{"paper_id":"85611151-3f7e-4625-bd7d-c690ee4de73a","body_text":"Folia Biologica (Praha) 61, 26-32 (2015)\nOriginal Article\nAntioxidant Status in Blood of Gynaecological Patients: \nInfluence of Diagnosis and Reproductive Factors\n(antioxidant\tenzymes\t/\treproductive\tfactors\t/\tgynaecological\tpatients)\nS.\tPEJIć,\t V .\tSTOJILJKOVIć,\tA.\tTODOROVIć,\tL.\tGA VRILOVIć,\tN.\tPOPOVIć,\t\nI.\tPA VLOVIć,\tS.\tB.\tPAJOVIć\nLaboratory\tof\tMolecular\tBiology\tand\tEndocrinology,\t“Vinča”\tInstitute\tof\tNuclear\tSciences,\tUniversity\t\nof Belgrade, Belgrade, Serbia\nAbstract. Cancer of the reproductive tract is an im-\nportant cause of morbidity and mortality among \nwomen worldwide. In this study we evaluated the in-\nfluence of diagnostic categories, age and reproduc-\ntive factors on antioxidant enzymes and lipid hy-\ndroperoxides in the blood of gynaecological patients \ndiagnosed with endometrial polyp, myoma, hyper -\nplasia simplex, hyperplasia complex and endometri-\nal adenocarcinoma. Multivariate regression analysis \nwas used to assess the association of diagnosis, age, \nparity, abortions and abnormal uterine bleeding \nwith the examined parameters. Diagnosis provided \nthe best predictive model for superoxide dismutase, \ncatalase and glutathione peroxidase activities, and \nalso for the lipid hydroperoxide level. Abortions fit-\nted the best predictive model for superoxide dis-\nmutase activity. A significant correlation was also \nfound between the predictor variables themselves. \nThis study showed that reproductive and other fac-\ntors may be associated, at least partially, with anti-\noxidant capacity and ability to defend against the \noxidative damage in gynaecological patients with \nvarious diagnoses.\nReceived\tAugust\t8,\t2014.\tAccepted\tSeptember\t29,\t2014.\nThis\twork\twas\tfinancially\tsupported\tby\tthe\tMinistry\tof\tEduca-\ntion,\tScience\tand\tTechnological\tDevelopment\t(Grants\t41027\tand\t\n41022).\nCorresponding\t author:\t Snežana\t Pejić,\t “Vinča”\tInstitute\t of\t Nu-\nclear\t Sciences,\t P .O.\tBox\t 522,\t 11001\t Belgrade,\t Serbia.\t Phone/\nFax:\t(+381)\t11\t6455\t561;\te-mail:\tsnezana@vinca.rs\nAbbreviations:\tACE\t –\t adenocarcinoma\t endometrii,\tAUB\t –\t ab-\nnormal\tuterine\tbleeding,\tAO\t–\tantioxidant,\tCA T\t–\tcatalase,\tCH\t–\t\nhyperplasia\tcomplex\tendometrii,\tEC\t–\tendometrial\tcancer,\tEH\t–\t\nendometrial\t hyperplasia,\t GPx\t –\t glutathione\t peroxidase,\t LOOH\t\n–\tlipid\thydroperoxide,\tPE\t–\tpolypus\tendometrii,\tROS\t–\treactive\t\noxygen\tspecies,\tSH\t–\thyperplasia\tsimplex\tendometrii,\tSOD\t–\tsu-\nperoxide\tdismutase,\tUM\t–\tuterus\tmyomatosus.\nIntroduction\nCancer of the reproductive tract is an important cause \nof morbidity and mortality among women worldwide, \nwith\tendometrial\tcancer\t(EC)\tas\tthe\tfourth\tmost\tcom-\nmon cancer among women in developed countries. So \nfar, it has been known that several factors may partici -\npate in pathogenesis of various gynaecological diseases. \nObesity is considered to be strongly associated with the \nrisk of developing endometrial cancer and it depends on \nthe\tobesity\tdegree,\texpressed\ton\tthe\trelative\t2–10\tscale\t\n(Olson\tet\tal.,\t1995;\tPurdie,\t2003).\tIt\tis\talso\tbelieved\tthat\t\nthe association between high body weight and EC is \nmore pronounced in postmenopausal women (Trentham-\nDietz\tet\tal.,\t2006);\thowever,\tabout\t5–30\t%\tof\twomen\t\nare pre- or perimenopausal at the time of diagnosis \n(Y amazawa\tet\tal.,\t2000;\tSoliman\tet\tal.,\t2005).\t\nDiagnoses before the age of 50 has been linked to a \nnumber of risk factors including age of menarche, pari-\nty,\tfailure\tto\tovulate\tand\ttamoxifen\tuse\t(Straughn\tand\t\nPartridge\t 2009;\t Zucchetto\t et\t al.,\t 2009).\t Other\t factors\t\nthat may contribute to increased EC risk are age, late \nmenopause,\thyperandrogenaemia\t(Cline,\t2004).\tFactors\t\naffecting\t EC\t and\t endometrial\t hyperplasia\t (EH)\t are\t\nknown\tto\tbe,\tat\tleast\tin\tpart,\tsimilar\t(Ricci\tet\tal.,\t2002).\t\nHowever, women with benign gynaecological condi-\ntions\tsuch\tas\tendometriosis,\tuterine\tfibroids\t(leiomyo-\nmas)\t or\t endometrial\t polyps\t may\t also\t experience\t in-\ncreased risk of developing hyperplasia and malignancy \n(Brinton\tet\tal.,\t2005;\tSilberstein\tet\tal.,\t2006;\tRowlands\t\net\tal.,\t2011).\tStudies\tindicate\tthat\tcompleted\tor\tuncom-\npleted pregnancy may be protective against EC but not \nagainst\tEH\t(Parslov\tet\tal.,\t2000;\tPike\tet\tal.,\t2004;\tXu\tet\t\nal.,\t2004).\tAssociation\tof\thistory\tof\tabortions\twith\tthe\t\nEC risk is still unclear since both positive and inverse \nrelationships\twere\treported\t(Xu\tet\tal.,\t2004).\t\nAbnormal\tuterine\tbleeding\t(AUB)\tis\tone\tof\tthe\tmost\t\ncommon symptoms of endometrial cancer in postmeno-\npausal\twomen\t(Epstein\tand\tV alentin,\t2004),\tbut\tit\talso\t\nrepresents a risk factor for the occurrence of endome -\ntrial hyperplasia in perimenopause (Farquhar et al., \n1999).\tAbout\t10\t%\tof\twomen\twho\thave\tirregular\tbleed-\n\nV ol.\t61\t 27\ning in postmenopause will have a diagnosis of endome-\ntrial cancer, the same percentage will have hyperplasia, \n60\t%\tof\twomen\twill\tbe\tdiagnosed\twith\tuterine\tatrophy,\t\nand\t10\t%\twill\tbe\tdiagnosed\twith\tpolyps\t(Karlsson\tet\tal.,\t\n1995).\t Bleeding\t in\t postmenopausal\t women\t may\t in-\ncrease the risk of developing endometrial cancer up to \n64\ttimes\t(Gull\tet\tal.,\t2003).\nOxidative\tstress\tmay\tplay\tan\timportant\trole\tin\tindi-\nvidual risk of developing many diseases, including can-\ncers.\t Cells\t developed\t an\t enzymatic\t antioxidant\t (AO)\t\npathway\tagainst\treactive\toxygen\tspecies\t(ROS),\twhich\t\nare\tgenerated\tin\toxidative\tmetabolism.\tSuperoxide\tdis-\nmutase\t(SOD)\tcatalyses\tdismutation\tof\tsuperoxide\tani-\non (O\n2\n.–)\tto\thydrogen\tperoxide\t(H2O2),\twhich\tin\tthe\tsec-\nond\t step\t is\t converted\t to\t water\t by\t catalase\t (CA T)\tor\t\nglutathione\tperoxidase\t(GPx).\tGPx\talso\treduces\torganic\t\nperoxides\tinto\talcohols,\tusing\tglutathione\tas\thydrogen\t\ndonor\t (Halliwell,\t 2006).\t The\t activity\t of\t the\t first-\tand\t\nsecond-step AO enzymes has to be balanced to prevent \npotential\toxidative\tdamage\tin\tcells.\tV ariations\tin\tAO\tca-\npacity\tmay\tinfluence\tindividual\tsusceptibility\tto\tpatho-\nlogical processes associated with the deleterious effects \nof\toxidative\treactions\t(Dalle-Donne\tet\tal.,\t2006;\tPagliu-\nso\tet\tal.,\t2008).\t\nIn our previous research, we have shown that the an-\ntioxidant\tstatus\tis\taltered\tin\tthe\tblood\tand\tendometrium\t\nof women with endometrial hyperplasia and adenocarci-\nnoma in comparison to those with polyps or leiomyo -\nmas.\tThe\t specific\tchanges\t were\t related\t to\t the\t enzyme\t\ntype\t and\t diagnosis;\t however,\tthe\t reduction\t in\t antioxi-\ndants\t and\t elevation\t of\t the\t lipid\t hydroperoxide\t level\t\nwere\tobserved\tin\tgeneral\t(Pejić\tet\tal.,\t2006,\t2009).\tIn\t\nthis study we sought to evaluate the association of age \nand reproductive factors such as parity, abortions and \nAUB with AO enzyme activities in the blood of these \npatients. \nMaterial and Methods \nSubjects\nThe\tmaterial\tused\tin\tthis\tstudy\tconsisted\tof\t88\tblood\t\nsamples of women admitted to the Department of \nGynaecology and Obstetrics for gynaecological evalua-\ntion within routine checkups or for abnormal uterine \nbleeding (Metrorrhagia prolongata, Metrorrhagia recidi-\nvans,\t Metrorrhagia\t postmenopausi).\t The\t specimens\t\nwere taken after obtaining the informed consent. The \nstudy was conducted prospectively and it was approved \nby the Human Studies Ethics Committee of the Clinical \nCentre. The protocol was consistent with the World \nMedical Association Declaration of Helsinki (Ethical \nPrinciples for Medical Research Involving Human \nSubjects).\tOn\tthe\tbasis\tof\tdiagnosis\tand\thistological\tex-\namination, subjects were divided into the following \ngroups:\t patients\t with\t polypus\t endometrii\t (PE),\t uterus\t\nmyomatosus\t (UM),\t patients\t with\t hyperplasia\t simplex\t\nendometrii\t(SH),\thyperplasia\tcomplex\tendometrii\t(CH),\t\nor\tadenocarcinoma\tendometrii,\tstage\tI\t(ACE).\tWith\tre-\ngard to the age, parity (nulliparous, primiparous and \nmultiparous)\tand\tabortions,\tpatients\twere\talso\tclassified\t\nin appropriate categories.\nSamples\nVenous blood samples were collected into heparin -\nized tubes on the same day as endometrial biopsy and \ncentrifuged\t at\t 2500\tg for 5 min. Plasma was used for \nLOOH\tconcentration\tmeasurement.\tFor\tSOD\tassay\t(Oxis\t\nInternational,\tInc.,\tPortland,\tOR),\tthe\tpellet\twas\tresus-\npended in four packed-cell volumes of ice-cold demine-\nralized ultrapure water (MilliQ reagent grade water sys-\ntem,\tMillipore\tCorp.,\tBedford,\tMA).\tAfter\taddition\tof\t\nethanol/chloroform\t extraction\t reagent\t (62.5/37.5\t vol/\nvol)\tto\tremove\thaemoglobin\tinterference,\tsamples\twere\t\ncentrifuged\tat\t3000\tg for 10 min (Eppendorf centrifuge \n5417,\tEppendorf\tAG,\tHamburg,\tGermany).\tThe\tupper\t\naqueous\tlayer\twas\tcollected\tand\tkept\tat\t-70\t°C\tuntil\tas-\nsay.\tThe\tactivities\tof\tCA T,\tGPx\tand\tGR\twere\tmeasured\t\nin blood lysates.\nThe\t enzyme\t activities\t and\t lipid\t hydroperoxide\t\n(LOOH)\t concentration\t were\t monitored\t spectrophoto-\nmetrically (Perkin Elmer Spectrophotometer, Lambda \n25,\tPerkin\tElmer\tInstruments,\tNorwalk,\tCT).\tThe\tspe-\ncific\tenzyme\t activities\t were\t expressed\t as\t units\t (U)\t or\t\nmU\tper\tmilligram\tof\ttotal\tcell\tprotein\t(U\tor\tmU/mg\tpro-\ntein).\tLOOH\tconcentration\twas\texpressed\tas\tnmol/mg\t\nprotein. Determination of protein concentration was \nperformed\t by\t the\t method\t of\t Lowry\t et\t al.\t (1951)\t and\t\nexpressed\tas\tmg/ml.\t\nAssays \nAssay of SOD activity. Determination of SOD activ-\nity\twas\tperformed\tusing\tOxis\tBioxytech®\tSOD-525™\t\nAssay\t(Oxis\tInternational,\tInc.).\tThe\tmethod\tis\tbased\ton\t\na\tSOD-mediated\tincrease\tof\tautoxidation\tof\t5,6,6a11b-\ntetrahydro-3,9,10-tryhydroxybenzo[c]fluorene\tin\t aque-\nous\talkaline\tsolution\tto\tyield\ta\tchromophore\twith\tmaxi-\nmum\t absorbance\t at\t 525\t nm.\t The\t SOD\t activity\t is\t\ndetermined\t from\t the\t ratio\t of\t autoxidation\t rates\t in\t the\t\npresence\t (Vs)\t and\t in\t the\t absence\t (Vc)\t of\t SOD.\t One\t\nSOD-525\tactivity\tunit\tis\tdefined\tas\tthe\tactivity\tthat\tdou-\nbles\tthe\tautoxidation\trate\tof\tthe\tcontrol\tblank.\t\nAssay of CAT activity. CAT activity was determined \nby\tthe\tmethod\tof\tBeutler\t(1982).\tThe\treaction\tis\tbased\t\non the rate of H2O2 degradation by catalase contained in \nthe\texamined\tsamples.\tThe\treaction\twas\tperformed\tin\t\nan\tincubation\tmixture\tcontaining\t1\tM\tTris-HCl,\t5\tmM\t\nEDTA,\tpH\t8.0,\tand\tmonitored\tspectrophotometrically\tat\t\n230\tnm.\tOne\tunit\tof\tCA T\tactivity\tis\tdefined\tas\t1\tµmol\t\nof H2O2 decomposed per minute under the assay condi-\ntions. \nAssay\tof\tGPx\tactivity.\tGPx\tactivity\twas\tassessed\tus-\ning\tthe\tOxis\tBioxytech®\tGPx-340™\tAssay\t(Oxis\tInter-\nnational,\tInc.),\tbased\ton\tthe\tprinciple\tthat\toxidized\tglu-\ntathione\t(GSSG)\tproduced\tupon\treduction\tof\tan\torganic\t\nperoxide\tby\tGPx\tis\timmediately\trecycled\tto\tits\treduced\t\nform\t(GSH)\twith\tconcomitant\toxidation\tof\tNADPH\tto\t\nNADP+.\tThe\toxidation\tof\tNADPH\twas\tmonitored\tspec-\nReproductive\tFactors/AO\tStatus\tin\tGynaecological\tPatients\n\n28\t V ol.\t61\ntrophotometrically\tas\ta\tdecrease\tin\tabsorbance\tat\t340\tnm.\t\nOne\tGPx-340\tunit\tis\tdefined\tas\t1\tµmol\tof\tNADH\toxi-\ndized per minute under the assay conditions. \nAssay of GR activity. Activity of GR was measured \nusing\tthe\tOxis\tBioxytech®\tGR-340™\tAssay\t(Oxis\tInter-\nnational,\t Inc.).\tThe\t assay\t is\t based\t on\t the\t oxidation\t of\t\nNADPH\t to\t NADP+\t during\t the\t reduction\t of\t oxidized\t\nglutathione\t(GSSG),\tcatalysed\tby\ta\tlimiting\tconcentra-\ntion\tof\tglutathione\treductase.\tThe\toxidation\tof\tNADPH\t\nwas monitored spectrophotometrically as a decrease in \nabsorbance\tat\t340\tnm.\tOne\tGR-340\tunit\tis\tdefined\tas\t1\t\nµmol\t of\t NADH\t oxidized\t per\t minute\t under\t the\t assay\t\nconditions. \nLipid\t hydroperoxides.\t The\t concentration\t of\t LOOH\t\nwas\t measured\t by\t Oxis\t Bioxytech®\t LPO-560™\tAssay\t\n(Oxis\tInternational,\tInc.),\twhich\tis\tbased\ton\tthe\toxida-\ntion of ferrous (Fe2+)\tions\tto\tferric\t(Fe3+)\tions\tby\thydro-\nperoxides\tunder\tacidic\tconditions.\tFerric\tions\tthen\tbind\t\nwith\tthe\tindicator\tdye,\txylenol\torange,\tand\tform\ta\tco-\nloured\t complex.\t The\t absorbance\t of\t the\t complex\t was\t\nmeasured\tat\t560\tnm.\tSince\thydrogen\tperoxide\tcontent\t\nin many biological samples is much higher than that of \nother\t hydroperoxides,\t samples\t were\t pre-treated\t with\t\ncatalase\tto\tdecompose\tthe\texisting\tH\n2O2\tand eliminate \nthe interference. \nStatistical analysis \nStatistical analysis was conducted using the SPSS \nsoftware package. The Pearson correlation method and \nmultivariate regression analysis were used to test the as-\nsociation of age, parity, abortions and AUB with the ac-\ntivities of AO enzymes. The stepwise logistic regression \nmodel, as the most sophisticated one, was used to ensure \nthe smallest possible set of predictor variables in the \nmodel. The principle was to enter each predictor in se-\nquence and to assess its value. If adding the variable \ncontributed to the model, then it was retained, and all \nother variables in the model were re-tested to see if they \nstill contributed to the success of the model. To perform \nthis analysis, variables were assigned certain numerical \nscores\t(Table\t1).\tStatistical\tsignificance\twas\tset\tat\tP\t<\t\n0.05.\nResults\nThe\tPearson\tcorrelation\tmatrix\tobtained\tbetween\t10\t\nvariables\t(5\tpredictors\tand\t5\tdependent\tvariables)\tis\tde-\npicted\tin\tTable\t2.\tIt\tis\tof\tinterest\tto\tnote\tthat,\texcept\tfor\t\nGR activity, all other AO enzymes and LOOH level \nwere correlated with different predictor factors. The ta-\nS.\tPejić\tet\tal.\nTable 1. Score assigned to different factors of multifactorial analysis\nFactor Score assigned\n1 2 3 4 5\nAge\t(years)\nN =\n25-35\n(8)\n36-46\n(24)\n47-57\n(45)\n58-68\n(7)\n69-79\n(4)\nParity\nN =\n0\n(3)\n1\n(7)\n2\n(33)\n3\n(24)\n>\t3\n(21)\nAbortions\nN =\n0\n(0)\n1\n(35)\n2\n(18)\n3\n(17)\n>\t3\n(18)\nAbnormal uterine bleeding*\nN =\nNone\n(30)\nMP\n(21)\nMR\n(16)\nMPM\n(21)\nDiagnosis\nN =\nPE\n(18)\nUM\n(12)\nSH\n(31)\nCH\n(22)\nACE\n(5)\n* MP – Metrorrhagia prolongata, MR – Metrorrhagia recidivans, MPM – Metrorrhagia postmenopausi\nTable 2. Variable correlation matrix in the blood\nVariable Age Parity Abortions Bleeding Diagnosis CuZnSOD CAT GPx GR LOOH\nAge 1.0 0.48*** 0.25** 0.32** 0.26** -0.22* 0.06 -0.07 -0.01 0.05\nParity 1.0 0.26** 0.28** 0.29** -0.23* 0.08 -0.20* -0.10 0.13\nAbortions 1.0 0.12 0.11 -0.27** 0.17* -0.07 -0.12 0.09\nBleeding 1.0 0.80*** -0.49*** 0.30** -0.55*** -0.02 0.35***\nDiagnosis -0.51*** 0.34*** -0.65*** 0.18* 0.39***\nCuZnSOD 1.0 -0.09 0.22* -0.08 -0.17\nCAT 1.0 -0.39*** 0.23** 0.42***\nGPx 1.0 -0.02 -0.34***\nGR 1.0 0.21*\nLOOH 1.0\n*P < 0.05, **P < 0.01, ***P < 0.001\n\nV ol.\t61\t 29\nble\talso\tshows\ta\tsignificant\tcorrelation\tbetween\tthe\tpre-\ndictor variables themselves as well as between the de-\npendant variables. This points to possible interactions \nbetween them in the prediction of AO enzyme activities \nof an individual through multiple regression. \nThe\t final\tpredictive\t model\t with\t multiple\t regression\t\nanalysis\tfor\tAO\tenzyme\tactivities\tis\tshown\tin\tTable\t3\t\nand Fig. 1. These results show that two factors (diagno-\nsis\t and\t abortions)\t fitted\tthe\t best\t predictive\t model\t for\t\nCuZnSOD activity (r\n2\t =\t 0.30,\t P\t <\t 0.001).\t Diagnosis\t\nalone\tcontributed\twith\t~6\t%\tand\tabortions\twith\t~24\t%\t\nto the total variations of the CuZnSOD activity. \nOne\tpredictive\tfactor\talone\t(diagnosis)\tprovided\tthe\t\nbest predictive model for CAT activity (r 2\t= 0.11, P = \n0.001)\t and\t GPx\t activity\t (r2\t =\t 0.42,\t P\t <\t 0.001).\t It\t ex-\nplained\t~11\t%\tof\ttotal\tvariations\tfor\tthe\tactivity\tof\tCA T\t\nand\t 42\t %\t of\t total\t variations\t for\t the\t activity\t of\t GPx.\t\nDiagnosis also scored alone for the predictive model of \nLOOH level (r\n2\t=\t0.15,\tP\t<\t0.001)\tand\tcontributed\twith\t\n15\t%,\twhile\tno\tpredictors\tmet\tcriteria\tfor\tthe\tGR\tactiv-\nity. A large part of variations of AO enzymes and LOOH \nlevel\tremained\tunexplained,\twhich\tprobably\tpoints\tto\ta\t\nrole of many other factors that were not considered in \nthis study or were unknown. \nDiscussion\nThe association of different clinical risk factors and \nvarious types of gynaecological pathologies is still not \nfully\tknown,\tsimilarly\tas\tthe\tinfluence\tthey\texert\ton\tthe\t\nAO status in these patients. In this study, AO enzyme \nactivities\tand\tthe\tlipid\tperoxidation\tlevel\tin\tthe\tblood\tof\t\nwomen with different gynaecological conditions and \nendometrial cancer were related to the diagnosis, AUB, \nage\t and\t reproductive\t factors\t (parity\t and\t abortions)\t to\t\nobserve the strength of the relationship among them and \nindependent association between AO enzymes and each \nindependent variable. \nThe\trelationship\tbetween\tantioxidants\tand\tpathologi-\ncal changes found in this study points to a role of the AO \ndefence mechanisms in the aetiology of various gynae-\ncological disorders. A strong reverse relationship be-\ntween\tSOD\tand\tGPx\tactivities\twith\tdiagnostic\tcatego-\nries was recorded, as well as a positive one between \ndiagnosis\tand\tCA T\tactivity/LOOH\tconcentration.\tThese\t\nobservations\tare\tin\taccordance\twith\tour\tprevious\tfind-\nings\t showing\t a\t decreasing\t trend\t of\t SOD\t and\t GPx\t ac-\ntivities in women with endometrial hyperplasia or ade-\nnocarcinoma in comparison to those with endometrial \npolyp\tor\tleiomyoma\t(Pejić\tet\tal.,\t2006).\tLowered\tSOD\t\nand\tGPx\tactivities\tin\tthe\tplasma\tof\tgynaecological\tpa-\ntients were also reported in other studies (Chiou and Hu, \n1999;\tManoharan\tet\tal.,\t2004).\nIt is known that SOD, as primary scavenger of super-\noxide\t anions,\t along\t with\t GPx\t has\t a\t protective\t role\t\nagainst\t lipid\t peroxidation.\t Thus,\t the\t observed\t reverse\t\nrelationship may be due to the increased endogenous \nproduction of ROS, as also evidenced by the recorded \npositive relationship of pathological changes in differ -\nent diagnosis and LOOH concentration. It is also known \nthat\tthe\tlevels\tof\tsuperoxide\tanion\tand\thydrogen\tperox-\nide increase in various pathological conditions and that \nsuperoxide\tanion\tinactivates\tGPx\t(Blum\tand\tFridovich,\t\n1985).\tIn\tsupport\tof\tthese\tfindings,\ta\tnegative\tcorrela-\ntion\tbetween\tSOD/GPx\tactivities\tand\tLOOH\tlevel\twas\t\nobserved\tin\tgynaecological\tpatients\t(Pejić\tet\tal.,\t2006).\t\nA positive relationship between diagnosis and CAT ac-\ntivity observed in this study indicate that CAT is less \nsensitive\tto\tthe\tredox\tchanges\tin\tthe\tblood\tof\tthe\texam-\nined women. Some studies point to a greater role of this \nenzyme\tin\tprotecting\terythrocytes\tagainst\tperoxidative\t\nstress\tthan\tGPx\t(Mueller\tet\tal.,\t1997).\tThe\tpositive\tcor-\nrelation that we previously recorded between lipid hy-\ndroperoxides\tand\tCA T\tactivity\talso\tsupports\tthis\tfinding\t\n(Pejić\tet\tal.,\t2006).\nWhen\t evaluating\t the\t influence\tof\t reproductive\t fac-\ntors, we found a negative association of abortions with \nSOD\tactivity\tonly,\twhile\tparity\thad\tno\tinfluence\ton\tAO\t\nenzymes\tor\tlipid\tperoxidation.\tStudies\thave\tconsistent-\nly shown an inverse relation between the risk of endo-\nmetrial cancer and the number of births (Cook et al., \n2006).\tHowever,\tdata\tabout\tassociation\tbetween\tone\tor\t\nmore\t incomplete\t pregnancies,\t differently\t defined\t in\t\nstudies as miscarriages or induced abortions, and endo-\nmetrial\tcancer\tare\tmixed\t(Xu\tet\tal.,\t2004;\tPocobelli\tet\t\nal.,\t2011).\tSince\tbenign\tgynaecologic\tdiseases\tand\thy-\nperplasia\t may\t progress\t to\t cancer\t (Ricci\t et\t al.,\t 2002;\t\nBrinton\tet\tal.,\t2005),\tthe\treproductive\tfactors\tare\tcon-\nTable 3. Multiple regression analysis (the strength of the model and independent contribution by significant predictor \nvariables to the AO enzyme activities in the blood)\nActivities of \nAO enzymes\nPredictors B β t P value β\t×\trxy F Model\nP value r2 Adjusted R2\nCuZnSOD Diagnosis -0.231 -0.483 -5.298 0.000 0.059 F2,85\t=\t18.45 0.000 0.303 0.286Abortions -0.094 -0.218 -2.392 0.019 0.244\nCAT Diagnosis 5.512 0.337 3.317 0.001 0.114 F1,86 = 11.00 0.001 0.113 0.103\nGPx Diagnosis -3.210 -0.649 -7.912 0.000 0.421 F1,86\t=\t62.59 0.000 0.421 0.415\nGR No predictors met criteria\nLOOH Diagnosis 0.028 0.386 3.878 0.000 0.149 F1,86\t=\t15.04 0.000 0.149 0.139\nB\t=\tunstandardized\tregression\tcoefficient,\tβ\t=\tstandardized\tregression\tcoefficient,\tF\t=\tF\tstatistics,\twhich\tevaluates\tthe\tmodel,\tr2\t= \nvariance in enzyme activity accounted for by the predictors, t = t statistics, which evaluates the predictor\nReproductive\tFactors/AO\tStatus\tin\tGynaecological\tPatients\n\n30\t V ol.\t61\nsidered to be related with hyperplastic changes as well \n(Epplein\tet\tal.,\t2008).\nMiscarriage and pregnancy appear to be associated \nwith\t increased\t oxidative\t stress.\t During\t uncomplicated\t\npregnancies, ROS levels are elevated at a certain time-\npoint and counterbalanced by the increased activity of \nantioxidants\t (Agarwal\t et\t al.,\t 2012).\t In\t recurrent\t preg-\nnancy\tloss,\tstudies\thave\tpointed\tto\ta\trole\tof\toxidative\t\nstress\t in\t its\t aetiology\t (Poston\t and\t Raijmakers,\t 2004;\t\nAgarwal\tet\tal.,\t2008).\tIn\tthese\tpatients,\tsignificantly\tlow\t\nlevels\tof\tSOD,\tGPx\tand\tCA T\twere\talso\tfound,\tin\taddi-\ntion to an increased malondialdehyde level (El-Far et \nal.,\t 2007).\t Spontaneous\t abortion\t is\t accompanied\t by\t a\t\nprofound\tdisruption\tof\tthe\tpro-oxidant-antioxidant\tho-\nmeostasis\ttowards\toxidative\tstress\t(Lagod\tet\tal.,\t2001)\t\nand\ta\tfirst-trimester\tmiscarriage\twas\tfound\tto\tbe\tassoci-\nated\twith\tsignificantly\treduced\tSOD\tlevels\t(Jenkins\tet\t\nal.,\t2000).\tThus,\ta\tnegative\trelationship\tof\tthe\tSOD\tac-\ntivity and spontaneous or induced abortions observed in \nthis\tstudy\talso\tsupports\tthe\trole\twhich\toxidative\tstress\t\nand AO defence may have in the aetiology of gynaeco -\nlogical disorders. Transformed tissues are known to pro-\nduce\thigh\tlevels\tof\tROS\tand\tare\tconstantly\tunder\toxida-\ntive\tstress\t(Hileman\tet\tal.,\t2001).\tThe\tincrease\tof\tROS,\t\nsuch\tas\tsuperoxide\tanion,\tis\table\tto\tstimulate\tcell\tcycle\t\nprogression and promote cell proliferation by molecular \nmechanisms that include oncogenic signals or respira-\ntory\tchain\tmalfunction\t(Pelicano\tet\tal.,\t2004).\tCell\tdam-\nage\tcaused\tby\tactivated\toxygen\tmetabolites\tand\taltered\t\nAO capacity might be responsible for biological dif-\nferences between transformed and normal tissues (Toyo-\nkuni,\t 2006).\tThe\t negative\t relationship\t that\t indicates\t a\t\nlower\t SOD\t activity\t and\t increased\t superoxide\t concen-\ntrations, observed in our study, implies that patients with \nbenign, premalignant and malignant gynaecological dis-\neases\tare\tlikely\tto\tbe\tunder\toxidative\tstress.\t\nFig. 1. Graphs showing the standard predictive value of the linear composite of predictors vs. CuZnSOD (A),\t CA T\t(B),\t\nGPx\t(C)\tactivities\tand\tLOOH\t(D)\tlevel\nS.\tPejić\tet\tal.\n\nV ol.\t61\t 31\nA\t large\t part\t of\t the\t examined\t correlations\t remained\t\nunexplained,\t which\t probably\t points\t to\t a\t role\t of\t other\t\nfactors that were not considered in this study or were \nunknown. However, this study shows that in gynaeco-\nlogical patients with various diagnoses, the reproductive \nand\tother\tfactors\tmay\tbe\tassociated\twith\tantioxidant\tca-\npacity\tand\tability\tto\tdefend\tagainst\toxidative\tdamage.\t\nThe correlations that were established between the pre-\ndictor variables also indicate possible interactions in the \nprediction\tof\tantioxidant\tenzyme\tactivities.\t\nReferences\nAgarwal,\tA.,\t Gupta,\t S.,\t Sekhon,\t L.,\t Shah\t R.\t (2008)\t Redox\t\nconsiderations in female reproductive function and assisted \nreproduction:\tfrom\tmolecular\tmechanisms\tto\thealth\timpli-\ncations. Antioxid. Redox Signal. 10,\t1375-1403.\nAgarwal, A., Aponte-Mellado, A., Premkumar, B. J., Shaman, \nA.,\tGupta\tS.\t(2012)\tThe\teffects\tof\toxidative\tstress\ton\tfemale\t\nreproduction:\ta\treview.\tReprod. Biol. Endocrinol. 10,\t49.\nBeutler,\tE.\t(1982)\tCatalase.\tIn:\tRed Cell Metabolism, a Ma-\nnual of Biochemical Methods,\ted.\tBeutler,\tE.,\tpp.\t105-106.\t\nGrune and Stratton, New York.\nBlum,\tJ.,\tFridovich,\tI.\t(1985)\tInactivation\tof\tglutathione\tper-\noxidase\t by\t superoxide\t dismutase\tradical.\tArch. Biochem. \nBiophys. 240,\t500-508.\nBrinton, L. A., Sakoda, L. C., Sherman, M. E., Frederiksen, \nK., Kjaer, S. K., Graubard, B. I., Olsen, J. H., Mellemkjaer, \nL.\t(2005)\tRelationship\tof\tbenign\tgynecologic\tdiseases\tto\t\nsubsequent risk of ovarian and uterine tumors. Cancer \nEpidemiol. Biomarkers Prev. 14,\t2929-2935.\nChiou,\tJ.\tF.,\tHu,\tM.\tL.\t(1999)\tElevated\tlipid\tperoxidation\tand\t\ndisturbed\t antioxidant\t enzyme\t activities\t in\t plasma\t and\t\nerythrocytes of patients with uterine cervicitis and myoma. \nClin. Biochem. 32,\t189-192.\nCline,\tJ.\tM.\t(2004)\tNeoplasms\tof\tthe\treproductive\ttract:\tthe\t\nrole\tof\thormone\texposure.\tILAR J. 45,\t179-188.\nCook,\t L.\t S.,\t Weiss,\tN.\t S.,\t Doherty,\t J.\t A.\t Chen,\t C.\t (2006)\t\nEndometrial\tcancer.\tIn:\tCancer Epidemiology and Preven-\ntion,\t eds.\t Schottenfeld,\t D.,\t Fraumeni\t J.\t F.\tJr.,\tpp.\t 1027-\n1044.\tOxford\tUniversity\tPress,\tNew\tY ork.\nDalle-Donne, I., Rossi, R., Colombo, R., Giustarini, D., \nMilzani,\tA.\t(2006)\tBiomarkers\tof\toxidative\tdamage\tin\thu-\nman disease. Clin. Chem. 52,\t601-623.\nEl-Far, M., El-Sayed, I. H., El-Motwally, Ael G., Hashem, I. \nA.,\tBakry,\tN.\t(2007)\tTumor\tnecrosis\tfactor-α\tand\toxidant\t\nstatus\tare\tessential\tparticipating\tfactors\tin\tunexplained\tre-\ncurrent spontaneous abortions. Clin. Chem. Lab. Med. 45, \n879-883.\nEpplein, M., Reed, S. D., V oigt, L. F., Newton, K. M., Holt, V . \nL.\tWeiss,\tN.\tS.\t(2008)\tRisk\tof\tcomplex\tand\tatypical\tendo-\nmetrial hyperplasia in relation to anthropometric measures \nand reproductive history. Am. J. Epidemiol. 168,\t563-570.\nEpstein, E., Valentin, L.\t (2004).\t Managing\t woman\t with\t the\t\npost-menopausal bleeding. Best Pract. Res. Clin. Obstet. \nGynaecol. 18,\t125-143.\nFarquhar, C. M., Lethaby, A., Sowter, M., Verry, J. Baranyai, \nJ.\t(1999)\tAn\tevaluation\tof\trisk\tfactors\tfor\tendometrial\thy-\nperplasia in premenopausal women with abnormal men-\nstrual bleeding. Am. J. Obstet. Gynecol. 181,\t525-529.\nGull,\t B.,\t Karlsson,\t B.,\t Milsom,\t I.,\t Granberg,\tS.\t (2003)\t Can\t\nultrasound\t replace\t dilation\t and\t curettage?\tA\t longitudinal\t\nevaluation of postmenopausal bleeding and transvaginal \nsonographic measurement of the endometrium as predic-\ntors of endometrial cancer. Am. J. Obstet. Gynecol. 188, \n401-408.\nHalliwell,\tB.\t(2006)\tReactive\tspecies\tand\tantioxidants.\tRedox\t\nbiology is a fundamental theme of aerobic life. Plant \nPhysiol. 141,\t312-322.\nHileman,\tE.\tA.,\tAchanta,\tG.,\tHuang,\tP .\t(2001)\tSuperoxide\tdis-\nmutase:\tan\temerging\ttarget\tfor\tcancer\ttherapeutics.\tExpert \nOpin. Ther. Targets 5,\t697-710.\nJenkins, C., Wilson, R., Roberts, J., Miller, H., McKillop, J. \nH.,\t Walker,\tJ.\t J.\t (2000)\tAntioxidants:\t their\t role\t in\t preg-\nnancy and miscarriage. Antiox. Redox Signal. 2,\t623-628.\nKarlsson, B., Granberg, S., Wikland, M., Ylostalo, P., Torvid, \nK.,\t Marsal,\t K.,\t V alentin,\tL.\t (1995)\t Transvaginal\t ultra-\nsonography of the endometrium in women with postmeno-\npausal bleeding – a Nordic multicenter study. Am. J. Obstet. \nGynecol. 172,\t1488-1494.\nLagod.\tL.,\tPaszkowski,\tT.,\tSikorski,\tR.,\tRola,\tR.\t(2001).\tThe\t\nantioxidant-prooxidant\tbalance\tin\tpregnancy\tcomplicated\t\nby spontaneous abortion. Ginekol. Pol. 72,\t1073-1078.\t(in\t\nPolish)\nLowry, O. H., Rosebrough, N. J., Farr, A. L., Randall, R. J. \n(1951)\tProtein\tmeasurement\twith\tthe\tFolin\tphenol\treagent.\t\nJ. Biol. Chem. 193,\t265-275.\t\nManoharan,\t S.,\t Kolanjiappan,\t K.,\t Kayalvizni,\t M.\t (2004)\t\nEnhanced\t lipid\t peroxidation\t and\t impaired\t enzymic\t anti-\noxidant\tactivities\tin\tthe\terythrocytes\tof\tpatients\twith\tcervi-\ncal carcinoma. Cell. Mol. Biol. Lett. 9,\t699-707.\nMueller,\tS.,\t Riedel,\t H.\t D.,\t Stremmel,\tW.\t(1997)\t Direct\t evi-\ndence for catalase as the predominant H\n2O2-removing en-\nzyme in human erythrocytes. Blood 90,\t4973-4978.\nOlson, S. H., Trevisan, M., Marshall, J. R., Graham, S., \nZielezny, M., Vena, J. E., Hellmann, R., Freudenheim, J. L. \n(1995)\tBody\tmass\tindex,\tweight\tgain,\tand\trisk\tof\tendome-\ntrial cancer. Nutr. Cancer \n23,\t141-149.\nPagliuso, R. G., Abbud-Filho, M., Alvarenga M. P. S., Fer -\nreira-Baptista M. A. S., Biselli J. M., Biselli P. M. Goloni-\nBertollo\tE.\tM.,\tPavarino-Bertelli,\tE.\tC.\t(2008)\tRole\tof\tglu-\ntathione S-transferase polymorphisms and chronic allograft \ndysfunction. Transplant. Proc. 40,\t743-745.\nParslov, M., Lidegaard, Ø., Klintorp, S., Pedersen, B., Jønsson, \nL.,\tEriksen,\tP .\tS.,\tOttesen,\tB.\t(2000)\tRisk\tfactors\tamong\t\nyoung\t women\t with\t endometrial\t cancer:\t a\t Danish\t case-\ncontrol study. Am. J. Obstet. Gynecol. 182,\t23-29.\nPejić,\t S.,\t Kasapović,\t J.,\t Todorović,\t A.,\t Stojiljković,\t V .,\t\nPajović,\t S.\t B.\t (2006)\t Lipid\t peroxidation\t and\t antioxidant\t\nstatus in blood of patients with uterine myoma, endome -\ntrial polypus, hyperplastic and malignant endometrium. \nBiol. Res. 39,\t619-629.\nPejić,\t S.,\t Todorović,\t A.,\t Stojiljković,\t V .,\tKasapović,\t J.,\t\nPajović,\tS.\tB.\t(2009)\tAntioxidant\tenzymes\tand\tlipid\tper-\noxidation\tin\tendometrium\tof\tpatients\twith\tpolyps,\tmyoma,\t\nhyperplasia and adenocarcinoma. Reprod. Biol. Endocrin. \n7, 149.\nPelicano,\tH.,\tCarney,\tD.,\tHuang,\tP .\t(2004)\tROS\tstress\tin\tcan-\ncer cells and therapeutic implications. Drug Resist. Updat. \n7, 97-110.\nReproductive\tFactors/AO\tStatus\tin\tGynaecological\tPatients\n\n32\t V ol.\t61\nPike,\t M.\t C.,\t Pearce,\t C.\t L.,\tWu,\tA.\t H.\t (2004)\t Prevention\t of\t\ncancers of the breast, endometrium and ovary. Oncogene \n23,\t6379-6391.\nPocobelli, G., Doherty, J. A., V oigt, L. F., Beresford, S. A., \nHill, D. A., Chen, C., Rossing, M. A., Holmes, R. S., Noor, \nZ.\t S.,\t Weiss,\tN.\t S.\t (2011)\tPregnancy\t history\t and\t risk\t of\t\nendometrial cancer. Epidemiology 22,\t638-645.\nPoston, L., Raijmakers, M. T. (2004)\t Trophoblast\toxidative\t\nstress,\t antioxidants\t and\t pregnancy\t outcome\t –\t a\t review.\t\nPlacenta 25(Suppl A),\tS72–S78.\nPurdie, D.\t M.\t (2003)\t Epidemiology\t of\t endometrial\t cancer.\t\nRev. Gynaecol. Pract. 3,\t217-220.\nRicci, E., Moroni, S., Parazzini, F., Surace, M., Benzi, G., \nSalerio,\tB.,\tPolverino,\tG.,\tLa\tV ecchia,\tC.\t(2002)\tRisk\tfac-\ntors\tfor\tendometrial\thyperplasia:\tresults\tfrom\ta\tcase-con-\ntrol study. Int. J. Gynecol. Cancer 12,\t257-260.\nRowlands, I. J., Nagle, C. M., Spurdle, A. B., Webb, P. M., \nAustralian National Endometrial Cancer Study Group, \nAustra\tlian\tOvarian\tCancer\tStudy\tGroup\t(2011)\tGyne\tcolo-\ngical conditions and the risk of endometrial cancer. \nGynecol. Oncol. 123,\t537-541.\nSilberstein, T., Saphier, O., van V oorhis, B. I., Plosker, S. M. \n(2006)\tEndometrial\tpolyps\tin\treproductive-age\tfertile\tand\t\ninfertile women. IMAJ J. 8,\t192-195.\nSoliman, P. T., Oh, J. C., Schmeler, K. M., Sun, C. C., \nSlomovitz, B. M., Gershenson, D. M., Burke, T. W., Lu, K. \nH. (2005)\t Risk\t factors\t for\t young\t premenopausal\t women\t\nwith endometrial cancer. Obstet. Gynecol. 105,\t575-580.\nStraughn,\tJ.\tM.\tJr.,\tPartridge,\tE.\tE.\t(2009)\tEndometrial\tcan-\ncer.\t In:\t General Surgery, Principles and International \nPractice, eds. Bland, K. I., Sarr, M. G., Büchler, M. W., \nCsendes,\t A.,\t Garden,\t O.\t J.,\t Wong,\t J.,\t pp.\t 1761-1771.\t\nSpringer-Verlag, London Ltd.\nToyokuni,\tS.\t(2006)\tNovel\taspects\tof\toxidative\tstress-associ-\nated carcinogenesis. Antioxid. Redox Signal. 8,\t1373-1377.\nTrentham-Dietz, A., Nichols, H. B., Hampton, J. M., New-\ncomb,\tP .\tA.\t(2006)\tWeight\tchange\tand\trisk\tof\tendometrial\t\ncancer. Int. J. Epidemiol. 35,\t151-158.\nXu,\tW.\tH.,\tXiang,\tY .\tB.,\tRuan,\tZ.\tX.,\tZoeng,\tW.,\tCheng,\tJ.\tR.,\t\nDai,\tQ.,\tGao,\tY .\tT.,\tShu,\tX.O.\t(2004)\tMenstrual\tand\trepro-\nductive\tfactors\tand\tendometrial\tcancer\trisk:\tresults\tfrom\ta\t\npopulation-based case-control study in urban Shanghai. \nInt. J. Cancer 108,\t613-619.\nYamazawa, K., Seki, K., Matsui, H., Kihara, M., Sekiya, S. \n(2000)\tPrognostic\tfactors\tin\tyoung\twomen\twith\tendome-\ntrial\tcarcinoma:\ta\treport\tof\t20\tcases\tand\treview\tof\tlitera-\nture. Int. J. Gynecol. Cancer 10,\t212-222.\nZucchetto, A., Serraino, D., Polesel, J., Negri, E., De Paoli, A., \nDal Maso, L., Montella, M., La Vecchia, C., Franceshi, S., \nTalamini,\tR.\t(2009)\tHormone-related\tfactors\tand\tgyneco-\nlogical conditions in relation to endometrial cancer risk. \nEur. J. Cancer Prev. 18,\t316-321.\nS.\tPejić\tet\tal.","source_license":"CC0","license_restricted":false}