the effect of these molecules in the environment, particularly in the hydrosphere. Realistic point of view ecological and easily controlled mesocosms to assess the long-term effects on the biotic and fate of chemicals in biota. In 2004, the OECD organized an international campaign of intercomparison of methods of analysis of vitellogenesis in the zebrafish (Brachydanio rerio).

Methods of analysis and detection

A first approach is to use different organisms as biosensors. The doses to certain hormones and endocrine disruptors may act are so low that their analysis poses a challenge. Some sensing systems can provide a response to steroid concentrations in the range 10 to 1000 times lower than the assay techniques for anabolic agents (technical responds well to concentrations of nanogram per milliliter). Moreover, low doses are often a complex mixture and the biosensor reveals all these substances.

Examples of biosensors can be used in human cells (MCF7) in culture or species, laboratory rats, yeast, bacteria, zebrafish. Each biosensor has its peculiarities and its ethical problems. For example, human MCF7 cells were used as a biosensor of estrogen compounds for almost 20 years. These cells are derived from breast cancer and they have retained several characteristics as being stimulated by the presence of estrogen. Estrogen, or substances that mimic, function by activating a receptor protein in the cell, which regulates a whole set of genes, just count the cells within a few days to determine their presence.

Another in vitro test used is based on a strain of yeast with a gene expressing the human estrogen receptor, said YES (yeast estrogen screen). The yeasts have the ability, through a series of enzymatic reactions, to produce β-galactosidase that turns yellow starting mixture of pink, which absorbs at 540 nm ultraviolet-visible spectroscopy 

Plasma vitellogenin in can be measured in certain types of fish (salmon trout). Vitellogenin is a yolk protein synthesized by mature female fish. It is also produced in the male fish exposed to endocrine disruptor. Male fish exposed to estrogen producing egg proteins have smaller gonads and, in more severe cases, produce eggs. The biological response of male fish which is the rate plasma vitellogenin was significantly increased at sites with high estrogen activity. To enable the analysis of plasma containing the vitellogenin using an ELISA (enzyme linked immunosorbent assay)  In addition to vitellogenin, it has been demonstrated that it can be disruptive to bioaccumulate in the liver, bile and fat 

The various tests usually take place in rivers because they are the place prepared for the discharge of sewage. For a river flow is strongly affected by precipitation. The distribution of chemicals is dependent on weather conditions (temperature and UV) and microbial activity. It is therefore difficult to determine a constant concentration to which organisms will face. It is therefore necessary to obtain a sample in time. The first method to overcome this difficulty is the use of biosensors as fish.

Several other techniques exist for extracting the raw water or filtered. Water can be a carrier of endocrine disruptors on two counts: the molecules are more soluble or more concentrated can be trained in dissolved form. Regarding soluble molecules or less prevalent for some time in the environment, they may be associated with soil particles and sediments suspended in water. The estimation of water contamination must appear in its entirety. Finally the evaluation of contamination of water can be seen in terms of concentration of water or endocrine disruptor in terms of exposure. To alleviate the concentration, an automatic sampler or instant (called active) and can be used to determine exposure, as the use of passive sensors is possible.

The liquid-liquid extraction with dichloromethane is the most widely used, since this method has good performance, a wide spectrum and this technique is easy to implement. It uses the physical and chemical properties of the molecule of dichloromethane are polar immiscible with water and lipophilic. Another technique involves cartridges (SPE). These traps set molecules when water is passed through. A mixture of solvents and can be used to elute the cartridge and collect the desired molecules. The solution is to lower volume and constitutes a step of preconcentration. The extraction is selective to specific molecules since all products are not necessarily held by the cartridge and eluted by the solvent used as eluent. These sensors are often automated tools for making data in time intervals. However, this type of aircraft is subject to major drawbacks as the use of energy source, the number of mechanical breakdowns, disruption due to weather.

Another method uses a recent technique is the use of passive sensor that integrates contamination over time. Exposure rather than the flow of contaminants is estimated. The devices come in the form of pocket SPMD (for semi-permeable membrane device) sensors or POCIS (componds for polar organic integrative samplers). In these sensors there is a resin with a capacity to absorb specific hydrophobicity of the organic pollutants. The SPMD are used for hydrophobic compounds (PCBs, organochlorine) while POCIS are for the most hydrophilic compounds such as pharmaceutical residues. The POCIS with a phase for the retention of compounds such as pesticides (particles Isolute ENV + and Ambersorb1500 dispercé on S-X3 BioBeads) showed a better selectivity for xenoestrogens 

To achieve the quantitative tests the GC-MS or LC / MS / MS can be used. One of the methods used by the Government of Quebec for the determination of endocrine disruptors is the GC / MS. Hormones and other substances are extracted using Oasis HLB column. The extraction takes a volume of 1 liter and 14 days is to be achieved, the column preconcentrates analytes in a volume of 500 mg. Subsequently, diethyl ether is used to wash this phase, and conditioned with methanol. Washed again with a mixture of water / methanol in proportions of 60: 40 and a further elution with diethyl ether / water 90: 10. Purification on silica gel with the same solvent elution is the next step. Finally, before analysis by GC / MS with BSTFA deviation is achieved. The characteristics of the column and the various experimental parameters can be observed in the corresponding external link.


Means of Control

The risk prevention requires an understanding of properties toxicity of molecules and the restriction or prohibition of their use, many research programs have been implemented, the Directive and REACH should broaden the scope of knowledge in this area Even if some products are immune (including pesticides).


Other states are taking stringent measures. For example, marketing, manufacturing, import and export of toys and childcare containing phthalates have been banned in France in 1999. The nonylphenol was banned in Germany in 2003. Some pesticides which chlordecone and the atrazine (which seriously disrupted the development of frogs) have been banned, to mention only the most publicized. The use of DDT was banned worldwide in 1973, but then again it is found in the environment and the fat of many organizations, pressures exist to reauthorize its use in poor countries.

Wastewater, contaminated with various endocrine disruptors (eg xenoestrogens or drugs) are treated through sewage treatment plants (WWTP). In these treatments, the compounds may be fully or partially degraded, creating sludge by their adsorption on airborne particles. Various factors such as type of treatment or the residence time may influence the efficiency of removal of pollutants through STEP. The problem stems from the inefficiency of STEP to retain certain substances, it follows that they are found in the treated water which will then contaminate surface waters. It has been demonstrated that the additional treatment of wastewater with ozone or activated carbon allows better removal of micropollutants. Unfortunately the economic efficiency of various options for upgrading sewage treatment plants encounter barriers at various levels of governments.


Strategies


Worldwide

Several states have programs on this subject (Japan, USA, Switzerland, France.). L' OCDE y travaille également, avec la commission européenne , dont sur les méthodes d'essai et d'évaluation des PE ; d'abord pour la santé humaine puis pour l'environnement. Various international commissions to include the protection of oceans (OSPAR, HELCOM ...) are also interested in these issues.

research usually covers the following topics:
The screening, ie the analytical methods (chemical, biological tests, biomarkers of effects) to determine the levels of these pollutants in the environment;
The extent of the distribution and life in the environment (water, soil, air, sediment, food ..) and / or living organisms (monitoring, biomonitoring ...)
Identifying hazards and assessing risks, measuring the degree of impact strength (in some species considered representative or easy to study);
Understanding the mechanisms of action (and possible effects epigenetic), but also natural degradation of these molecules by écocinétiques and toxicological studies of PE;
The study of metabolic pathways in the body and the key stages of biodegradation;

Population studies and landscape epidemiology have highlighted long-term effects. They have developed animal models and begin to develop models of the effects of endocrine disruptors.


United States: For example, the EPA (Agency for Environmental Protection of the United States) has set a target to test thousands of different chemicals in the coming years to verify the effects of endocrine disruptors.

Canada: Measures of polyethoxylated alkylphenols used in the manufacture of pesticides have been collected from several streams.

Switzerland in the early 2000s, the Federal Council on National Fund has launched a research program in Switzerland (NRP 50 to develop strategies to assess the risks and dangers of life exposure to endocrine disrupters, for 2001-2007; 17 projects funded in 2001 and another 7 in 2003, leading to conclusions and recommendations for authorities and industries to limit the negative effects of these substances and compounds developed future.

France: A national program of research on endocrine disrupters "was launched in 2005, in conjunction with the NRA and the AFSSET, following the recommendations of the Committee of Prevention and Precaution (CPP) following a referral on endocrine disruptors (findings published in December 2003), and the announcement in June 2004, minister in charge of the Environment S. Lepeltier. A new call for projects was launched mid-2008 targeting "neglected issues", including the impact on thyroid function or immune system.


European Union

The EU has established a "Community Strategy for Endocrine Disrupters" to "address an urgent problem of endocrine disrupters harmful to health and the environment" . This strategy provides for three types of measures: short, medium and long term.

In the short term, the EU is to evaluate a priority list of molecules ( "priority list PE) highly suspicious of hormonal effects, identifying gaps in knowledge. The EU wants to use existing legislative instruments, including REACH (Regulation on risk assessment) and the directive on classification of dangerous substances, to establish monitoring programs that must estimate the doses and durations of exposure to PE substances "priority" and study its effects. It is also intended to identify possible cases of individuals and groups vulnerable to certain substances (eg, children ...) if these substances are not already regulated, they should be included on the priority list of PE; networks of exchange, information and consultation of stakeholders (international, Commission, Member States, industry, etc..) are also provided. The EU wants a "fully informed" of the population, including the activities undertaken, the mechanisms, the priority list PE. The EU is funding several studies since the 2000s, including through interregional Interreg funds.
In 2000, 553 artificial substances and 9 synthetic hormones and natural have been listed in the "priority list". Priority actions have been listed to evaluate the role of these substances, and a conference was organized by the Commission on endocrine disrupters (18-20 June 2001, Sweden). A call for research proposals in May 2001 has allocated a budget of 20 million euros. The White Paper on the strategy for future policy on chemicals includes the EP and the forthcoming directive on general product safety should allow for easier "emergency measures at Community level. A Community Strategy on Mercury, which is also an endocrine disruptor has also been a communication 

In the medium term, the EU wants to help identify and assess endocrine disrupters, develop alternative products and tests to identify the EP, including through the Fifth Framework Program for Research and Development and private initiatives.

In the long term, the EU is to adapt / amend the legislation to take into account the EP, including through the Council Regulation (EEC) No 793/93 on the evaluation of risks and Directive 67/548/EEC on the Classification hazardous substances. The good ecological status under the Framework Directive on water must incorporate aspects PE.