Chemo Defence System

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Concise review paper / Point sur

Chemo-defence system

Andr Rico*

24, rue Balard, 75015 Paris, FranceReceived 22 May 2000; accepted 16 October 2000Communicated by Roger Monier

Abstract By analogy with the immune defence system, the existence is suggested of achemo-defence system protecting living organisms against toxic substances, whethernatural or man-made, that are present in the environment. This paper deals first with thevarious facets of such a system: mechanisms involving, among others, lipophilic com-pounds, hydrophilic compounds, oxidants, acidosis, genotoxics and metals; second,with the biological characteristics of the system and a comparison with the immunedefence system : partial immaturity of the young, inducibility, non-specificity andspecificity, and saturability; we will show that the two systems share many common

features; third, with the evolution of the system, which demonstrates that the system isvery old and suggesting that it came into existence before the immune defence system;and fourth, with some of its consequences: estimation of the toxic effects of low doses,hormesis, impact of a vegetable diet on health. Finally, it could be emphasised that life iswell protected against chemicals by its chemo-defence system, which appeared veryearly with the first living organisms on the earth. 2001 Acadmie des sciences/ditionsscientifiques et mdicales Elsevier SAS

chemo-defence / immune defence / biotransformations / low doses / hormesis / fruit and vegetable diet

Rsum Systme de chimio-dfense. Par analogie avec le systme immunitaire, onpeut estimer quil existe un systme de chimio-dfense protgeant les tres vivants des

innombrables toxiques chimiques naturels ou de synthse prsents dans leur environ-nement. Dans une premire partie, sont discuts les principaux mcanismes de cesystme, prsents suivant certaines caractristiques physico-chimiques ou biologiquesdes toxiques : composs lipophiles, composs hydrophiles, oxydants, acidifiants, muta-gnes, mtaux. Dans une deuxime partie, sont dcrites les caractristiques biologiquesdu systme : partielle immaturit chez les jeunes et inductibilit, non-spcificit etspcificit, saturabilit. Une comparaison est faite avec le systme immunitaire montrantque ces deux systmes sont de fait biologiquement assez proches. Dans une troisimepartie est aborde lvolution du systme dmontrant aussi sa ralit. La dernire partietraite de quelques consquences de lexistence de ce systme : effets des faiblesdoses , concept dhormesis et hypothse daction protectrice dun rgime vgtarien, enparticulier sur le dveloppement des cancers. Finalement, il apparat que la vie est bienprotge des toxiques chimiques par son systme de chimio-dfense apparu trs tt surterre avec les premiers organismes vivants. 2001 Acadmie des sciences/ditionsscientifiques et mdicales Elsevier SAS

chimio-dfense / systme immunitaire / biotransformations / faibles doses / hormesis / rgimesvgtariens

* Correspondence and reprints.E-mail address:[email protected] (A. Rico).

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C.R. Ac ad . Sc i. Paris, Sc ienc es de la vie / Life Sc ienc es 324 (2001) 97106 2001 Ac ad m ie des sc ienc es/ d itions sc ientifiq ue s et md ica les Elsev ier SAS. Tou s d roits rservsS0764446900012816/ FLA


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Version abrge

La vie sest dveloppe depuis son origine dans unenvironnement chimiquement et physiquement hos-

tile : agents physiques (rayons UV solaires, rayonne-ments ionisants), composs chimiques prsents dans lemilieu (mtaux lourds, oxygne) ou synthtiss pardautres organismes vivants pour agresser les proies ou

se dfendre contre les prdateurs.Il est donc normal quau cours de lvolution soientapparus des mcanismes de dfense. Lobjet de cettenote est dindiquer les analogies qui existent entre lesystme immunitaire qui nous protge des agressionsdes organismes pathognes et certains mcanismes dedfense contre les toxiques chimiques qui peuvent treconsidr comme partie intgrante dun systme homo-logue.

Dans cette note, nous illustrerons ce concept parquelques exemples. Nous proposerons une classifica-tion des mcanismes en cause, nous soulignerons lesanalogies existantes entre systme immunitaire et sys-tme de chimio-dfense et nous montrerons que laprsence de ces mcanismes infirme la navet delhypothse selon laquelle toute quantit, si faiblesoit-elle, de toxiques chimiques fait courir un risque lorganisme vivant, ce qui amnerait la conclusionquil faut faire entrer les tres vivants dans des coconsprotecteurs les mettant labri de tous toxiques physi-ques ou chimiques.

Lintroduction de ce concept amne, au contraire, se demander si, comme pour le systme immunitaire,la prsence de petites quantits de produits toxiquesnest pas ncessaire pour assurer le fonctionnementcorrect du systme chimio-dfense grce des stimu-lations priodiques et pour assurer son adaptation aumilieu.

Les produits chimiques peuvent tre classs en fonc-tion de leurs proprits physiques, chimiques ou bio-logiques. Les composs dits lipophiles ont les plusgrandes potentialits toxiques. Leur lipophilie leur per-met de traverser aisment les membranes, donc dtrefacilement absorbs par lintestin, la peau ou le pou-mon, sils sont volatiles. Ils pntrent aussi aismentdans les cellules et sils ne sont pas biotransforms ilssaccumulent dans les graisses, voire dans le cerveau. Ilexiste heureusement des processus de biotransforma-tions impliquant en particulier les cytochromes P 450,qui agissent sur leur structure pour les rendre pluspolaires et faciliter leur limination par la bile et lurine.

ces biotransformations, il faut ajouter les transpor-teurs multidrogues (MDT) existant dans toutes lesmembranes des cellules des tres vivants et plus parti-culirement exprims dans les pithliums, structurestissulaires en contact avec les substances chimiqueslors de leur absorption (tractus digestif) ou de leurexcrtion (rein). Les MDT sont non spcifiques et

expliquent la rsistance exceptionnelle aux antibioti-ques de certains microorganismes tels que Pseudomo-

nas aerginosaposant un rel problme de sant publi-que.

Les composs hydrophiles sont nettement moinsbiodisponibles que les prcdents, mais certains sonttoxiques aprs injection. Cest le cas, par exemple, descurares. Ces composs hydrophiles subissent aussi des

processus de biotranformations non-cytochrome P 450dpendants, assurant aussi leur dtoxification et leurlimination.

Loxygne etles espces oxydantes associes, commelion superoxyde ou leau oxygne, sont des toxiques.Les oxydations dgradent les substrats, augmententlentropie du systme donc le dsordre. La respirationcellulaire arobie peut tre assimile un mcanismede dtoxification des potentialits toxiques de loxy-gne avec rcupration deffets biologiques positifs(synthse ATP). Quant aux superoxydes dismutases oucatalases, elles permettent de lutter contre les hyper

oxydants et compltent laction de la respiration cellu-laire.La rgulation acido-basique du pH sanguin peut tre

aussi considre comme un processus de dtoxifica-tion en particulier vis--vis des acides.

Les gnotoxiques sont des composs dangereuxcapables daltrer lADN. Cependant, dans les cas desfaibles doses, il existe des systmes de rparationefficaces et varis. Ils sont constitutifs et inductibles. Parcontre, si la dose ou le dbit de dose est lev, laprobabilit de rparation fautive saccrot rapidement.

Les mtaux sont aussi dangereux, mais ils peuvent

tre dtoxifis par les organismes vivants par com-plexation avec en particulier les mtallothionines,protines soufres rencontres chez toutes les espces.

Le systme de chimio-dfense prsente un certainnombre de caractristiques biologiques analogues celles du systme immunitaire. Le systme de chimio-dfense est en partie immature chez les jeunes, ilsexprime par adaptation des organismes dans leurmilieu soit par expression gnique soit par induction.Une question qui se pose est de savoir si l inductiondoit tre considre, comme cest souvent le cas entoxicologie classique, comme un effet non dsir. Cet

aspect est discut. Le systme de chimio-dfensecomme le systme immunitaire comprend une compo-sante non spcifique et une composante scifique. Enparticulier, une analogie et une comparaison des struc-tures entre cytochromes P 450 et immunoglobulines

peuvent tre faites, expliquant dans les deux cas lagrande spcificit de substrats exprime par ces deuxtypes de molcules.

Enfin le systme de chimio-dfense, comme le sys-tme immunitaire, est saturable. Cest la dose qui fait lepoison, cest le quantum bactrien ou viral qui peut

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dclencher la maladie. Les deux systmes prsententaussi des dsordres, par exemple de type hrditaire.

Ltude de lvolution de ce systme avec le tempsest aborde. Elle est intressante en ce sens quelledmontre sa ralit et certainement son antriorit ausystme immunitaire.

De lexistence de ce systme de chimio-dfense, onpeut tirer diffrentes consquences. Trois dentre elles

sont voques. a) Effets des faibles doses : il est clairque lvaluation toxicologique actuelle gnralementfonde s u r lextrapolation linaire est discutable etconduit au final des limites tolrables plus dordreanalytique que toxicologique avec tous les inconv-nients que cela entrane, en particulier une chimiopho-bie de la socit. b) Le concept dhormesis : il indiqueque les faibles doses peuvent avoir des effets bnfi-ques, ce qui peut tre considr comme un corollaire

de lexistence dun systme de chimio-dfense. c) Lerle anticancreux tabli des rgimes base de fruits etde lgumes : celui-ci est gnralement considrcomme li lexistence dantioxydants en nombreimportant dans ces aliments. Cependant, pris isolmentces composs (-carotnes, vitamines E, vitamines C,etc.) ne manifestent pas exprimentalement de tellesproprits. Une autre approche consisterait dire que

cest la complexit des structures chimiques rencon-tres chez les vgtaux par rapport celles rencontresdans les produits animaux, qui, en stimulant notre

systme de chimio-dfense, nous protge efficacement.

Au total, on peut donc dire que la vie est bien

protge des substances chimiques par son systme dechimio-dfense apparu trs tt sur terre avec les pre-miers tres vivants.

1. IntroductionWe are surrounded by bacteria, viruses, parasites and

prions, but we survive, mainly because our body is pro-tected by an immune defence system. We live in an oceanof chemical substances (carcinogens, mutagens, terato-gens, endocrine disrupters, etc.). Most of these are naturalxenobiotics which can be highly toxic (mycotoxines, alka-loids, heterosides, etc.). Man-made chemicals (pesticides,veterinary drugs, etc.) are also present in our environment.Sometimes in pathology problems arise from abnormalmetabolites. Nevertheless, we survive. How?

By analogy with protection against infectious diseases,we would like to suggest the existence of a system thatincludes a large number of mechanisms and that we couldcall the chemo-defence system.

When living organisms appeared on the earth, theywere confronted with a very hostile environment with ahigh flux of UV light and ionising radiation and manyinorganic and some organic chemicals. In order to survive,the living organisms developed defence mechanisms suchas DNA repair systems.

When oxygen, which is toxic and was present at tracelevels in the beginning of life, became more quantitativelyimportant, some bacteria detoxified it by way of aerobiccellular respiration. The chemical medium became morecomplex with the plants. Thus, to face the many neworganic chemicals, initial cytochrome P 450s, devoted atthe beginning to metabolic pathways, became more diver-sified to detoxify complex liphophilic chemical structures.Subsequently, the multi-cellular organisms used all thesechemo-defence mechanisms and organised them so as tosurvive. It is this complex organisation that could bedefined as the chemo-defence system.

I introduced this new concept very recently but onlyrelatively briefly [1]. In this basic conceptual research

paper, I would like to go into more scientific details tosupport my conceptual model.

This paper will deal with the nature and function of thechemo-defence system, together with its biological char-acteristics, evolution and consequences. However, it willnot cover all the toxic effects due to chemicals in detail.The same applies for references; it is not possible to cite allthe articles devoted to the different sections. A choice hasbeen made, but as in all choices, it could be open tocriticism.

2. Nature and functions

Chemicals can be classified according to their physicalproperties, chemical properties or biological effects. Weshall use this classification to discuss the major differentcategories of toxic products and the processes of detoxi-fication associated with them.

2.1. Lipophilic compounds

In general lilophilic compounds present the greatestpossibility of toxicity. Their lipophilic properties facilitatetheir penetration into the lipid membranes. This meansthat they are readily absorbed by the gut, they can gothrough the skin and, if they are volatile, they can rapidly

penetrate the lung tissue.If they are not rapidly detoxified, some (e.g. organochlo-

ride insecticides such as DDT) may accumulate in fattytissues, others (such as methyl selenium) can go throughthe bloodbrain barrier, often with highly toxic effects.They may also penetrate the cells. Their volumes of distri-bution, in terms of pharmacokinetics, are always high tovery high.

2.1.1. Biotransformations

Biotransformation processes exist in some organs, par-ticularly the liver, but also in the intestine, lung, kidney,

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skin and testes. These biotransformation processes areable to transform the structures of lipophilic xenobiotics,thereby increasing their polarity, facilitating their elimina-tion by bile and urine and most often decreasing theirtoxicity [2, 3].

Classically there are two phases in this process.

Phase I: this is a structural biotransformation involvingan enzyme system referred to as the mixed function oxy-dase (MFO) system. In this system, cytochrome P 450mono-oxygenases play an important role. They are con-stituted by a super family of more than 400 hemothiolate-enzymes, with a great affinity for lipophilic substances.These enzymes permit the introduction of hydroxyl groupsintothe structure of lipophilic compounds, which increasestheir polarity and facilitates excretion. Phase II, or conjugated phase: the biotransformationproducts and others, if their structures are adequate,become conjugated with polar agents of conjugation,which increase their urinary and biliary excretions. This isthe case with glucuronic acid, sulphates, etc.

Almost all these enzymes are inducible.

The cytochrome P 450s are very efficient and tworemarks can be made about their action.

In general, biotransformations are processes of detoxifi-cation. In some cases the action of cytochrome P 450s canlead to bioactivation and produce aggressive electrophilicmetabolites [3]. These metabolites are attracted by thenucleophilic sites, present for example in proteins andDNA, and can bind covalently with these targets and oftenexhibit toxic effects. They are suspected of genotoxicityand carcinogenicity. However, at low dose levels, theseelectrophiles are trapped by the glutathion present in thecell cytoplasm. As for the residues of pesticides or veteri-

nary drugs, the process of bioactivation leads to the for-mation of covalently bound residues which are consid-ered not to be of toxicological significance for consumersof food containing these residues [4].

Some cytochrome P 450s are not involved in detoxifi-cation but are essential to the synthesis of steroid hor-mones catalysing, for example, the aromatisation reactionthat produces oestrogen in the female sex organs but alsoin the oxidative metabolism of fatty acids, prostaglandins,leukotrienes, biogenic amines and pheromones.

To summarise, it would seem that the biotransforma-tions of lipophilic chemicals by the MFO system globallyrepresent much more a process of detoxification thantoxification at least at low dose levels.

2.1.2. Multidrugtransporters (MDT)

In organisms ranging from bacteria to humans, processtransmembrane transportersare present which confer resis-tance to toxic compounds [5]. On the basis of bioenergeticand structural criteria these transport systems can bedivided into two categories:

ATP-binding cassette (ABC) transporters, which use therelease of phosphate bond-energy by ATP hydrolysis topump drugs out of the cell;

secondary transporters, which mediate the extrusion ofdrugs from the cell in a coupled exchange with ions.

P glycoproteins (Pgp) are plasma glycoproteins of about170 kDa and belong to the superfamily of ATP-bindingcassette transporters. Humans have two Pgp genes, MDR1(class I) and MDR3 (also named MDR2, class II) located onchromosome 7. They are expressed to a greater of lesserextent in many normal tissues. High levels are found intissues with absorption or excretory functions, especiallyin the apical plasma membrane of intestinal epithelialcells and the proximal tubules of the kidney. These couldexplain the resistance of cancers derived from epitheliumcells to chemotherapy. On the contrary, the high sensitiv-ity of CF-1 mice to the toxicity of avermectins, used asantiparasitic drugs and pesticides, is associated with Pgpdeficiency in the small intestine and the capillary endot-helial cells of the bloodbrain barrier [6, 7].

Finally, the resistance of micro-organisms to antibioticsis in part related to the existence of MDT [8, 9]. It is thesame for the chemo-resistance of cancer cells [10].

Such systems could be considered as an important part

of the chemo-defence system [11] and are also usuallydirected more specifically against lipophilic compoundsthat enter the cells [12].

2.2. Hydrophilic compounds

These compounds, because of their structure, showlimited, if any, absorption by the gut or through the skin. Ifthey penetrate the body after parenteral administration, forexample, they do not penetrate the cells, which are pro-tected by their lipophilic membranes. In terms of pharma-cokinetics, their volumes of distribution are low to verylow. They are rapidly excreted by the kidney and are not

accumulated. However, they can be toxic in the body,often by acting on the cell membrane receptors as someanalogous polar hormones (e.g. 2 agonists) [13]. Curaresare good examples. Their oral bioavailability is very low,so they are not toxic by this route, but have a potentparalysing action on the muscles after injection. Strepto-mycin is also a highly polar compound and is not absorbedby the gut or active by oral route.

These compounds are also subjected to biotransforma-tions, not involving cytochrome P 450s,but other enzymes,such as epoxidehydrolase, esterase and amidase, or areconjugated. These processes decrease the toxicity of these

compounds and facilitate their excretion. Some of themare also inducible.

2.3. Oxidants and oxidative species

Oxidations, or at least full oxidations, can be viewed astoxic aggressions. These reactions are exergonic reactions,which destroy substrates, increase the entropy of the sys-tem, and in other words introduce disorder.

Substrate dehydrogenations are in fact also oxidationprocesses. Oxygen is interesting to consider. Nature hasfound an efficient process to combat its toxicity: the cellu-

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lar aerobic respiration that takes place in the mitochondria[14, 15]. The oxidative phosphorylation process consumesoxygen, reducing it in sequential steps to produce waterand create adenosine triphosphate (ATP). This high-energymolecule is then used in the endergonic reactions involvedin substrate synthesis. These endergonic reactions decreasethe entropy and disorder of the system. Cellular aerobicrespiration thus provides a perfect illustration of the poten-tial of living organisms to transform what could be toxic

effects into beneficial effects.By-products of oxidation such as the oxidising radicals

superoxide anion and H2O2 are also very aggressive,leading to what is known as oxidative stress [16]. How-ever, several means of defence against oxidative stressexist which maintain such oxidants at a low steady state.The superoxide anion is eliminated in the mitochondria bya specific Mn-containing superoxide dismutase, similar tothe bacterial forms. This enzyme is inducible. Theseenzymes also exist in the cytoplasm but they containcopper and zinc.

H2O2 can be eliminated by glutathion peroxidase and

catalase. Catalases are present in great quantities in theleukocytes and it should be noted that H

2O

2is formed

during inflammation.These protective mechanisms are very important

because it is now very well known that processes ofoxidation are involved in many toxic or natural effectssuch as mutations, carcinogenesis, damage to lipids anddegenerative diseases associated with ageing [17].

2.4. Acidosis

In some diseases such as diabetes mellitus or in shockphenomena, a syndrome of acidosis occurs by the accu-mulation of acid metabolites, which can lead to death.This syndrome can be compared to a toxic effect. It is wellknown that the blood pH is maintained between 7.30 and7.35. A decrease in the pH to 7.20 is lethal. Two mecha-nisms of regulation maintain this pH at the normal level.One is a physicochemical system constituted by the buffersystems in the plasma (carbonic acid/bicarbonate, etc.).This system becomes active immediately, but its effect islimited.The second is a physiological mechanism involvedin the excretion of acids by the kidney, which eliminatesprotons, and the lung, which eliminates carbon dioxide.These two systems work synergically to efficiently regulate

the blood pH and can be viewed as detoxifying mecha-nisms against acidic toxics.

2.5. Mutagens

Mutagenicity can be defined as an irreversible alterationof the DNA molecule which may, for example, lead tocarcinogenicity. Many natural and synthetic mutagensexist in our environment. In toxicology one importantquestion concerns the bioactivation process [2, 3]; bioac-tivations can lead to highly reactive electrophilic metabo-lites which are able to attack DNA. It should also be

stressed, however, that such bioactivations are located inthe endoplasmic reticulum, so that after formation theseelectrophiles have to go through the cytoplasm, penetratethe nuclear membrane, go through nucleoproteins, basicproteins with many nucleophilic sites, in order to reachand attack the DNA. On their way, the electrophilesencounter numerous nucleophilic sites present in the pro-teins or specific detoxifying substrates such as glutathion.In fact, we can say that the nucleus is the safe box of the

cell, in which the most important molecule for its survivaland future is DNA and that this molecule is very wellprotected [18, 19].

So if production of electrophilic compounds is certainlydose dependent, this is not the case for their mutageniceffects, because at low dose the flux of electrophilescreated by bioactivation is too low and has little chance ofmodifying DNA. This strongly suggests a threshold for thiskind of toxic metabolite. The existence of a threshold formutagenicity and carcinogenicity has already beenstressed by many authors. Several papers have challengedthe validity of the linear no threshold relationship (LNT),

concerning radiation [20, 21] and chemicals [22].Even if a modification of DNA takes place it is now wellestablished that numerous and various DNA repair sys-tems exist that are able to correct errors of replication orexternal aggressions. They are very efficient, present inalmost all tissues, including skin, and are also inducible[19].

2.6. Metals

Metals may be toxic [23]; however, it is important tostress that in fact the term toxicity in the case ofa metal isnot toxicologically correct. What is important as regards

metal toxicity is what we term speciation (the chemicalform of the metal) and the bioavailability (absorption afteringestion). These aspects are very wellknown, for example,for mercury, lead, chromium, arsenic, etc.

The toxicity of metals is related to their ionic form; andsuch toxicity is lost when they are chelated or complexed.

Substances able to chelate metals and thus to detoxifythem exist in the body. This is the case of metallothioneins,sulphur proteins formed in water organisms, mammalsand birds [24]. These molecules are able to chelate heavymetals and decrease their toxicity (mercury, cadmium,etc.). Ferritin and hemosiderin act in the same way for

iron. The metallothioneins are inducible.In summary, numerous mechanisms exist that candetoxify the chemicals which penetrate the body. In fact,these mechanisms do not work separately but together andoften synergistically. For example, when the MDT pumpthe toxic compounds in the blood outside of the cells,these compounds will be detoxified through biotransfor-mations, specifically in the liver and the kidney, andexcreted. These mechanisms can be also complementaryor redundant. They are complementary when during aero-bic respiration some reactive oxygen species appear inmitochondria. These species are eliminated by the super-

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oxide dismutase enzyme. They are redundant in the pro-tection of the genome: in the case of mutation, the variousDNA repair systems act to restore the integrity of themolecule. However, other mechanisms participate in thisrestoration, such as the arrest of cellular division, to givetime to the cells to repair the lesions before mitosis (controlpoints in G

2and in phase M in the cellular cycle) [25, 26],

or apoptosis, which eliminates the abnormal cells [27].

3. Biological characteristics

3.1. Immaturity and inducibility

It is well known that many biological differences existbetween the infant, the young and the adult [28]. In termsof chemo-defence the very young are partially immature.

The metabolism of xenobiotic compounds in the new-born is both qualitatively and quantitatively different fromthatin adults. The system changes progressively and adjustsfollowing contact with toxic compounds of the environ-ment. This can be considered as a consequence of theexpression of existing genes and inducibility.

Many of the enzymes participating in the chemo-defence system are inducible. This is particularly the caseof cytochrome P 450s, some conjugases, DNA repairenzymes, metallothioneins, etc. The question is whetherinduction has a beneficial, adverse or toxic effect. Inexperimental toxicology, in long-term toxicology tests,this kind of biological effect is often associated with anincrease in liver weight. Its significance is a matter ofjudgement. It seems that when this effect is not associatedwith histopathological changes or any other biologicaleffect, we can consider that it is an adaptive if not benefi-

cial effect, especially if we accept the fact that the cyto-chrome P 450s have specific substrates, as will be seen inthe next chapter.

Remark: These characteristics are very similar to thoseof the immune defence system which is also immature inthe newborn and is also inducible. Even more so, thepresence of antibodies, which are induced by pathogens,is not always a sign of illness but rather a sign of the bodyscontact with the germ.

3.2. Non-specificity and specifity

The functions of the immune system are provided bytwo major mechanisms: a non-specific or constitutivemechanism not requiring prior contact with the inducingagent and lacking specificity, and a specific or adaptivemechanism directed against and specific for the elicitingagent [29, 30]. The same is true for the chemo-defencesystem.

3.2.1. Non-specificity

Multi-drug transporters are non-specific mechanisms.As it was stressed above, they exist more or less in alltissues and there are no really specific inhibitors showing

that the effect is not related to any specific structure [6].This explains in part the difficulties of chemotherapists tofight against chemo-resistant cancer cells [31].

3.2.2. Specificity

3.2.2.1. Organ specificity

Not all organs or tissues are involved in the same way inthe process of chemo-defence, especially in the case of

biotransformations, which are pivotal in terms of detoxifi-cation. The liver is the main organ. All the cells in thisorgan are able to participate in the process, and are able toextract a considerable quantity of toxic agents and excretethem via the bile. The importance of this organ is notsurprising, since a primary function of the liver is toreceive and process chemicals absorbed from the gastro-intestinal tract.

But we also find processes of detoxification to a lesserextent than in the liver in those organs involved in theabsorption of toxic compounds (lung with clara cells,intestine, skin), in their elimination (kidney, lung), but also

in the testes where there is continuous production ofspermatozoa essential to species reproduction and sur-vival. These organs constitute an organ defence systemsimilar to that of the immune system.

3.2.2.2. Substrate specificity

Many enzymes involved in detoxifying processes exhibitchemical function specificity but not substrate specificity.This is true, for example, of esterases, amidases, epoxihy-drolases, etc. The original consensus was that the cyto-chrome P 450 systems were not highly specific to sub-strates, but were essential to transform lipophiliccompounds. However, in the light of recent research, a

brief discussion of this point may be of interest [32, 33]. Itis becoming clear that all cytochrome P 450s possess aconserved structural core. This core contains the activesite and heme pocket, and the parts of the protein involvedin the integration of the enzyme into the phospholipidmatrix of the endoplasmic reticulum. This environmentallipid matrix explains the affinity of the system for lipophiliccompounds.

However, although the structural core is apparentlyconserved among P 450s, there is sufficient variabilitybetween the protein sequences for it to be said that nocytochrome P 450s are structurally identical. This variabil-

ity is without any doubt related to substrate specificity,which certainly exists, but the situation is so complex thatup to now it has been very difficult to identify it clearly.However, in the future it should be possible to say: Tellme what you eat and I will tell you what specific cyto-chrome P 450s are in your liver.

An interesting point is to compare the cytochrome P 450structure with that of the immunoglobulin molecules [30].An immunoglobulin consists of four peptide chains, twolight chains and two heavy chains, held together by disul-phide bonds. Furthermore, each heavy and light chain issubdivided into a variable and a constant region. It is the

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variable region that determines the molecular specificityfor antigens while the constant region of the heavy chainsis responsible for the biological activity of the molecule.

The structure of cytochrome P 450s is similar to that ofimmunoglobulins, againsupporting an homology betweenthe immune defence system and the chemo-defence sys-tem.

3.3. Saturability

The chemo-defence system is saturable. In other words,when the quantity of toxic compounds becomes too high,the system is unable to control them, and toxic effectsoccur. This aspect is well illustrated by the basic toxico-logical principle: It is the dose, which makes the poison,stated first by Paracelse and later emphasised by ClaudeBernard

But this is also true of the immune system. It is thebacteria or virus quantum that can launch the illness. Thequantum is dependent on the nature of the agent and itspathogenicity in the same way as the dose in toxicology isa function of the nature of the toxic compound togetherwith its potential toxicity. The targets for the immunesystem are living organisms, whereas for the chemo-defence system they are chemical products. But in bothcases, the aim is the same: to neutralise the undesirableelements, by killing them or changing them structurally todecrease their toxicity.

3.4. Dysfunction

Some dysfunctions of the system are due to the absenceof some part of the process. For example, xerodermapigmentosum is a hereditary disorder due to a deficiencyin some DNA repair mechanisms in the skin. This disordercauses an increased sensitivity to UV light, with a greatly

increased incidence of skin cancer and premature death.The absence of certain enzymes can also explain thegreat sensitivity to certain compounds in different species.The cat is deficient in sulfotransferase, so it is much moresensitive than other species to phenol compounds. Dys-functions also exist in the immune defence system ashypersensitivity, allergy or autoimmunity.

To summarise, the chemo-defence system presents bio-logical characteristics that are similar from many points ofview to the immune defence system: partial immaturity inthe newborn, inducibility, non-specificity and specificityof organs and substrates, saturability and dysfunctions.These biological characteristics could support the exist-

ence of a chemo-defence system.

4. Evolution

We emphasise that almost all of the mechanismsdescribed in section 2 appeared with the first living organ-isms on the earth.

4.1. Cytochrome P 450s

The structure, evolution and regulation of P450 genesare clearly described by Nebert and Gonzales [34]. Theystated:

In the present review we describe the gene structureand evolution of the 10 known P450 gene families, eightof which exist in mammals. Unless otherwise indicated,percent similarity always refers to the comparison offull-length amino acidsequencesderivedin a fewinstancesfrom protein sequencing but more than 90% of the timededuced from the cDNA nucleotide sequence. With thesedata, several conclusions about P450 gene evolution areapparent. 1. The P450 superfamily comprises at least 10

gene families. 2. The P450 superfamily is ancient and hasexpanded via divergent evolution. 3. The ancestral P450gene, present probably more than 1.5 billion years ago,had a minimum of 22 exons. 4. Estimation of the unitevolutionary period (UEP; millions of years required for1% divergence in amino acid sequence) may be difficultdue to several instances of gene conversion betweenhomologous P450 genes. 5. Compared with all of themicrosomal P450 proteins, two mitochondrial P450 pro-teins, encoded by nuclear DNA, are more similar to theprokaryotic P450 protein.

Most drugs and combustion products are derived from

plants or are similar in chemical structure to plant metabo-lites (phytoalexins). It is proposed that four of these P450gene families (1, 11, Ill and IV) have evolved and divergedin animals due to their exposure to plant metabolites anddecayed plant products during the last one billion years.The overlapping substrate specificities of the P450enzymes are remarkable. There are numerous examples ofdrugs and other foreign chemicals that are good substratesfor the enzymes encoded by two or more P450 genefamilies gene families that have diverged so long ago asto be chromosomally non-linked.

4.2. Multidrug transporters (MDT)

It is clear that the MDT appeared early in micro-organisms living in hostile chemical environments. Wereproduce here the concluding remarks of the paper citedin section 2.1.2 [5]. MDT are present in all living cells.This review shows that the function and structure of somemultidrug transporters is conserved from bacteria tohumans. The short generation time, and the ease of mutantselection and DNA manipulation associated with micro-organisms offers great advantages over mammalian cells.Therefore, studies on multidrug transporters in microor-ganisms could bring valuable information about the gen-eral molecular mechanism and physiological role of these

transport systems, and their implications in human dis-ease. Much more, appreciation of the mechanism of suchtransporters is crucial for the development of effective newdrugs able to suppress multidrug resistance in clinicalsettings.

In a recent paper concerning the genome sequence ofPseudomonas aeruginosa [35], it was stated Pseudomo-nas aeruginosa is a ubiquitous environmental bacteriumthat is one of the top three causes of opportunistic humaninfections. A major factor in its prominence as a pathogenis its intrinsic resistance to antibiotics and disinfectants.Here we report the complete sequence of P.aeruginosa

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strain PA01. At 6.3 million base pairs, this is the largestbacterial genome sequenced, and the sequence providesinsights into the basis of the versatility and intrinsic drugresistance of P. aeruginosa. Consistent with its largergenome size and environmental adaptability, P.aeruginosacontains the highest proportion of regulatory genesobserved for a bacterial genome and a large number ofgenes involved in the catabolism, transport and efflux oforganic compounds as well as four potential chemotaxis

systems. We propose that the size and complexity of theP.aeruginosa genome reflect an evolutionary adaptationpermitting it to thrive in diverse environments and resistthe effects of a variety of antimicrobial substances.

4.3. Oxidant and oxidative species

We indicated in section 2.3 that cellular aerobic respi-ration taking place in the mitochondria is an efficientprocess to combat the toxicity of oxygen. In the presenceof oxygen, somemicro-organisms developed cellular aero-bic respiration. Mitochondria share many aspects withbacteria, so it was hypothesised that it was a transcript of a

primitive bacteria. This bacteria was phagocyted by alarger cell and established an endosymbiosic relationshipwith the host. A recent paper [36] illustrates well thishypothesis: We conclude that MsFtsZ-mtis likely to havebeen acquired from an endosymbiotic -proteobacteriumthat was the ancestor of the present-day mitochondrion.We suggest that in the course of evolution. the gene wastransferred from mitochondrion to nucleus and that thenuclear-encoded protein is now targeted back to the mito-chondrion to play a role in the division of the organelle.This is the first identification of a eukaryotic ftsZ whoseprotein seems to be specifically targeted to the mitochon-

drion, and which may thus be related to the earliestmitochondrial division genes.

4.4. Mutagenicity and DNA repair

Life appeared and developed in a bath of ionisingradiation, which was more intense a billion years ago thannow. The micro-organisms living at that time were underthis flux. Consequently many mutations of these micro-organisms certainly occurred. But also at this time, bacte-ria developed DNA repair systems, which are found nowfrom bacteria to humans.

At a recent symposium [37], it was stressed that repair

processes act according to a precise sequence of reactionsin response to DNA lesions and such a programmedsequence is exquisitely regulated in bacteria. In humans,the example of xeroderma pigmentosum is a good illustra-tion of the importance of the reparation systems in bothbacteria and humans.

To summarise, it must be stressed that the mechanismsprotecting living organisms against chemicals are very old.They took place in the first living organisms, bacteria, andwere later transmitted to other flourishing organisms onthe earth. This original chemo-defence system, whichbegan with bacteria, was certainly developed before the

immune defence system, which came after the appear-ance of eukaryotic and multi-cellular organisms. Perhapsit is for this reason that the immune defence system seemsmore integrated and more complex than the chemo-defence system. Consequently, it is clear that it is notcytochrome P 450s that are modelled on immunoglobu-lins but rather the contrary.

Thus, this section on evolution supports and confirmsthe existence of a chemo-defence system.

5. Some consequences

5.1. Low doses/high doses

As the system is saturable, it functions correctly for lowdoses. It might be less effective for medium doses and evenless so for high doses. Unfortunately in toxicologicalexperiments, the effects of low doses are the black box ofthis area. Until now, it has been very difficult to explore themechanism of the effects of low levels of toxic agents.

Today, the estimation of risk for humans is generallybased on the effects observed at medium doses and high

doses. From these results, an extrapolation is made, takinginto account what we call safety factors. This is a veryconservative approach, often scientifically false, whichtotally ignores the chemo-defence system. Now, with theapplication of the precautionary principle, we are movingmore and more towards analytical tolerances, ignoring thetoxicological significance of these values, for syntheticchemicals such as pesticides, veterinary drugs and foodcontaminants. By doing this we are creating fear in thepublic, increasing chemo-phobia and spending a greatdeal of money for the control of problems that are far morea consequence of psychological poisons than real poi-sons. This was very recently the case of dioxins detected in

chickens in France and Belgium, which had a consider-able economic impact for nothing.

Research efforts must be made in toxicology to explorethe black box of low dose effects. We now dispose ofvaluable analytical techniques with limits of detection ofng or pg/kg and also of suitably adapted tools in molecularbiology and other fields (e.g. gene identification andsequences, their expression, computed modellising struc-tures, 2D electrophoresis, etc.) to be able to investigate indepth the mechanisms of action of toxic agents.

5.2. Hormesis concept

Hormesis can be defined as the adaptive effects of lowlevels of toxic agents. Although the situation was firstdescribed for radiation [38, 39], and also for chemicalagents [40, 41] recent papers are now demonstrating thevalidity of this concept [42] and its applications.

However, until now, this new concept, which supportsthe fact that toxic agents that are detrimental to human andanimal health above certain threshold levels may inducepositive effects at a dose that is significantly lower than theclassical no effect level, has not been wholly accepted bya large part of the scientific community and the regulatoryagencies.

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The existence of a chemo-defence system certainlyprovides a valid explanation of the hormesis concept. Theinduction of detoxifying enzymes and other proteins bylow doses would support the data that imply a hormesisresponse.

However, it is not surprising that the national authoritiesdo not take this concept into account in assessing the risks,at least in terms of food contaminants. Risk assessmentsmust determine whether and how to incorporate this

information into public health decisions in a mannerconsistent with regulations, public safety and public trust.Nevertheless, in the future a change in policy will cer-tainly take place to incorporate this new scientific concept[43].

5.3. Anti-carcinogenic effects of fruits and vegetables

Ames has reviewed the role of dietary factors that areanti-tumorigenic and thus included fruits and vegetables,folic acids, various antioxidants, some vitamins and tracesof metals [44]. Recently, the following topics were dis-cussed [45]: a) the anti-oestrogenic and anti-tumoric activ-ity of diindolymethane present in cruciferous vegetables,b) the chemo-preventive action of organosulphur com-pounds in allium, c) the mechanism of action and chemo-preventive activity of the isoflavonoid genistein, and d)polyphenolic compounds in green tea.

The protective effects of carotenoids [46], vitamins Cand E, etc., as antioxidants have also been put forward.

However, when the specific compounds suspected ofbeing involved in anti-carcinogenic or protective effectsare tested separately using classical toxicological tests, theresults are more often negative. In the case of high doses ofcarotenoids, these compounds are even suspected of car-

cinogenicity [47].So the available information suggests that the efficacy of

fruits and vegetables in preventing cancer is not linked toone or other specific compound. In plants but not inanimals, many chemical structures are suspected of toxic-ity. In terms of natural pesticides, for example, which arenot the only chemical products in vegetables, it has beenestimated that the human diet contains roughly 5 000 to10 000 different natural pesticides and their breakdownproducts [48].

So why is a plant diet in fact effective against cancers?Probably because it brings together many different chemi-cal structures that are more or less toxic. These complexchemical mixtures ingested at low doses may stimulate thechemo-defence system and enhance the resistance of thehost.

Remark: This concept of the chemo-defence systemcould help change attitudes in our society, which currentlyconsiders chemicals to be poisonous, dangerous anddestructive to humans and the environment [49].

6. Conclusion

It appears that a chemo-defence system exists in living

organisms in the same way as the immune defence system.In terms of evolution, the chemo-defence system mostlikely came into existence before the latter. The chemo-defence system seems to be the ancestor of the immunedefence system, which was developed when multi-cellularorganisms confronted the problem of pathogens. Thus, it isnot surprising that the two systems have many commonfeatures and complementary roles to fight against chemi-cals and pathogens. The system can be stimulated by allthe chemicals present in our environment, whether natu-ral or synthetic. There is abundant proof that we canincrease the performance of the muscles or cardio-vascularsystem by exercise and sport and of the brain by intellec-

tual effort. It is even better known that a physiological orbiochemical system, which is not stimulated, progres-sively loses its properties. So, why should it not be possiblefor an organism to stimulate its defence against the chemi-cals found in our environment?

The aim of this paper is not to report original data, but tosupport, by the introduction of the chemo-defence systemconcept, the idea that living organisms are well armed tofight against chemicals, at least at low or medium doses.

Certain details in this conceptual paper may be criti-cised and some basic aspects may be controversial. This isa good thing, because it will open up a new area ofdiscussion and should initiate new scientific investiga-tions: for example, into the role of the nucleus as the safebox of the cells in preventing aggression against DNA, onthe substrate specificity of cytochrome P 450s, on theeffects of low doses more specifically on the validity of theLNT hypothesis, on the concept of hormesis and, last butnot least, on the protective mechanisms of the effects offruits and vegetables in our diet.

Finally, it must be stressed that life is very well protectedagainst chemicals by its chemo-defence system, whichappeared very early with the first living organisms on theearth.

Acknowledgements. I am extremely grateful to Profes-

sor M. Tubiana and Professor R. Monier for criticallyreviewing this manuscript and for their suggestions andadvice.

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