Fyziologie působení farmak a toxických látek Přednáška č.5 Endokrinní disrupce u obratlovců II. Modulace funkcí RAR/RXR, TR a PPAR - deregulace vývoje organismu, modulace endokrinních signálu a karcinogenní účinky; Efekty spojené s deregulací hladiny retinoidů: • Funkce RA; • Vznik končetin; • Vývoj nervové soustavy; • Vývojové abnormality obojživelníků; • Narušení hladin vitamínu A; o ^^ 3,4-didehydro-RA ^-^ 3,4~didehydro-retinol / 4 18 retinol 14-hydroxy-reíro-retingl 0-C-fCH2)7' o i^Yvvv\Atí ^_ ^-"^ retina! + 1S-OH-RA OH Struktura a syntéza kyseliny retinové die^'^-RCHO-CRBP<"^ROH-c retiny i pa Imitate |l-carútône 9-cŕs-RA target cell FIG. 3. Absorption, distribution, and metabolism of naturally occurring retinoids. on 9,13-di-tľŕS-RA [■'[(i. 2. Structures of naturally occurring retinoids. A mm m ■---------■ A j B C D E F Transcriptional activation domains (AF-1) DNA binding domain (RARE) Hinge region Ligand-binding Heierodimerization Transcription activation (AF-2) ■ N-Cor?SMRT binding domains ? B 5' tí AGGTCA - AGGTCA- -AGGTCA-AGGTCA (WW RARE Early response genes fc Secondary response genes y t Gene product (e.g. transcription factors STATs, RARs, c/EBP, etc.) Inhibition of cell growth, Induction of differentiation, apoptosis Fig. 1 - Structure and functions of retinoid receptors. A) Schematic representation of retinoid receptor protein depicting various functional domains. B) A molecular model for retinoid action. The Iiganded RAR forms heterodimer with RXR, binds to specific regulatory sequences (RARE) in the promoter region of target genes. Transactivation of such early response genes is a primary event of retinoid action. In addition to this, the products of early response genes can activate the transcription of secondary genes. Transactivation of these genes therefore represents secondary action of retinoids since their transcription requires protein synthesis. This cascade of gene events leads to secondary and tertiary events that eventually produce a phenotype that is characteristic of retinoid action. • Abnormalities caused by exogenous agents (certain chemicals or viruses, radiation, or hyperthermia) are called developmental disruptions. The agents responsible for these disruptions are called teratogens. Most teratogens produce their effects only during certain critical periods of development. The most critical time for any organ is when it is growing and forming its structures. Different organs have different critical periods, but the time from period from day 15 through day 60 of gestation is critical for many human organs. • Retinoic acid is important in forming the anterior-posterior axis of the mammalian embryo and also in forming the limbs. In these instances, retinoic acid is secreted from discrete cells and works in a small area. However, if retinoic acid is present in large amounts, cells that normally would not receive such high concentrations of this molecule will respond to it. Inside the developing embryo, vitamin A and 13-cis-retinoic acid become isomerized to the developmentally active forms of retinoic acid, all-/ra/75-retinoic acid and 9-c/5-retinoic acid. Some of the Hox genes have retinoic acid response elements in their promoters. • In the early 1980s, the drug Accutane® (the trade name for isoretinoin, or 13-c/5-retinoic acid) was introduced as a treatment for severe acne. Women who took this drug during pregnancy had an increased number of spontaneous abortions and children born with a range of birth defects. RA reguluje vznik a vývoj končetin There are discrete positions where limb fields are generated. Researchers have precisely localized the limb fields of many vertebrate species. Interestingly, in all land vertebrates, there are only four limb buds per embryo, and they are always opposite each other with respect to the midline. Although the limbs of different vertebrates differ with respect to which somite level they arise from, their position is constant with respect to the level of Hox gene expression along the anterior-posterior axis. For instance, in fishes (in which the pectoral and pelvic fins correspond to the anterior and posterior limbs, respectively), amphibians, birds, and mammals, the forelimb buds are found at the most anterior expression region of Hoxc-6, the position of the first thoracic vertebra. Retinoic acid appears to be critical for the initiation of limb bud outgrowth, since blocking the synthesis of retinoic acid with certain drugs prevents limb bud initiation, suggested that a gradient of retinoic acid along the anterior-posterior axis might activate certain homeotic genes in particular cells and thereby specify them to become included in the limb field. Legs regenerating from retinoic acid-treated tadpole tail. (A) The tail stump of a balloon frog tadpole treated with retinoic acid after amputation will form limbs from the amputation site. (B) Normal tail regeneration in a Rana temporäriatadpole 4 weeks after amputation. A small neural tube can be seen above a large notochord, and the muscles are arranged in packets. No cartilage or bone is present. (C) A retinoic acid-treated tadpole tail makes limb buds (arrows) as well as pelvic cartilage and bone. The cartilaginous rudiment of the femur can be seen in the right limb bud. pottftior Dfoaaphila Hox complex a lab pb |bh/ Dfd Ser (Hz) Antp Ubx abó-A AbtS-B -G-----D- T í I t M «íl ľ f JJínif? ítnv? UmJ Uínif. Hox J Hotó HcuŕJÍ Hoií Hourä HoxS (central) ancestral __| I I I I I I I I L Ho* complex ^ n-*r X mammalian Hcuf 7 (posterior) n----- .•..■■•-. JT i. -i.. ■",: >4S 47 A3 A10 Alt Al 3 BI 32 83 Bi BS BS B? BS rV HOXB B13 -O Ci C5 CS CS C9 CIO C11 C}2 A13 HOXC CH HOXD 03 D4 -D-D- DS OS D10 DM D12 DÍ3 filndbrain and spinal cord mBsodwrn posterior The Hox complex of an insect and the Hox complexes of a mammal compared and related to body regions. Hoxb-4 dorsal view aide víew dorsal view k de view Expression domains of Hox genes in a mouse. The photographs show whole embryos displaying the expression domains of two genes of the HoxB complex (blue stain). These domains can be revealed by in situ hybridization or, as in these examples, by constructing transgenic mice containing the control sequence of a Hox gene coupled to a LacZreporter gene, whose product is detected histochemically. Each gene is expressed in a long expanse of tissue with a sharply defined anterior limit. The earlier the position of the gene in its chromosomal complex, the more anterior the anatomical limit of its expression. Thus, with minor exceptions, the anatomical domains of the successive genes form a nested set, ordered according to the ordering of the genes in the chromosomal complex. 04 Teratogenesis in frogs. (A) Wild green frog {Rana clamifans) with an eye deformity, collected in New Hampshire in 1999 by K. Babbitt. (B) Xenopustadpole with eye deformities caused by incubating newly fertilized eggs in water containing methoprenic acid, a by-product of methoprene. (C) One of several pathways by which methoprene can decay into teratogenic compounds such as methoprenic acid. (D) Ar\ isomer of retinoic acid showing the structural similarities to methoprenic acid. RA reguh je vývoj CNS Top panel: At left, retinoic acid activates gene expression in a subset of cells in the normal developing forebrain of a midgestation mouse embryo (blue areas indicate ß-galactosidase reaction product, an indicator of gene expression in this experiment); at right, after maternal ingestion of a small quantity of retinoic acid (0.00025 mg/g of maternal weight), gene expression is ectopically activated throughout the forebrain. Bottom panel: At left, the brain of a normal mouse at term; at right, the grossly abnormal brain of a mouse whose mother ingested this same amount of retinoic acid at mid-gestation. Nťuťal ŕiest Nťura] h.iťe Sonk hedgehog, netirtoir arid, noggin,-. CniscH ill., i 1995 Beclcmen el al., 11997) Brouweret al., (1989) D<- swan .-i «I.. (low i. ioo.it Jenssen et al., (1995) Hall h al.,(1998) thyroid glan d lliMiiid abscesses thyroid adenoma plasma I retinol l IT,". TT3f pl.ism.i ■I retinol l TT,, IT/-, TTg plasma I retinol I tt4, ny plasma i TT,. IT, plasma ■lTTg:TT4 TABLE 3. Alterations in thyroid inland morphology and retinoid levels in lish associated with exposure to poMialogenared aromatic hydrocarbons and polynuclear aromatic hulrm-irbmiN. Species Study location \,.,..,.,i.,l contaminants 1 ÍS3UI v.....| -1- -1 TliMniiln liniiiil changes References Salmon species Great hikes unknown Factor thyroid gland thyroid hypertrophy, hy- Sonstegard el al.. Í197ÍÍ) (OtlCůHuflduU sp.t perplasia White sucker Montreal. Qik-Ikv coplanar liver I retinol Spear el ill. \'.)'.)2 Catostomus commeno ,„ ľ( ľ- ■ i retiny] palmitate Hrancliauilel al.,(l995) Lake sturgeon Montreal. (,)iu-Imi- ľi ľ, intestine J retinyl palmitate Ndaviliaiiini.l.il. 1001 (Aclpenser fiilvescens 1 dehydroretiny] palmitate T l\.V' metabolism Like sturgeon Si. Lawrence River, coplanar PCBs liver Doyon el al., (1999) (Aclpenser fiilvescens) Quebec I retinoids Brown bullheads Great Ijiki-s PAHŕ liver ■I retinyl palmitate Arcand-Hoyetal., (1999) Aiiwiunis nebidosus) i (k'hulimiviiml 56 �35284 52 52 418 487746 70 5985 TABLE 1. Alterations in thyroid gland morphology, aromatic hydrocarbons. thyroid hormones und retinoid IiavIs in free-ranging avian species associated with exposure to polyhalogenated Species Study location Associated contaminants s.....pled Thrvoid/relinoid changes References 1 [erring gulls (Lotus argpntatus I 1 [erring nulls (Larta argpntatus) 1 U'ľľini; pills (Lan« argeiiíarus) 1 [erring ^iills (Larta argpntatus) Great hlín* lierons Anas herodlas) < cormorants (Phalacrocorax carba) 1 [erring pills Joints argpntatus) Caspian terns (Sterna caspia) Common l en is Strnul liíritliiln) 1 [iTľiliH pills •.hints argentatus) Tree swallowsP (Tachyclneta bicolor Great l-akt-s I'lIAlhv" thyroid T thyroid mass thyroid hyperplasia Mocda et al., (lySfi) Great Likes I'll Úl- liver . retiny] palmitate Government of Canada (1991 Great Lakes 2.:l.7.S-Tcnnl' liver i IVlillol i retinyl palmitate Spear et al.. {ii)HI\ [992 Lakes Huron Erie, •1.^.7.H-vam ■ :gg -i ■ ill T retlnol: retinyl palmitate Spearetal.,(1990) Ontario iPCDDs + PCDFŕ dioxin rEQsd SI. Lawrence River, iPCBs 111.) + US egg yolk i retinyl palmitate Boilyetal., [994 Quebec IPClis US + TEQsf 1 l\ Netherlands PCBsS . rgg ) ■ 'Ik i IT.,1' van den Berg ei al.. l'< 1)1). liver T EROD1 ■ 1 !)!>(' PCDFs plasma Greal Lakes ]'( IS,' plasma* ■l ľi-linol Grasmanel al.. UiKXi* Belgi..... Nillui lands Ot-al Ijikes Great 1-akes St Lawrence River liasin mono-cirllio l'< Uis PCDDs l'<:m-s t*j5; Volk plasma liver II lis l>l>l -.li. 1.1..........\ liv.-i All-inducing chemiealsfl livei i retinyl palmitate' i TLÝ". TT.,". VY., T plasma retino] li> \olk s.ii-retiny] palmitate i retiny] palmitate I retino! T EROD Mnrkftiil.. (lööß) Foxel al.. (1998) Bishopel al. (1999) 78 PHAHs modulují hladiny retinoidu - mobilizace zásob vitaminu A v játrech c o o o c 300 250 200 - 150 S 100 a. ťD X 50 -:■::-! iii :■:■■:-:■-:■:? ¥:>»-:*' u*d r-'iJiiU mm mí mm všX-šy-i ■-"/í I I k0 WM 111 ■:-:-:-:*:-;«■ :■:--:•: «í: £221, Dams Fetus Maid Female Mate Female GD20 PND21 PND90 FIG. 3. Hepatic retinol concentrations, expressed as percentage of control values, mean ± SEM, from dams, their fetuses (N = 6), male and female neonates {N = 8-10), and adult offspring (JV = 10) following maternal exposure to 0 Q, 5 M, or 25 O mg Aroclor l254/kg on Days 10-16 of gestation. ^Indicates a significant difference from controls, p < 0.05. GD20, Gestation Day 20; PND21, Postnatal Day 21; PND90, Postnatal Day 90, BPA moduluje hladiny retinoidních receptoru v průběhu embryogenéze - myši rr^S OH p-Alky I phenols OCH3 OCH3 p.p-Methoxychlor OH i OH 1 rS n U ĺ jŕ H3C ~~ C ~~ CH j f C "1 */*Y K V CI Ôh Hydroxy-PCB Bisphenol-A ^COjCHjCeHj KC02(CHľ}3CH3 Diethylstilaestrol Benzyl bulyIphthalate Figure 2 Structures of some xc n oestrogen s. Funkce thyroidních hormonů v ontogenezí a vliv organických polutantů: • Funkce thyroidních hormonů v metamorfóze • Funkce thyroidních hormonů ve vývoji nervové soustavy; Hypotéza - environmentálni polutanty jako kauzální faktor neurologických poruch (autismus, poruchy učení, hyperaktivita, nádorová onemocnění, juvenilní formy diabetes); • Toxické látky narušující thyroidní regulace; T3 a T4 mají zásadní význam pro iniciaci metamorfózy obojživelníků Temp Light Nutr. etc, ^ J&&, S ---------Hypothalamus TRH \ CRF Pítuary PROLACTIN + V e F e I b a c k O m Embryo Aduti Fig, I. Schematic representation of the hormonal regulation of amphibian metamorphosis. In response to environmental cues, the dormant thyroid gland of the tadpole is activated to produce the thyroid hormones T4 and T3 by the hypothalamic and pituitary hormones TR.F, C'Rľ and TSll. "1'hyroid hormone (Til) is obligatorily required to initiate and maintain the metamorphosis, its action being potentiated by glucocorticoid hormone and retarded by prolactin. Nuti\, nutritional factors; TRH, thyrotrophin-releasing hormone; CR1\ cortcotrophin-releasing factor; TSE J, thyoid -stimulating hormone; T4, l-thyroxine; T3, triíodo-L-thyroníne; GC, glucocorticoid hormone. Table 1 Diversity of morphological and biochemicai responses ro thyroid hormone during amphibian metamorphosis Tissue Response Morphological Biochemical Brain Liver Eye Skin Limb bud, king Tail, gills Intestine, pancreas Immune system Muscle Restructuring; axon guidance and growth; cell turnover Functional differentiation; restructuring Repositioning; new retinal neurones; altered lens Restructuring; keratinisation; granular gland formation De novo morphogenesis of bone, skin, muscle, nerve, etc. Total tissue regression and removal Major remodelling of tissues Redistribution of immune cell populations Growth, differentiation, apoptosis Cell division; apoptosis; protein synthesis Induction of albumin and urea cycle enzymes; larval adult haemoglobin switch Visual pigment switch; induction of fi- crystal lín Induction of collagen, 63 kDa keratin, magainin Cell proliferation; gene expression Programmed cell death; induction of lytic enzymes New structural and functional constituents Aquisition of new immunocompetence Induction of myosin heavy chain Conception 12 weeks 24 weeks Birth Maternal thyroid hormone _________—-----------—— ^~~~ Thyroid receptors flu, p p Irs -------------------—------------------- ' ___-------——— ~~ Fetal thyroid hormone 1 D WEEKS ----—— 4 we e ks Hypothalamus G weeks Cochlea 18 weeks 7 weeks Hippocampus 5 weeks Synaptogenesis, myelinogenesis., gliogenesis 7 weeks Sexual differentiations—urogenital Figure 1. Role of thyroid hormones in fetal neurologic development in relation to timing of several landmark stages of development. Figure adapted from Howdeshell (2002). Although it has been known for a century that hypothyroidism leads to retardation and other serious developmental effects, the role of thyroid hormones in brain development is still not completely understood. It is also accepted that thyroid hormones transferred from the mother to the embryo and fetus are critical for normal brain development, even though the thyroid gland of a fetus starts producing thyroid hormones at about 10 weeks. We now recognize that only a slight difference in the concentration of thyroid hormones during pregnancy can lead to significant changes in intelligence in children. Možné mechanismy disrupce funkce thyroidních hormonů • Inhibition of active transport of inorganic iodide into the follicular cell • Interference with the sodium/iodide transporter system • Inhibition of thyroid peroxidases to convert inorganic iodide into organic iodide to couple iodinated tyrosyl moieties into thyroid hormone • Damage to follicular cells • Inhibition or enhancement of thyroid hormone release into the blood • Inhibition or activation of the conversion of T4 to T3 by 5'-monodeiodinase at various sites in the body, for example, the fetal brain • Enhancement or interference of the metabolism and excretion of thyroid hormone by liver uridine diphosphate • Interference with transport of thyroid hormones • Vitamin A (retinol) disturbances • Blocking of or interfering with thyroid receptors 22 Mechanisms of Action of Thyroid-Disrupting Chemicals The complexity of the development of both the neurologic and thyroid systems offers numerous opportunities for chemicals to interfere as the systems develop, mature, and function. Briefly, there are chemicals that interfere with iodine uptake (the herbicides 2,4-D and mancozeb, several PCB congeners, and thiocyanates) and peroxidation at the molecular level (the herbicides aminotriazole and thioureas, the insecticides endosulfan and malathion, and PCBs). They also interfere with the protein transporter that provides a pathway for iodine to enter the cell (military and aerospace chemicals, Perchlorates). Certain antagonists (PCBs, the herbicides aminotriazole and dimethoate, and the insecticide fenvalerate) prevent the release of thyroid hormone from the cell and inhibit conversion of T4 to triiodothyronine (T3). Various chemicals enhance excessive excretion of thyroid hormones, some through activation of the cytochrome P450 system (dioxin, hexachlorobenzene, and fenvalerate). Some PCBs, phthalates, and other widely used chemicals compete for sites on the thyroid transport proteins that deliver thyroid hormones throughout the body. New research focuses on the role of chemicals as they interfere with vitamin A (retinols). retinols, a process essential for thyroid hormone expression. Hydroxylované PCB During normal enzyme detoxification of PCBs in the maternal liver, certain PCB congeners are hydroxylated. This metabolic step enhances the binding affinity of the hydroxylated PCBs to TTR. Through their high-affinity binding the hydroxylated ci congeners displace essential f T4 that must get to the fetal brain to be converted to fT3. Hydroxylated PCBs also interfere with the normal excretion of thyroid hormones by inhibiting their sulfation. PCB hydroxylates also have estrogenic and antithyroid properties. Thyroidní disrupce u volně žijících obratlovců: Obojživelníci Gut leb and co-workers did a series of exposure studies with Xenopus laevis ar\d Rana femporaria. They found increased incidence of mortality in tadpoles weeks after they ceased dosing the animals. Over an 80-day period, 47.5% of the tadpoles died. The X. laevisexposed to 7.7 pM and 0.64 nM PCB 126 exhibited swimming disorders prior to death. Both increased mortality and reduced T4 concentrations occurred in a dose- response manner in X. laevis. Severe eye and tail malformations increased in the froglets in a dose-response manner after approximately 60-68 days. Ptáci Thyroid hormones in birds have been investigated for their role in migration and courtship. Preventing migrating species from breeding out of season is especially critical for their survival. From the 1950s through to the 1970s, fish-eating birds in the Great Lakes were experiencing very poor reproductive success. Keith suggested that the high embryo mortality and low chick survival in herring gulls nesting in upper Green Bay in the mid 1960s was both the result of a) the effects of the chemical residues from the mother on the embryo and b) the effects of the adult's contamination on its parental behavior. Ryby Migration of salmonids is linked with THs effecting a sequence of behaviors. In the laboratory, increases in T4 led to less display of aggressive behavior such as territoriality. Elevated concentrations of both T3 and T4 reduced the fishes' preference for shade to more open areas (phototaxis). T3 treatment caused the fish to swim with the current rather than against the flow (rheotaxis). Savci PCBs and dioxins have been shown to alter thyroid function in rodents by multiple mechanisms, including direct toxic effects on the thyroid gland, induction of thyroid hormone metabolism via the UDP-glucuronyl transferases, and interactions with thyroid hormone plasma transport proteins, particularly transthyretin. A number of investigators have evaluated the effects of maternal PCB exposure on thyroid function of rat pups. Pup serum thyroxine (T4) levels are markedly reduced by PCB or dioxin exposure, but the levels of the active form of the hormone, triiodothyronine (T3), are generally unchanged, or only slightly reduced. A relationship between exposure to dioxins and PCBs and alterations in thyroid hormones has also been reported in human infants. Infants exposed to higher levels of PCBs and dioxins had lower free T4 levels and higher thyroid-stimulating hormone levels. 22 PPAR • Deregulace PPAR a reprodukce • PPAR a karcinogenita Ligand-índependent activation domain (AFI) I DBD DNA Binding Domain (2 zinc fingers) Ligand Li gand-dependent activation domain (AF-2) LBD-Dim. Ligand Binding and Dimerization Domains 9-ŕľiV jietinoic acid AGGTCANAGGTCAa TCCAGTNTCCAGT ■ r CiglitEizone 'XX NU ľioĽUla/i>RĽ ch Troglitaironc tr-r^a 1 Rosiglitazone FIG. 1. Genera] structure and mechanism of action of PPAR». PPAR isoforms share a common domain structure and molecular mechanism of action. Membrane phospholipids I Plrospho lipase A2 Arachidonic acid Linoteic acid OxLDL Cycloxygenase Lipoxygenases Lipoxygenases 15-deqxy PG-J2 LTB4, 8-HETE, 1S-HETE 9-HODE, 13-HODE Glitazones PPAR COjH linoleic acid 15-deo!cy-Al2l4-PGJ2 'C02H HO 9-HODE ,CO;>H eicosapentaenoie acid /-COjH h y— 0H 8(S)-HETE 13-HODE Mo no ethyl (M EP) Monobutyl(MBP) ľvlonopentyl i'MPP'i O ^ChL Mo no methyl í M M Pi O H o n o h exy I í M H Pi Mono p no pyl (MPrP) Monü-(2-ethylhexyl)(MEHP) Figure 1. Structurally related phthalate monoesters. Diesters of o-phthalic acid are quickly metabolized in vivo to their active metabolites, the monesters. The length and structure of the side chain is important for toxicity. Ftaláty jako Ugandy PPAR - efekty na samčí a samicí reprodukční systém TABLE 1 A Summary o ľ Effects o ť in Utero Exposure to Phthu la tes on the PPAR: RXR ER: E R Developing Mule Reproductive Tract Ph t h li kites-» 0 oo Endpoinl measured" DBP DEHP BBP DIN P \/ , Up- (if down- Testis regulation ... Weight i i i i i i — 1 1 1 1 Sperm number 1 1 | | Degeneration/atrophy of i i i ERE Regulated Gene seminiferous Lubules Leydigcell hyperplasia/aggregates i i i D PhťhaLates Leydigcell adenoma i + Cryptorchidism i i i 1 RXR Sex org Lin s Epididymis: ... wl, agenesis/malformed i i i _ PPAR 0-0-0 \ TRorRAE Penis: delayed/incomplete preputial i i i - / separation, hypospadias, ... wl of glans i i i ^ / Down- Prostate: . wt, agenesis Regulation Seminal vesicle: ... wl 1 + l l 1 1 i i 1 T J £ U 1 J ■ 1 1 | | Vasdeierens: , wl, mal lor med/ agenesis Miscellaneous TREot RARE Re WADH + O I R —CHj—C—CH;—C —SCoA O Ihiolasc HSCííA Ü I R —CHj —C —SCoA + H,C —C —SCoA AíetylCoA Acyl CůA ahartariud by two tarbůn at&ma ■H,O s"? Oxidation of fatty acids by peroxisomes. Peroxisomes degrade fatty acids with more than 12 carbon atoms by a series of reactions similar to those used by liver mitochondria. In peroxisomes, however, the electrons and protons transferred to FAD and NAD+ during the oxidation reactions are subsequently transferred to oxygen, forming H202 >* ' , 1 Éi^ r4 ■ »4k . m PPARcc a karcinogeneze S. Bosgra et a L / Toxicology 206 (2005) 309-323 PP-----^(ŕPÄíSí)—^^P^^g^)^(gxR) TGACCTnTGACCT PPRE i gene regulation -* peroxisome proliferation -+. ß-oxidation (ACO, BFE, THL)' • co-oxidation (CYP4A) cell proliferation Ui apoptosis | '} ■*- H2O2 initiated hepatocytes i liver tumours Peroxisome proliferation ■ Liver growth - hypertrophy - hyperplasia ■ Induction of liver enzymes - peroxisomal enzymes (peroxisome proliferation) - P450 - the CYP4 genes ■ Proliferation of the Endoplasmic Reticulum and peroxisomes ■ Hypolipidaemia Oxidative Theory Fatty acids increase in peroxisomal ß-oxidation without catalase (<2 fold) Increase H202^^^^^^ genotoxic DNA damage Cancer Úloha Kupfferových buněk Kupffer cells Hepatocytes (RX PPRE i gene regulation ■+ peroxisome proliferation ■* ^oxidation (ACO, BFE, THLJ' (it-oxidation (CYP4 A) cell proliferation t **■ apoptcsis I ■* HřO? } initiated hepatocytes liver tumours Species Differences and Human Risk Assessment ■ PPARa exists m mouse, rat, guinea pig and human ■ Low hepatic levels \r\ human and g-pig ■ Human liver - No peroxisome proliferation - No induction of liver growth ■ If PPARs cause cancer m rats, do they cause cancer \r\ humans? Therefore, no risk of cancer???