Research ArticleAssociation of IGF-1 CA(n) and IGFBP3 rs2854746Polymorphisms with Endometrial Polyp Risk

Pedro Leopoldo Silva Doria 1 Thomas Moscovitz1 Marcos Tcherniakovsky1

Cesar Eduardo Fernandes1 LucianoMelo Pompei1 MiltonWajman1

Angela Van Nimwegen1 and Sergio Haimovich23

1Department of Obstetrics and Gynecology Faculdade de Medicina do ABC Santo Andre SP Brazil2Head of the Hysteroscopy Unit Del Mar University Hospital Barcelona Spain3Head of Gynecology Ambulatory Surgery Hillel Yaffe Medical CenterTechnion-Israel Technology Institute Hadera Israel

Correspondence should be addressed to Pedro Leopoldo Silva Doria pedrodoria81gmailcom

Received 13 June 2018 Accepted 15 October 2018 Published 13 December 2018

Guest Editor Valentina L La Rosa

Copyright copy 2018 Pedro Leopoldo Silva Doria et al This is an open access article distributed under the Creative CommonsAttribution License which permits unrestricted use distribution and reproduction in any medium provided the original work isproperly cited

Introduction Insulin-like growth factor 1 (IGF-1) is a peptide growth factor that promotes cell proliferation and inhibits apoptosisThe bioavailability of IGF-1 is regulated by the insulin-like growth factor binding protein 3 (IGFBP3) Genetic variations influencethe levels of IGF-1 and IGFBP3 The purpose of this study was to examine the association of polymorphisms IGF-1 CA(n) andIGFBP3 rs2854746 with risk of endometrial polyps Materials and Methods Case control observational study composed of 104women with antecedent of endometrial polyp (case group) and 81 postmenopausal women without antecedent of endometrialdiseases (control group) Genotyping of IGF-1 CA(n) was performed by PCR and fragment analysis by capillary electrophoresisand genotyping of IGFBP3 rs2854746 was performed by PCR-HRM Odds ratios (ORs) and 95 confidence intervals (CIs) werecalculated by logistic regression Results The genotype IGF-1 CA(19)CA(19) was associated with an increased endometrial polyprisk (OR=257 IC 95= 109 - 601) this was also found when combining it with CA(gt19)CA(n) genotypes (OR=218 IC 95=106-447)The IGFBP3 rs2854746 analyses showed the CG genotype having a protective effect for endometrial polyp (OR=037 IC95= 019-073) fact also observed when grouping CG and GG carriers (OR=051 IC 95= 028-093) Conclusion The genotypesCA(19)CA(19) and CA(19)CA(19) + CA(gt19)CA(n) of the IGF-1 CA(n)may be considered a risk for endometrial polyp whereasthe genotypes CG and CG + GG of IGFBP3 rs2854746 polymorphism have an inverse effect of endometrial polyp risk

1 Introduction

Endometrial polyps are defined as nonmalignant peduncu-lated or sessile nodules composed of either functional orbasal endometrium or a combination of the two [1] Theprevalence of endometrial polyps varies between 6 and32 depending on the definition of a polyp the diagnosticmethod used (transvaginal sonography hysteroscopy andorsonohysterography) and the population studied [2]

The pathogenesis of endometrial polyp is multifactorialand imbalance between proliferation and apoptosis plays apivotal role in the process Cell proliferation and apoptosisin the endometrium are complex events involving several

signaling pathways including the insulin-like growth factor[3]

The IGF system includes insulin-like growth factors 1 and2 (IGF-1 and IGF-2) their receptors (IGF-1R and IGF-2R) andsix binding proteins (IGFBP 1-6) Both IGF-1 and IGFBP-3 aregrowth hormone dependent [4]

IGFBP-3 is themajor IGF-binding protein in serum and itserves as a reservoir for the IGFs in circulation [5ndash8] IGFBP-3 carries IGF-1 in circulation and directs it to target tissuesprotects it from proteolytic degradation and regulates itsinteraction with the IGF-1R Additionally IGFBP-3 has itsown IGF-independent apoptotic effects mediated through aspecific cell surface receptor [9]

HindawiBioMed Research InternationalVolume 2018 Article ID 8704346 6 pageshttpsdoiorg10115520188704346

2 BioMed Research International

Both IGF-1 and IGFBP-3 genes are polymorphic in humanpopulations [10] A polymorphism in the IGF-I gene hascomprising a variable length of a CA repeat sequence (IGF-1 CA( n)) [11]

The IGF system is an important regulator of cell prolifera-tion differentiation and apoptosis in various tissues Severalstudies suggest that the imbalance between serum levels ofIGF-1 and IGFBP3 could trigger abnormal cell proliferationincreasing the risk of neoplastic diseases [12]

The polymorphisms that alter gene expression or proteinfunction may result in increased or decreased circulatinglevels of IGF-1 and IGFBP-3 and therefore influence the riskof endometrial disease [7]

The aim of the present study was to investigate the rela-tionship between risk of endometrial polyp and genotypes ofIGF-1 CA( n) and IGFBP-3 rs2854746 polymorphisms

2 Material and Methods

21 Study Population A hundred and eighty-five womenwere evaluated in this molecular epidemiological studyWomen were divided into two groups study and controlgroups The study group consisted of 104 patients with pre-vious history of hysteroscopic polypectomy that underwentsurgery between 2012 and 2015 All of themwere symptomatic(infertility menorrhagia or postmenopausal vaginal bleed-ing) before surgery and had confirmed endometrial polypby histological examination The control group consistedof 81 postmenopausal women without previous history ofendometrial pathology and endometrial thickness less than5 millimeters (transvaginal ultrasonography) that came tooutpatient office for routine gynecologic visit

Patients with a history of any cancer or tamoxifen wereexcluded from the study

The fact that all patients in the control group aremenopausal reinforces the influence of the polymorphism onthe genesis of the polyp

Women fromboth groups were asked to participate in thestudy and after their written informed consent a short ques-tionnaire and peripheral blood sample were obtained from allparticipants The study was approved by the Research EthicsCommittee of the Institution (CAAE 08657412500000082)

22 Genotyping DNA was extracted by standard protocolsfrom peripheral blood as previously described [13] Con-centration and purity were verified by spectrophotometry(NanoDrop USA) Genotyping of the IGF-1 microsatellitepolymorphism (cysteine-alanine or CA repeat) was deter-mined by PCR amplification of the polymorphic regionfollowed by capillary electrophoresis analyses using theABI 3500 DNA Sequencer (Applied Biosystems Foster CityCA) as previously described [14] PCR primers were for-ward 51015840-GCTAGCCAGCTGGTGTTATT-31015840 and Reverse 51015840-ACCACTCTGGGAGAAGGGTA-31015840 the primer forward was51015840-labeled with a fluorescent dye (6-FAM) Fragment sizingwas determined by Genescan analyses software (ABI AppliedBiosystems) The fragments ranged in size from 174 to202 base pairs depending on the number of CA repeatsRepresentative homozygotes (1818 1919 2020 and 2121)

were sequenced to determine (CA)n repeat number frombase pair length Quality control procedures were inclusionof positive and negative controls in each assay run and 20samples were repeated blindly to validate the genotypingprocedures The concordance for the blinded repeat sampleswas 100

Genotyping of rs2854746 polymorphism in IGFBP3 wasdone by PCR-HRM (High Resolution Melting) PCR primersused to amplify the mutation were forward 51015840-CTGGGC-CGCTGCGCTGACTCT-31015840 and reverse 51015840-GCTCGCAGC-GCACCACGGGAC-3 PCR reaction contained 0128mM offorward and reverse primers 125ul of Type-it HRM PCRkit EVA GREEN and 20 120583l do genomic DNA (20ng) Totalvolume per reaction was 25120583l Amplification was carriedout at Rotor Gene 6000 (Qiagen USA) using the followingprogram preincubation for 5 min at 95∘C and amplificationfor 40 cycles of 30 s at 95∘C 30 s at 584∘C and 45 s at 72∘Cafter which a high resolution melting curve was generatedusing the following protocol 5 s at 95∘C 1 min at 60∘Cfollowed by a gradual increase in temperature from 60∘C to97∘C using a ramp rate of 01∘C per s with one measurementper 2 s Quality control included in all assay run included apreviously sequenced homozygous C and G allele samplesheterozygous CG sample and a negative sample and 20samples were repeated blindly to validate the genotypingprocedures The concordance for the blinded repeat sampleswas 98

23 Statistical Analysis ldquoGenotypic counts of controls weretested for HardyndashWeinberg equilibrium using Chi-square(1205942) test Allele estimates were determined as well as thefrequencies of the most common alleles for gene IGF-1and IGFBP3 Linkage disequilibrium (LD) statistics werecomputed using Haploview 40 Logistic regression wasperformed with the presence of polyp as a dependent variableand polymorphisms as independent variablesrdquo [15] Age sexand body mass index were used as confounders Odds ratios(ORs) and 95 confidence intervals (CI) were estimatedfor each polymorphism reference categories were wild typeGene-gene interactions were investigated by estimating mod-els with two polymorphisms one of each of the two genesLikelihood ratio tests were conducted to compare a modelwith interaction effect between the two polymorphisms toa model without interaction term Analysis of the data wasperformed using the software SPSS for Windows version 17All P values are two-sided P values lt 005 were considered tobe statistically significant

3 Results

There were 185 women included in this study They weredivided into two groups control (n=81) and study (n=104)Personal and lifestyle characteristics of both groups areshown in Table 1 Significant differences between the groupswere observed in three variables age prevalence of highblood pressure and previous use of hormonal therapy 46patients (44) of the polyp group are menopausal and 56are in the reproductive period

BioMed Research International 3

Table 1 Epidemiological characterization of patients with arterial hypertension use of hormone replacement therapy (HRT) age groupdiabetes and evaluation of body mass index (BMI)

Control Polyp OR 95 CI P-ValueArterial hypertensionno 44(56) 74(71)yes 35(44) 30(29) 0509 (0275 -0941) 0031no answer 2 0HRTno 44(56) 89(87)yes 34(44) 13(13) 0189 (0090 -0393) lt0001no answer 3 2Age range29-39 3 (4) 36(35)40-49 8 (10) 22(21) 0229 (0054 -0956) 004350-59 52 (65) 18(17) 0028 (0007 -0105) lt000160-80 17 (21) 28(27) 0137 (0036 -0515) 0003no answer 1 0Diabetesno 72 (92) 96 (92)yes 6 (8) 8 (8) 0980no answer 2 0BodyMass Indexnormal 30 (39) 36(37)overweight 25 (33) 35(35)obese 21 (28) 27(28) 0912no answer 5 6

All IGF-1 and IGFBP3 alleles were in HardyndashWeinbergequilibrium in the control population (IGF-1 p = 048 andIGFBP3 p = 032)

There were seven different IGF-1 alleles ranging from 16to 22 CA repeats The IGF-1 CA(19) was the most commonallele in both groups (control 51 and study 56) followedby CA(20) allele (control 15 and study 18) CA(18) allele(control 19 and study 10) and CA(21) allele (control9 and study 11) No other allele frequency exceeded 5(data not shown) Overall the CA(19)CA(19) genotype wasmost common (control 238 e study 32) Next mostcommon genotypes were CA(19)CA(20) (control 225 estudy 17)CA(18)CA(19) (control 163 e study 11) andCA(19)CA(21) (control 125 e study 11) IGF-1 CA(n)genotypes were grouped in three different ways as an attemptto identify the influence of allele length Regression analysisshowed that homozygous CA(19) genotype is associated withendometrial polyp risk (OR=257 IC 95= 109-601 p=02)Further analyses grouping homozygous 19 CA genotype withgenotypes with one allele longer than CA(19) also representeda risk for endometrial polyp CA(gt19)CA(n) (OR= 2181 IC95= 106-447 p =003) (Table 2)

For IGFBP3 the C allele had a frequency of control65 and study 70 The G allele frequency was 35 incontrol and 30 in the study group The most commongenotypes were CG in control group (48) and CC in studygroup (57) Regression analyses showed that CG genotype

has a protective effect on endometrial polyp development(OR= 03 IC 95= 0195-0730 p=0003) Further analysesgrouping GG and CG genotypes showed similar results (OR=051 IC 95= 0284-0937 p=0029) (Table 3)

Interaction among IGF-1 CA(n) and IGFBP3 rs2854746were also investigated The results showed that homozygousCA(19) + genotypes with one allele longer than CA(19) +CC genotype were significant associated with endometrialpolyp risk (OR= 427 IC 95= 164-1109 p=0002) (Table 4)Further analyses correcting the results for confounders (agehigh blood pressure and previous use of hormonal ther-apy) showed similar results (OR= 371 IC 95= 138-100p=0009) The CG genotype appeared as a protective factor(OR= 016 IC 95= 006-040 p=00001) (Table 5)

4 Discussion

Genetic polymorphisms are natural variations in the genomicDNA sequence present in more than 1 of the populationIGF-1 plays an important role in the regulation of cellproliferation differentiation and apoptosis with a recognizedeffect on tumor growth [16]

The IGF-1 gene is located on chromosome 12 (12q 22ndash241)It contains in the promoter region amicrosatellite comprisinga variable length of CA repeat sequence which ranges from10 to 24 [14]

4 BioMed Research International

Table 2 Characterization of the sample for IGF-1 genotyping

Control Polyp OR 95 CI P-ValueGrouping 1CA(19)CA(19) 19 (23) 34 (33)CA(19)CA(n) 44 (54) 49 (47) 0382CA(n)CA(n) 18 (22) 21 (20)Grouping 2CA(19)CA(19) 19 (23) 34 (33) 257 (109-601) 0024CA(19)CA(lt19) + CA(lt19)CA(lt19) 23 (28) 16 (15) 1CA(gt19)CA(n) 39 (48) 54 (52) 199 (093-425) 0076Grouping 3CA(19)CA(lt19) + CA(lt19)CA(lt19) 23 (28) 16 (15) 1CA(19)CA(19) + CA(gt19)CA(n) 58 (72) 88 (85) 218 (106-447) 0033Total 81 104A alanineC cysteineCA allele cysteine-alanine

Table 3 Characterization of the sample for IGFBP3 (rs2854746)genotyping

IGFBP3rs2854746 Control Polyp OR 95 CI P-Value

CC 32 (41) 58 (57) 1CG 38 (48) 26 (26) 037 019-073 0003GG 9 (11) 18 (18) 110 044-273 0831GG + CG 47 (59) 44 (43) 051 028-093 0029No answer 2 2C cysteineG glycineCC allele cysteine-cysteineGG allele glycine-glycineCG allele cysteine-glycine

The importance of IGF-1 CA(n) polymorphism relieson its association with IGF-1 levels This effect seems tobe dependent on the number of CA repeats with highercirculating IGF-1 levels for the CA(19) and CA(20) repeatsalleles while both alleles shorter than CA(19) and longer thanCA(20) repeats seem to have lower circulating IGF-1 levels[10 17]

In the present study we found a statistically significantassociation between endometrial polyp risk and homozy-gous CA(19) genotype Furthermore grouping homozygousCA(19) genotype with genotypes with one allele longer thanCA(19) also represented a risk for endometrial polyp Theseresults suggest that endometrial polyp risk is dependent onthe number of CA repeats [7] So there is a lack of infor-mation regarding the effect of genetic factors on endometrialpolyp risk Therefore our results give new insight overpotential heritable factors associated with the development ofendometrial polyp

The underlying mechanisms by which some genotypes ofIGF-1 CA(n) polymorphism increases the risk of endometrialpolyp was not addressed in our study Nevertheless we

speculates that CA(19) and CA(gt19) alleles may be relatedto higher circulating IGF-1 and consequently augment ofendometrial proliferative activity This hypothesis is based onother studies that found a relationship between high IGF-1levels and abnormal endometrial proliferative activity [18 19]

The IGFBP3 gene is located on chromosome 7 (7p13)and contains five exons In exon 1 there is a nonsynony-mous amino acid change glycine to alanine This changeoccurs at residue 32 in the protein structure a region thathas been shown in fragment analyses to contain a high-affinity binding region for IGF-1 [20] This polymorphism(rs2854746) may have an effect on the concentration ofcirculating IGFBP3 with IGFBP3 levels increasing from CC997888rarr GC 997888rarr GG in cancer-free individuals [21]

We found a statistically significant association betweenIGFBP3 rs2854746 polymorphism and endometrial polyprisk with CG genotype having a protective effect GroupingCG and GG genotype carriers also showed significant inverseassociation with endometrial polyp risk Explanation for thisassociation stem from previously demonstrated relationshipbetween IGFBP3 rs2854746 polymorphism and IGFBP3 lev-els as the presence of G allele displayed higher IGFBP3 levelswhen compared with C allele [21]

In our study we also examined the interactions amongvariants of the two polymorphisms and endometrial polypriskThe results showed that the association of IGF-1homozy-gous CA(19) or genotypes with one allele longer than CA(19)and CC IGFBP3 rs2854746 represents a risk for endometrialpolyp This suggests that associations of endometrial polypwith IGF hormones may be causal and it is not restricted toone member of IGF family Furthermore the disequilibriumbetween IGF-1 and IGFBP3 levels could be the triggeringfactor for endometrial polyp development

Some limitations of this study should be considered (1)the number of participants per group did not allow us toverify the relationship between each homozygous genotypewith endometrial polyp risk (2) ethnicity of study popula-tion could not be determined due to Brazilian population

BioMed Research International 5

Table 4 Results of interactions between IGF-1 CA(n) and IGFBP3 rs2854746

Interaction IGFBP3 e IGF1 Control Polyp OR 95 CI P-ValueCA(lt19)CA(n) + CC 16 (203) 11 (108)CA(lt19)CA(n) + CG 5 (63) 2 (2) 0581 009-355 0557CA(lt19)CA(n) + GG 2 (25) 3 (29) 2181 031-1528 0432CA(19)CA(19)+ CA(gt19)CA(n) + GG 7 (89) 15 (147) 3116 095-1015 0059CA(19)CA(19)+ CA(gt19)CA(n) + CC 16 (203) 47 (461) 4272 164-1109 0002CA(19)CA(19)+ CA(gt19)CA(n) + CG 33 (418) 24 (235) 1057 041-268 0905No answer 2 2Total 81 104

Table 5 Multivariate logistic regression between the IGF-1 CA(n)and IGFBP3 rs2854746 polymorphisms adjusted for hypertensionage range and hormone replacement therapy (HRT) use

OR 95 CI P-ValueHRT

nao 1Sim 02462 (0096-0631) 0003

Arterial hypertensionnao 1sim 08201 (0341-1968) 0657

Age range29-39 140-49 02012 (0042-0946) 004250-59 00326 (0007-0143) 000560-80 01387 (0029-0647) 0012

IGF-1 CA(19)CA(lt19)CA(n) 1CA(19)CA(19) +CA(gt19)CA(n) 37191 (1380-10020) 0009

IGFBP3CC 1CG 01607 (0063-0409) 0001GG 06648 (0192-2298) 0518

admixture and (3) significant differences in confounders(eg age hormonal therapy use and high blood pressure)between control and study groups Women from our studygroup were younger with lower prevalence of high bloodpressure and hormonal therapy use thanwomen from controlgroup However it should be noted that results from mul-tiple logistic regression showed no influence of high bloodpressure on the association between variants of the studiedpolymorphisms and endometrial polyp risk The strength ofour study is that controls were known to be polyp-free andwith no history of endometrial diseases at the time of bloodsampling Our decision to select postmenopausal women ascontrol group was based on their lifetime exposure to riskfactors for endometrial diseaseswithout developing themWespeculate that women with this profile might have some kindof protection against endometrial polyp risk factors

Themultiple logistic regressions showed protective influ-ence of the advancement of age with endometrial polypbecause our control patients were all menopausal and there-fore this may have interfered with this result Somethingsimilar occurs with the use of hormone replacement therapywhere the majority of patients who used the therapy werefrom the control group and thismade the therapy a protectiveeffect in relation to the endometrial polyp

The strength of our study is that controls were knownto be polyp-free and with no history of endometrial dis-eases at the time of blood sampling Our decision to selectpostmenopausal women as control group was based on theirlifetime exposure to risk factors for endometrial diseaseswithout developing them We speculate that women withthis profile might have some kind of protection againstendometrial polyp risk factors

Our study reinforces the importance of polymorphism inthe genesis of the endometrial polyp and genetic variabilitygains more force as an important risk factor The clinicalimportance of this study is that with it we can say that somepolymorphisms are considered risk factors for endometrialpolyp

The results suggest that some genotypes of the IGF-1 CA(n) polymorphism have a risk ratio for endometrialpolyp However some genotypes of the IGFBP3 polymor-phism rs2854746 are inversely related to endometrial polypTherefore it is possible to consider them as risk or protectionfactors according to the genotype expression in question

Data Availability

The data used to support the findings of this study areavailable from the corresponding author upon request

Conflicts of Interest

The authors declare that they have no conflicts of interest andnothing to disclose

Authorsrsquo Contributions

Pedro Leopoldo Silva Doria Thomas Moscovitz MarcosTcherniakovsky and Angela Van Nimwegen contributedequally to this work Cesar Eduardo Fernandes Luciano

6 BioMed Research International

Melo Pompei Milton Wajman and Sergio Haimovich alsocontributed equally to this work

Acknowledgments

We would like to thank all the academic colleagues andpartners of the laboratory who collaborated directly in thedevelopment and creation of this scientific work

References

[1] M TMazur andR J KurmanDiagnosis of Endometrial Biopsiesand Curettings Springer Publishing Company NY USA 1995

[2] EDreisler S Stampe Sorensen PH Ibsen andG Lose ldquoPreva-lence of endometrial polyps and abnormal uterine bleeding in aDanish population aged 20ndash74 yearsrdquo Ultrasound in Obstetricsamp Gynecology vol 33 no 1 pp 102ndash108 2009

[3] M J Arends ldquoApoptosis in the endometriumrdquo Histopathologyvol 35 no 2 pp 174ndash178 1999

[4] H Yu and T Rohan ldquoRole of the insulin-like growth factorfamily in cancer development and progressionrdquo Journal of theNational Cancer Institute vol 92 no 18 pp 1472ndash1489 2000

[5] M Pollak ldquoInsulin-like growth factor physiology and cancerriskrdquo European Journal of Cancer vol 36 no 10 pp 1224ndash12282000

[6] N Vaessen P Heutink J A Janssen et al ldquoA polymorphismin the gene for IGF-I functional properties and risk for type 2diabetes and myocardial infarctionrdquo Diabetes vol 50 no 3 pp637ndash642 2001

[7] M McGrath I-M Lee J Buring and I De Vivo ldquoCommongenetic variationwithin IGFI IGFII IGFBP-1 and IGFBP-3 andendometrial cancer riskrdquo Gynecologic Oncology vol 120 no 2pp 174ndash178 2011

[8] J Pavelic B Radakovic and K Pavelic ldquoInsulin-like growthfactor 2 and its receptors (IGF 1R and IGF 2Rmannose6-phosphate) in endometrial adenocarcinomardquo GynecologicOncology vol 105 no 3 pp 727ndash735 2007

[9] E Marshman and C H Streuli ldquoInsulin-like growth factorsand insulin-like growth factor binding proteins in mammarygland functionrdquo Breast Cancer Research vol 4 no 6 pp 231ndash239 2002

[10] C Deal J Ma F Wilkin et al ldquoNovel promoter polymorphismin insulin-like growth factor-binding protein-3 Correlationwith serum levels and interaction with known regulatorsrdquoTheJournal of Clinical Endocrinology amp Metabolism vol 86 no 3pp 1274ndash1280 2001

[11] J L Weber and P E May ldquoAbundant class of human DNApolymorphisms which can be typed using the polymerase chainreactionrdquo American Journal of Human Genetics vol 44 no 3pp 388ndash396 1989

[12] J Ma M N Pollak and E Giovannucci ldquoProspective study of356 colorectal cancer risk in men and plasma levels of insulin-like growth factos IGF-1 and IGF binding protein-3rdquo JNCIJournal of the National Cancer Institute vol 91 no 23 pp 620ndash625 1999

[13] Embrapa Fundamentos teorico-praticos e protocolos deextracao e de amplificacao de DNA por meio da tecnicade reacao em cadeia da polimerase Sao Carlos Jun 2007Avaliable from httpwwwcppseembrapabrservicospublic-acaogratuitae-booksLVFundDNACited 18 January 2018

[14] T O Keku A Vidal S Oliver et al ldquoGenetic variants in IGF-IIGF-II IGFBP-3 and adiponectin genes and colon cancer riskin African Americans and Whitesrdquo Cancer Causes amp Controlvol 23 no 7 pp 1127ndash1138 2012

[15] E Feik A Baierl B Hieger et al ldquoAssociation of IGF1 andIGFBP3 polymorphisms with colorectal polyps and colorectalcancer riskrdquo Cancer Causes amp Control vol 21 no 1 pp 91ndash972010

[16] M Harrela H Koistinen J Kaprio et al ldquoGenetic and environ-mental components of interindividual variation in circulatinglevels of IGF-I IGF-II IGFBP-1 and IGFBP-3rdquo The Journal ofClinical Investigation vol 98 no 11 pp 2612ndash2615 1996

[17] I Rietveld J A M J L Janssen E F C Van Rossum et al ldquoApolymorphic CA in the IGF-I gene is associated with gender-specific differences in body height but has no effect on thesecular trend in body heightrdquoClinical Endocrinology vol 61 no2 pp 195ndash203 2004

[18] E-M Rutanen ldquoInsulin-like growth factors and insulin-likegrowth factor binding proteins in the endometrium Effect ofintrauterine levonorgestrel deliveryrdquoHuman Reproduction vol15 no 3 pp 173ndash181 2000

[19] M Nagamani C A Stuart P A Dunhardt and M G DohertyldquoSpecific binding sites for insulin and insulin-like growth factorI in human endometrial cancerrdquo American Journal of Obstetricsamp Gynecology vol 165 no 6 pp 1865ndash1871 1991

[20] T Zou A S Fleisher D Kong et al ldquoSequence alterations ofinsulin-like growth factor binding protein 3 in neoplastic andnormal gastrointestinal tissuesrdquoCancer Research vol 58 no 21pp 4802ndash4804 1998

[21] L M Morimoto P A Newcomb E White J Bigler and J DPotter ldquoVariation in plasma insulin-like growth factor-1 andinsulin-like growth factor binding protein-3 Genetic factorsrdquoCancer Epidemiology Biomarkers amp Prevention vol 14 no 6pp 1394ndash1401 2005

BioMed Research International 3

Table 1 Epidemiological characterization of patients with arterial hypertension use of hormone replacement therapy (HRT) age groupdiabetes and evaluation of body mass index (BMI)

Control Polyp OR 95 CI P-ValueArterial hypertensionno 44(56) 74(71)yes 35(44) 30(29) 0509 (0275 -0941) 0031no answer 2 0HRTno 44(56) 89(87)yes 34(44) 13(13) 0189 (0090 -0393) lt0001no answer 3 2Age range29-39 3 (4) 36(35)40-49 8 (10) 22(21) 0229 (0054 -0956) 004350-59 52 (65) 18(17) 0028 (0007 -0105) lt000160-80 17 (21) 28(27) 0137 (0036 -0515) 0003no answer 1 0Diabetesno 72 (92) 96 (92)yes 6 (8) 8 (8) 0980no answer 2 0BodyMass Indexnormal 30 (39) 36(37)overweight 25 (33) 35(35)obese 21 (28) 27(28) 0912no answer 5 6

All IGF-1 and IGFBP3 alleles were in HardyndashWeinbergequilibrium in the control population (IGF-1 p = 048 andIGFBP3 p = 032)

There were seven different IGF-1 alleles ranging from 16to 22 CA repeats The IGF-1 CA(19) was the most commonallele in both groups (control 51 and study 56) followedby CA(20) allele (control 15 and study 18) CA(18) allele(control 19 and study 10) and CA(21) allele (control9 and study 11) No other allele frequency exceeded 5(data not shown) Overall the CA(19)CA(19) genotype wasmost common (control 238 e study 32) Next mostcommon genotypes were CA(19)CA(20) (control 225 estudy 17)CA(18)CA(19) (control 163 e study 11) andCA(19)CA(21) (control 125 e study 11) IGF-1 CA(n)genotypes were grouped in three different ways as an attemptto identify the influence of allele length Regression analysisshowed that homozygous CA(19) genotype is associated withendometrial polyp risk (OR=257 IC 95= 109-601 p=02)Further analyses grouping homozygous 19 CA genotype withgenotypes with one allele longer than CA(19) also representeda risk for endometrial polyp CA(gt19)CA(n) (OR= 2181 IC95= 106-447 p =003) (Table 2)

For IGFBP3 the C allele had a frequency of control65 and study 70 The G allele frequency was 35 incontrol and 30 in the study group The most commongenotypes were CG in control group (48) and CC in studygroup (57) Regression analyses showed that CG genotype

has a protective effect on endometrial polyp development(OR= 03 IC 95= 0195-0730 p=0003) Further analysesgrouping GG and CG genotypes showed similar results (OR=051 IC 95= 0284-0937 p=0029) (Table 3)

Interaction among IGF-1 CA(n) and IGFBP3 rs2854746were also investigated The results showed that homozygousCA(19) + genotypes with one allele longer than CA(19) +CC genotype were significant associated with endometrialpolyp risk (OR= 427 IC 95= 164-1109 p=0002) (Table 4)Further analyses correcting the results for confounders (agehigh blood pressure and previous use of hormonal ther-apy) showed similar results (OR= 371 IC 95= 138-100p=0009) The CG genotype appeared as a protective factor(OR= 016 IC 95= 006-040 p=00001) (Table 5)

4 Discussion

Genetic polymorphisms are natural variations in the genomicDNA sequence present in more than 1 of the populationIGF-1 plays an important role in the regulation of cellproliferation differentiation and apoptosis with a recognizedeffect on tumor growth [16]

The IGF-1 gene is located on chromosome 12 (12q 22ndash241)It contains in the promoter region amicrosatellite comprisinga variable length of CA repeat sequence which ranges from10 to 24 [14]

4 BioMed Research International

Table 2 Characterization of the sample for IGF-1 genotyping

Control Polyp OR 95 CI P-ValueGrouping 1CA(19)CA(19) 19 (23) 34 (33)CA(19)CA(n) 44 (54) 49 (47) 0382CA(n)CA(n) 18 (22) 21 (20)Grouping 2CA(19)CA(19) 19 (23) 34 (33) 257 (109-601) 0024CA(19)CA(lt19) + CA(lt19)CA(lt19) 23 (28) 16 (15) 1CA(gt19)CA(n) 39 (48) 54 (52) 199 (093-425) 0076Grouping 3CA(19)CA(lt19) + CA(lt19)CA(lt19) 23 (28) 16 (15) 1CA(19)CA(19) + CA(gt19)CA(n) 58 (72) 88 (85) 218 (106-447) 0033Total 81 104A alanineC cysteineCA allele cysteine-alanine

Table 3 Characterization of the sample for IGFBP3 (rs2854746)genotyping

IGFBP3rs2854746 Control Polyp OR 95 CI P-Value

CC 32 (41) 58 (57) 1CG 38 (48) 26 (26) 037 019-073 0003GG 9 (11) 18 (18) 110 044-273 0831GG + CG 47 (59) 44 (43) 051 028-093 0029No answer 2 2C cysteineG glycineCC allele cysteine-cysteineGG allele glycine-glycineCG allele cysteine-glycine

The importance of IGF-1 CA(n) polymorphism relieson its association with IGF-1 levels This effect seems tobe dependent on the number of CA repeats with highercirculating IGF-1 levels for the CA(19) and CA(20) repeatsalleles while both alleles shorter than CA(19) and longer thanCA(20) repeats seem to have lower circulating IGF-1 levels[10 17]

In the present study we found a statistically significantassociation between endometrial polyp risk and homozy-gous CA(19) genotype Furthermore grouping homozygousCA(19) genotype with genotypes with one allele longer thanCA(19) also represented a risk for endometrial polyp Theseresults suggest that endometrial polyp risk is dependent onthe number of CA repeats [7] So there is a lack of infor-mation regarding the effect of genetic factors on endometrialpolyp risk Therefore our results give new insight overpotential heritable factors associated with the development ofendometrial polyp

The underlying mechanisms by which some genotypes ofIGF-1 CA(n) polymorphism increases the risk of endometrialpolyp was not addressed in our study Nevertheless we

speculates that CA(19) and CA(gt19) alleles may be relatedto higher circulating IGF-1 and consequently augment ofendometrial proliferative activity This hypothesis is based onother studies that found a relationship between high IGF-1levels and abnormal endometrial proliferative activity [18 19]

The IGFBP3 gene is located on chromosome 7 (7p13)and contains five exons In exon 1 there is a nonsynony-mous amino acid change glycine to alanine This changeoccurs at residue 32 in the protein structure a region thathas been shown in fragment analyses to contain a high-affinity binding region for IGF-1 [20] This polymorphism(rs2854746) may have an effect on the concentration ofcirculating IGFBP3 with IGFBP3 levels increasing from CC997888rarr GC 997888rarr GG in cancer-free individuals [21]

We found a statistically significant association betweenIGFBP3 rs2854746 polymorphism and endometrial polyprisk with CG genotype having a protective effect GroupingCG and GG genotype carriers also showed significant inverseassociation with endometrial polyp risk Explanation for thisassociation stem from previously demonstrated relationshipbetween IGFBP3 rs2854746 polymorphism and IGFBP3 lev-els as the presence of G allele displayed higher IGFBP3 levelswhen compared with C allele [21]

In our study we also examined the interactions amongvariants of the two polymorphisms and endometrial polypriskThe results showed that the association of IGF-1homozy-gous CA(19) or genotypes with one allele longer than CA(19)and CC IGFBP3 rs2854746 represents a risk for endometrialpolyp This suggests that associations of endometrial polypwith IGF hormones may be causal and it is not restricted toone member of IGF family Furthermore the disequilibriumbetween IGF-1 and IGFBP3 levels could be the triggeringfactor for endometrial polyp development

Some limitations of this study should be considered (1)the number of participants per group did not allow us toverify the relationship between each homozygous genotypewith endometrial polyp risk (2) ethnicity of study popula-tion could not be determined due to Brazilian population

BioMed Research International 5

Table 4 Results of interactions between IGF-1 CA(n) and IGFBP3 rs2854746

Interaction IGFBP3 e IGF1 Control Polyp OR 95 CI P-ValueCA(lt19)CA(n) + CC 16 (203) 11 (108)CA(lt19)CA(n) + CG 5 (63) 2 (2) 0581 009-355 0557CA(lt19)CA(n) + GG 2 (25) 3 (29) 2181 031-1528 0432CA(19)CA(19)+ CA(gt19)CA(n) + GG 7 (89) 15 (147) 3116 095-1015 0059CA(19)CA(19)+ CA(gt19)CA(n) + CC 16 (203) 47 (461) 4272 164-1109 0002CA(19)CA(19)+ CA(gt19)CA(n) + CG 33 (418) 24 (235) 1057 041-268 0905No answer 2 2Total 81 104

Table 5 Multivariate logistic regression between the IGF-1 CA(n)and IGFBP3 rs2854746 polymorphisms adjusted for hypertensionage range and hormone replacement therapy (HRT) use

OR 95 CI P-ValueHRT

nao 1Sim 02462 (0096-0631) 0003

Arterial hypertensionnao 1sim 08201 (0341-1968) 0657

Age range29-39 140-49 02012 (0042-0946) 004250-59 00326 (0007-0143) 000560-80 01387 (0029-0647) 0012

IGF-1 CA(19)CA(lt19)CA(n) 1CA(19)CA(19) +CA(gt19)CA(n) 37191 (1380-10020) 0009

IGFBP3CC 1CG 01607 (0063-0409) 0001GG 06648 (0192-2298) 0518

admixture and (3) significant differences in confounders(eg age hormonal therapy use and high blood pressure)between control and study groups Women from our studygroup were younger with lower prevalence of high bloodpressure and hormonal therapy use thanwomen from controlgroup However it should be noted that results from mul-tiple logistic regression showed no influence of high bloodpressure on the association between variants of the studiedpolymorphisms and endometrial polyp risk The strength ofour study is that controls were known to be polyp-free andwith no history of endometrial diseases at the time of bloodsampling Our decision to select postmenopausal women ascontrol group was based on their lifetime exposure to riskfactors for endometrial diseaseswithout developing themWespeculate that women with this profile might have some kindof protection against endometrial polyp risk factors

Themultiple logistic regressions showed protective influ-ence of the advancement of age with endometrial polypbecause our control patients were all menopausal and there-fore this may have interfered with this result Somethingsimilar occurs with the use of hormone replacement therapywhere the majority of patients who used the therapy werefrom the control group and thismade the therapy a protectiveeffect in relation to the endometrial polyp

The strength of our study is that controls were knownto be polyp-free and with no history of endometrial dis-eases at the time of blood sampling Our decision to selectpostmenopausal women as control group was based on theirlifetime exposure to risk factors for endometrial diseaseswithout developing them We speculate that women withthis profile might have some kind of protection againstendometrial polyp risk factors

Our study reinforces the importance of polymorphism inthe genesis of the endometrial polyp and genetic variabilitygains more force as an important risk factor The clinicalimportance of this study is that with it we can say that somepolymorphisms are considered risk factors for endometrialpolyp

The results suggest that some genotypes of the IGF-1 CA(n) polymorphism have a risk ratio for endometrialpolyp However some genotypes of the IGFBP3 polymor-phism rs2854746 are inversely related to endometrial polypTherefore it is possible to consider them as risk or protectionfactors according to the genotype expression in question

Data Availability

The data used to support the findings of this study areavailable from the corresponding author upon request

Conflicts of Interest

The authors declare that they have no conflicts of interest andnothing to disclose

Authorsrsquo Contributions

Pedro Leopoldo Silva Doria Thomas Moscovitz MarcosTcherniakovsky and Angela Van Nimwegen contributedequally to this work Cesar Eduardo Fernandes Luciano

6 BioMed Research International

Melo Pompei Milton Wajman and Sergio Haimovich alsocontributed equally to this work

Acknowledgments

We would like to thank all the academic colleagues andpartners of the laboratory who collaborated directly in thedevelopment and creation of this scientific work

References

[1] M TMazur andR J KurmanDiagnosis of Endometrial Biopsiesand Curettings Springer Publishing Company NY USA 1995

[2] EDreisler S Stampe Sorensen PH Ibsen andG Lose ldquoPreva-lence of endometrial polyps and abnormal uterine bleeding in aDanish population aged 20ndash74 yearsrdquo Ultrasound in Obstetricsamp Gynecology vol 33 no 1 pp 102ndash108 2009

[3] M J Arends ldquoApoptosis in the endometriumrdquo Histopathologyvol 35 no 2 pp 174ndash178 1999

[4] H Yu and T Rohan ldquoRole of the insulin-like growth factorfamily in cancer development and progressionrdquo Journal of theNational Cancer Institute vol 92 no 18 pp 1472ndash1489 2000

[5] M Pollak ldquoInsulin-like growth factor physiology and cancerriskrdquo European Journal of Cancer vol 36 no 10 pp 1224ndash12282000

[6] N Vaessen P Heutink J A Janssen et al ldquoA polymorphismin the gene for IGF-I functional properties and risk for type 2diabetes and myocardial infarctionrdquo Diabetes vol 50 no 3 pp637ndash642 2001

[7] M McGrath I-M Lee J Buring and I De Vivo ldquoCommongenetic variationwithin IGFI IGFII IGFBP-1 and IGFBP-3 andendometrial cancer riskrdquo Gynecologic Oncology vol 120 no 2pp 174ndash178 2011

[8] J Pavelic B Radakovic and K Pavelic ldquoInsulin-like growthfactor 2 and its receptors (IGF 1R and IGF 2Rmannose6-phosphate) in endometrial adenocarcinomardquo GynecologicOncology vol 105 no 3 pp 727ndash735 2007

[9] E Marshman and C H Streuli ldquoInsulin-like growth factorsand insulin-like growth factor binding proteins in mammarygland functionrdquo Breast Cancer Research vol 4 no 6 pp 231ndash239 2002

[10] C Deal J Ma F Wilkin et al ldquoNovel promoter polymorphismin insulin-like growth factor-binding protein-3 Correlationwith serum levels and interaction with known regulatorsrdquoTheJournal of Clinical Endocrinology amp Metabolism vol 86 no 3pp 1274ndash1280 2001

[11] J L Weber and P E May ldquoAbundant class of human DNApolymorphisms which can be typed using the polymerase chainreactionrdquo American Journal of Human Genetics vol 44 no 3pp 388ndash396 1989

[12] J Ma M N Pollak and E Giovannucci ldquoProspective study of356 colorectal cancer risk in men and plasma levels of insulin-like growth factos IGF-1 and IGF binding protein-3rdquo JNCIJournal of the National Cancer Institute vol 91 no 23 pp 620ndash625 1999

[13] Embrapa Fundamentos teorico-praticos e protocolos deextracao e de amplificacao de DNA por meio da tecnicade reacao em cadeia da polimerase Sao Carlos Jun 2007Avaliable from httpwwwcppseembrapabrservicospublic-acaogratuitae-booksLVFundDNACited 18 January 2018

[14] T O Keku A Vidal S Oliver et al ldquoGenetic variants in IGF-IIGF-II IGFBP-3 and adiponectin genes and colon cancer riskin African Americans and Whitesrdquo Cancer Causes amp Controlvol 23 no 7 pp 1127ndash1138 2012

[15] E Feik A Baierl B Hieger et al ldquoAssociation of IGF1 andIGFBP3 polymorphisms with colorectal polyps and colorectalcancer riskrdquo Cancer Causes amp Control vol 21 no 1 pp 91ndash972010

[16] M Harrela H Koistinen J Kaprio et al ldquoGenetic and environ-mental components of interindividual variation in circulatinglevels of IGF-I IGF-II IGFBP-1 and IGFBP-3rdquo The Journal ofClinical Investigation vol 98 no 11 pp 2612ndash2615 1996

[17] I Rietveld J A M J L Janssen E F C Van Rossum et al ldquoApolymorphic CA in the IGF-I gene is associated with gender-specific differences in body height but has no effect on thesecular trend in body heightrdquoClinical Endocrinology vol 61 no2 pp 195ndash203 2004

[18] E-M Rutanen ldquoInsulin-like growth factors and insulin-likegrowth factor binding proteins in the endometrium Effect ofintrauterine levonorgestrel deliveryrdquoHuman Reproduction vol15 no 3 pp 173ndash181 2000

[19] M Nagamani C A Stuart P A Dunhardt and M G DohertyldquoSpecific binding sites for insulin and insulin-like growth factorI in human endometrial cancerrdquo American Journal of Obstetricsamp Gynecology vol 165 no 6 pp 1865ndash1871 1991

[20] T Zou A S Fleisher D Kong et al ldquoSequence alterations ofinsulin-like growth factor binding protein 3 in neoplastic andnormal gastrointestinal tissuesrdquoCancer Research vol 58 no 21pp 4802ndash4804 1998

[21] L M Morimoto P A Newcomb E White J Bigler and J DPotter ldquoVariation in plasma insulin-like growth factor-1 andinsulin-like growth factor binding protein-3 Genetic factorsrdquoCancer Epidemiology Biomarkers amp Prevention vol 14 no 6pp 1394ndash1401 2005

4 BioMed Research International

Table 2 Characterization of the sample for IGF-1 genotyping

Control Polyp OR 95 CI P-ValueGrouping 1CA(19)CA(19) 19 (23) 34 (33)CA(19)CA(n) 44 (54) 49 (47) 0382CA(n)CA(n) 18 (22) 21 (20)Grouping 2CA(19)CA(19) 19 (23) 34 (33) 257 (109-601) 0024CA(19)CA(lt19) + CA(lt19)CA(lt19) 23 (28) 16 (15) 1CA(gt19)CA(n) 39 (48) 54 (52) 199 (093-425) 0076Grouping 3CA(19)CA(lt19) + CA(lt19)CA(lt19) 23 (28) 16 (15) 1CA(19)CA(19) + CA(gt19)CA(n) 58 (72) 88 (85) 218 (106-447) 0033Total 81 104A alanineC cysteineCA allele cysteine-alanine

Table 3 Characterization of the sample for IGFBP3 (rs2854746)genotyping

IGFBP3rs2854746 Control Polyp OR 95 CI P-Value

CC 32 (41) 58 (57) 1CG 38 (48) 26 (26) 037 019-073 0003GG 9 (11) 18 (18) 110 044-273 0831GG + CG 47 (59) 44 (43) 051 028-093 0029No answer 2 2C cysteineG glycineCC allele cysteine-cysteineGG allele glycine-glycineCG allele cysteine-glycine

The importance of IGF-1 CA(n) polymorphism relieson its association with IGF-1 levels This effect seems tobe dependent on the number of CA repeats with highercirculating IGF-1 levels for the CA(19) and CA(20) repeatsalleles while both alleles shorter than CA(19) and longer thanCA(20) repeats seem to have lower circulating IGF-1 levels[10 17]

In the present study we found a statistically significantassociation between endometrial polyp risk and homozy-gous CA(19) genotype Furthermore grouping homozygousCA(19) genotype with genotypes with one allele longer thanCA(19) also represented a risk for endometrial polyp Theseresults suggest that endometrial polyp risk is dependent onthe number of CA repeats [7] So there is a lack of infor-mation regarding the effect of genetic factors on endometrialpolyp risk Therefore our results give new insight overpotential heritable factors associated with the development ofendometrial polyp

The underlying mechanisms by which some genotypes ofIGF-1 CA(n) polymorphism increases the risk of endometrialpolyp was not addressed in our study Nevertheless we

speculates that CA(19) and CA(gt19) alleles may be relatedto higher circulating IGF-1 and consequently augment ofendometrial proliferative activity This hypothesis is based onother studies that found a relationship between high IGF-1levels and abnormal endometrial proliferative activity [18 19]

The IGFBP3 gene is located on chromosome 7 (7p13)and contains five exons In exon 1 there is a nonsynony-mous amino acid change glycine to alanine This changeoccurs at residue 32 in the protein structure a region thathas been shown in fragment analyses to contain a high-affinity binding region for IGF-1 [20] This polymorphism(rs2854746) may have an effect on the concentration ofcirculating IGFBP3 with IGFBP3 levels increasing from CC997888rarr GC 997888rarr GG in cancer-free individuals [21]

We found a statistically significant association betweenIGFBP3 rs2854746 polymorphism and endometrial polyprisk with CG genotype having a protective effect GroupingCG and GG genotype carriers also showed significant inverseassociation with endometrial polyp risk Explanation for thisassociation stem from previously demonstrated relationshipbetween IGFBP3 rs2854746 polymorphism and IGFBP3 lev-els as the presence of G allele displayed higher IGFBP3 levelswhen compared with C allele [21]

In our study we also examined the interactions amongvariants of the two polymorphisms and endometrial polypriskThe results showed that the association of IGF-1homozy-gous CA(19) or genotypes with one allele longer than CA(19)and CC IGFBP3 rs2854746 represents a risk for endometrialpolyp This suggests that associations of endometrial polypwith IGF hormones may be causal and it is not restricted toone member of IGF family Furthermore the disequilibriumbetween IGF-1 and IGFBP3 levels could be the triggeringfactor for endometrial polyp development

Some limitations of this study should be considered (1)the number of participants per group did not allow us toverify the relationship between each homozygous genotypewith endometrial polyp risk (2) ethnicity of study popula-tion could not be determined due to Brazilian population

BioMed Research International 5

Table 4 Results of interactions between IGF-1 CA(n) and IGFBP3 rs2854746

Interaction IGFBP3 e IGF1 Control Polyp OR 95 CI P-ValueCA(lt19)CA(n) + CC 16 (203) 11 (108)CA(lt19)CA(n) + CG 5 (63) 2 (2) 0581 009-355 0557CA(lt19)CA(n) + GG 2 (25) 3 (29) 2181 031-1528 0432CA(19)CA(19)+ CA(gt19)CA(n) + GG 7 (89) 15 (147) 3116 095-1015 0059CA(19)CA(19)+ CA(gt19)CA(n) + CC 16 (203) 47 (461) 4272 164-1109 0002CA(19)CA(19)+ CA(gt19)CA(n) + CG 33 (418) 24 (235) 1057 041-268 0905No answer 2 2Total 81 104

Table 5 Multivariate logistic regression between the IGF-1 CA(n)and IGFBP3 rs2854746 polymorphisms adjusted for hypertensionage range and hormone replacement therapy (HRT) use

OR 95 CI P-ValueHRT

nao 1Sim 02462 (0096-0631) 0003

Arterial hypertensionnao 1sim 08201 (0341-1968) 0657

Age range29-39 140-49 02012 (0042-0946) 004250-59 00326 (0007-0143) 000560-80 01387 (0029-0647) 0012

IGF-1 CA(19)CA(lt19)CA(n) 1CA(19)CA(19) +CA(gt19)CA(n) 37191 (1380-10020) 0009

IGFBP3CC 1CG 01607 (0063-0409) 0001GG 06648 (0192-2298) 0518

admixture and (3) significant differences in confounders(eg age hormonal therapy use and high blood pressure)between control and study groups Women from our studygroup were younger with lower prevalence of high bloodpressure and hormonal therapy use thanwomen from controlgroup However it should be noted that results from mul-tiple logistic regression showed no influence of high bloodpressure on the association between variants of the studiedpolymorphisms and endometrial polyp risk The strength ofour study is that controls were known to be polyp-free andwith no history of endometrial diseases at the time of bloodsampling Our decision to select postmenopausal women ascontrol group was based on their lifetime exposure to riskfactors for endometrial diseaseswithout developing themWespeculate that women with this profile might have some kindof protection against endometrial polyp risk factors

Themultiple logistic regressions showed protective influ-ence of the advancement of age with endometrial polypbecause our control patients were all menopausal and there-fore this may have interfered with this result Somethingsimilar occurs with the use of hormone replacement therapywhere the majority of patients who used the therapy werefrom the control group and thismade the therapy a protectiveeffect in relation to the endometrial polyp

The strength of our study is that controls were knownto be polyp-free and with no history of endometrial dis-eases at the time of blood sampling Our decision to selectpostmenopausal women as control group was based on theirlifetime exposure to risk factors for endometrial diseaseswithout developing them We speculate that women withthis profile might have some kind of protection againstendometrial polyp risk factors

Our study reinforces the importance of polymorphism inthe genesis of the endometrial polyp and genetic variabilitygains more force as an important risk factor The clinicalimportance of this study is that with it we can say that somepolymorphisms are considered risk factors for endometrialpolyp

The results suggest that some genotypes of the IGF-1 CA(n) polymorphism have a risk ratio for endometrialpolyp However some genotypes of the IGFBP3 polymor-phism rs2854746 are inversely related to endometrial polypTherefore it is possible to consider them as risk or protectionfactors according to the genotype expression in question

Data Availability

The data used to support the findings of this study areavailable from the corresponding author upon request

Conflicts of Interest

The authors declare that they have no conflicts of interest andnothing to disclose

Authorsrsquo Contributions

Pedro Leopoldo Silva Doria Thomas Moscovitz MarcosTcherniakovsky and Angela Van Nimwegen contributedequally to this work Cesar Eduardo Fernandes Luciano

6 BioMed Research International

Melo Pompei Milton Wajman and Sergio Haimovich alsocontributed equally to this work

Acknowledgments

We would like to thank all the academic colleagues andpartners of the laboratory who collaborated directly in thedevelopment and creation of this scientific work

References

[1] M TMazur andR J KurmanDiagnosis of Endometrial Biopsiesand Curettings Springer Publishing Company NY USA 1995

[2] EDreisler S Stampe Sorensen PH Ibsen andG Lose ldquoPreva-lence of endometrial polyps and abnormal uterine bleeding in aDanish population aged 20ndash74 yearsrdquo Ultrasound in Obstetricsamp Gynecology vol 33 no 1 pp 102ndash108 2009

[3] M J Arends ldquoApoptosis in the endometriumrdquo Histopathologyvol 35 no 2 pp 174ndash178 1999

[4] H Yu and T Rohan ldquoRole of the insulin-like growth factorfamily in cancer development and progressionrdquo Journal of theNational Cancer Institute vol 92 no 18 pp 1472ndash1489 2000

[5] M Pollak ldquoInsulin-like growth factor physiology and cancerriskrdquo European Journal of Cancer vol 36 no 10 pp 1224ndash12282000

[6] N Vaessen P Heutink J A Janssen et al ldquoA polymorphismin the gene for IGF-I functional properties and risk for type 2diabetes and myocardial infarctionrdquo Diabetes vol 50 no 3 pp637ndash642 2001

[7] M McGrath I-M Lee J Buring and I De Vivo ldquoCommongenetic variationwithin IGFI IGFII IGFBP-1 and IGFBP-3 andendometrial cancer riskrdquo Gynecologic Oncology vol 120 no 2pp 174ndash178 2011

[8] J Pavelic B Radakovic and K Pavelic ldquoInsulin-like growthfactor 2 and its receptors (IGF 1R and IGF 2Rmannose6-phosphate) in endometrial adenocarcinomardquo GynecologicOncology vol 105 no 3 pp 727ndash735 2007

[9] E Marshman and C H Streuli ldquoInsulin-like growth factorsand insulin-like growth factor binding proteins in mammarygland functionrdquo Breast Cancer Research vol 4 no 6 pp 231ndash239 2002

[10] C Deal J Ma F Wilkin et al ldquoNovel promoter polymorphismin insulin-like growth factor-binding protein-3 Correlationwith serum levels and interaction with known regulatorsrdquoTheJournal of Clinical Endocrinology amp Metabolism vol 86 no 3pp 1274ndash1280 2001

[11] J L Weber and P E May ldquoAbundant class of human DNApolymorphisms which can be typed using the polymerase chainreactionrdquo American Journal of Human Genetics vol 44 no 3pp 388ndash396 1989

[12] J Ma M N Pollak and E Giovannucci ldquoProspective study of356 colorectal cancer risk in men and plasma levels of insulin-like growth factos IGF-1 and IGF binding protein-3rdquo JNCIJournal of the National Cancer Institute vol 91 no 23 pp 620ndash625 1999

[13] Embrapa Fundamentos teorico-praticos e protocolos deextracao e de amplificacao de DNA por meio da tecnicade reacao em cadeia da polimerase Sao Carlos Jun 2007Avaliable from httpwwwcppseembrapabrservicospublic-acaogratuitae-booksLVFundDNACited 18 January 2018

[14] T O Keku A Vidal S Oliver et al ldquoGenetic variants in IGF-IIGF-II IGFBP-3 and adiponectin genes and colon cancer riskin African Americans and Whitesrdquo Cancer Causes amp Controlvol 23 no 7 pp 1127ndash1138 2012

[15] E Feik A Baierl B Hieger et al ldquoAssociation of IGF1 andIGFBP3 polymorphisms with colorectal polyps and colorectalcancer riskrdquo Cancer Causes amp Control vol 21 no 1 pp 91ndash972010

[16] M Harrela H Koistinen J Kaprio et al ldquoGenetic and environ-mental components of interindividual variation in circulatinglevels of IGF-I IGF-II IGFBP-1 and IGFBP-3rdquo The Journal ofClinical Investigation vol 98 no 11 pp 2612ndash2615 1996

[17] I Rietveld J A M J L Janssen E F C Van Rossum et al ldquoApolymorphic CA in the IGF-I gene is associated with gender-specific differences in body height but has no effect on thesecular trend in body heightrdquoClinical Endocrinology vol 61 no2 pp 195ndash203 2004

[18] E-M Rutanen ldquoInsulin-like growth factors and insulin-likegrowth factor binding proteins in the endometrium Effect ofintrauterine levonorgestrel deliveryrdquoHuman Reproduction vol15 no 3 pp 173ndash181 2000

[19] M Nagamani C A Stuart P A Dunhardt and M G DohertyldquoSpecific binding sites for insulin and insulin-like growth factorI in human endometrial cancerrdquo American Journal of Obstetricsamp Gynecology vol 165 no 6 pp 1865ndash1871 1991

[20] T Zou A S Fleisher D Kong et al ldquoSequence alterations ofinsulin-like growth factor binding protein 3 in neoplastic andnormal gastrointestinal tissuesrdquoCancer Research vol 58 no 21pp 4802ndash4804 1998

[21] L M Morimoto P A Newcomb E White J Bigler and J DPotter ldquoVariation in plasma insulin-like growth factor-1 andinsulin-like growth factor binding protein-3 Genetic factorsrdquoCancer Epidemiology Biomarkers amp Prevention vol 14 no 6pp 1394ndash1401 2005

BioMed Research International 5

Table 4 Results of interactions between IGF-1 CA(n) and IGFBP3 rs2854746

Interaction IGFBP3 e IGF1 Control Polyp OR 95 CI P-ValueCA(lt19)CA(n) + CC 16 (203) 11 (108)CA(lt19)CA(n) + CG 5 (63) 2 (2) 0581 009-355 0557CA(lt19)CA(n) + GG 2 (25) 3 (29) 2181 031-1528 0432CA(19)CA(19)+ CA(gt19)CA(n) + GG 7 (89) 15 (147) 3116 095-1015 0059CA(19)CA(19)+ CA(gt19)CA(n) + CC 16 (203) 47 (461) 4272 164-1109 0002CA(19)CA(19)+ CA(gt19)CA(n) + CG 33 (418) 24 (235) 1057 041-268 0905No answer 2 2Total 81 104

Table 5 Multivariate logistic regression between the IGF-1 CA(n)and IGFBP3 rs2854746 polymorphisms adjusted for hypertensionage range and hormone replacement therapy (HRT) use

OR 95 CI P-ValueHRT

nao 1Sim 02462 (0096-0631) 0003

Arterial hypertensionnao 1sim 08201 (0341-1968) 0657

Age range29-39 140-49 02012 (0042-0946) 004250-59 00326 (0007-0143) 000560-80 01387 (0029-0647) 0012

IGF-1 CA(19)CA(lt19)CA(n) 1CA(19)CA(19) +CA(gt19)CA(n) 37191 (1380-10020) 0009

IGFBP3CC 1CG 01607 (0063-0409) 0001GG 06648 (0192-2298) 0518

admixture and (3) significant differences in confounders(eg age hormonal therapy use and high blood pressure)between control and study groups Women from our studygroup were younger with lower prevalence of high bloodpressure and hormonal therapy use thanwomen from controlgroup However it should be noted that results from mul-tiple logistic regression showed no influence of high bloodpressure on the association between variants of the studiedpolymorphisms and endometrial polyp risk The strength ofour study is that controls were known to be polyp-free andwith no history of endometrial diseases at the time of bloodsampling Our decision to select postmenopausal women ascontrol group was based on their lifetime exposure to riskfactors for endometrial diseaseswithout developing themWespeculate that women with this profile might have some kindof protection against endometrial polyp risk factors

Themultiple logistic regressions showed protective influ-ence of the advancement of age with endometrial polypbecause our control patients were all menopausal and there-fore this may have interfered with this result Somethingsimilar occurs with the use of hormone replacement therapywhere the majority of patients who used the therapy werefrom the control group and thismade the therapy a protectiveeffect in relation to the endometrial polyp

The strength of our study is that controls were knownto be polyp-free and with no history of endometrial dis-eases at the time of blood sampling Our decision to selectpostmenopausal women as control group was based on theirlifetime exposure to risk factors for endometrial diseaseswithout developing them We speculate that women withthis profile might have some kind of protection againstendometrial polyp risk factors

Our study reinforces the importance of polymorphism inthe genesis of the endometrial polyp and genetic variabilitygains more force as an important risk factor The clinicalimportance of this study is that with it we can say that somepolymorphisms are considered risk factors for endometrialpolyp

The results suggest that some genotypes of the IGF-1 CA(n) polymorphism have a risk ratio for endometrialpolyp However some genotypes of the IGFBP3 polymor-phism rs2854746 are inversely related to endometrial polypTherefore it is possible to consider them as risk or protectionfactors according to the genotype expression in question

Data Availability

The data used to support the findings of this study areavailable from the corresponding author upon request

Conflicts of Interest

The authors declare that they have no conflicts of interest andnothing to disclose

Authorsrsquo Contributions

Pedro Leopoldo Silva Doria Thomas Moscovitz MarcosTcherniakovsky and Angela Van Nimwegen contributedequally to this work Cesar Eduardo Fernandes Luciano

6 BioMed Research International

Melo Pompei Milton Wajman and Sergio Haimovich alsocontributed equally to this work

Acknowledgments

We would like to thank all the academic colleagues andpartners of the laboratory who collaborated directly in thedevelopment and creation of this scientific work

References

[1] M TMazur andR J KurmanDiagnosis of Endometrial Biopsiesand Curettings Springer Publishing Company NY USA 1995

[2] EDreisler S Stampe Sorensen PH Ibsen andG Lose ldquoPreva-lence of endometrial polyps and abnormal uterine bleeding in aDanish population aged 20ndash74 yearsrdquo Ultrasound in Obstetricsamp Gynecology vol 33 no 1 pp 102ndash108 2009

[3] M J Arends ldquoApoptosis in the endometriumrdquo Histopathologyvol 35 no 2 pp 174ndash178 1999

[4] H Yu and T Rohan ldquoRole of the insulin-like growth factorfamily in cancer development and progressionrdquo Journal of theNational Cancer Institute vol 92 no 18 pp 1472ndash1489 2000

[5] M Pollak ldquoInsulin-like growth factor physiology and cancerriskrdquo European Journal of Cancer vol 36 no 10 pp 1224ndash12282000

[6] N Vaessen P Heutink J A Janssen et al ldquoA polymorphismin the gene for IGF-I functional properties and risk for type 2diabetes and myocardial infarctionrdquo Diabetes vol 50 no 3 pp637ndash642 2001

[7] M McGrath I-M Lee J Buring and I De Vivo ldquoCommongenetic variationwithin IGFI IGFII IGFBP-1 and IGFBP-3 andendometrial cancer riskrdquo Gynecologic Oncology vol 120 no 2pp 174ndash178 2011

[8] J Pavelic B Radakovic and K Pavelic ldquoInsulin-like growthfactor 2 and its receptors (IGF 1R and IGF 2Rmannose6-phosphate) in endometrial adenocarcinomardquo GynecologicOncology vol 105 no 3 pp 727ndash735 2007

[9] E Marshman and C H Streuli ldquoInsulin-like growth factorsand insulin-like growth factor binding proteins in mammarygland functionrdquo Breast Cancer Research vol 4 no 6 pp 231ndash239 2002

[10] C Deal J Ma F Wilkin et al ldquoNovel promoter polymorphismin insulin-like growth factor-binding protein-3 Correlationwith serum levels and interaction with known regulatorsrdquoTheJournal of Clinical Endocrinology amp Metabolism vol 86 no 3pp 1274ndash1280 2001

[11] J L Weber and P E May ldquoAbundant class of human DNApolymorphisms which can be typed using the polymerase chainreactionrdquo American Journal of Human Genetics vol 44 no 3pp 388ndash396 1989

[12] J Ma M N Pollak and E Giovannucci ldquoProspective study of356 colorectal cancer risk in men and plasma levels of insulin-like growth factos IGF-1 and IGF binding protein-3rdquo JNCIJournal of the National Cancer Institute vol 91 no 23 pp 620ndash625 1999

[13] Embrapa Fundamentos teorico-praticos e protocolos deextracao e de amplificacao de DNA por meio da tecnicade reacao em cadeia da polimerase Sao Carlos Jun 2007Avaliable from httpwwwcppseembrapabrservicospublic-acaogratuitae-booksLVFundDNACited 18 January 2018

[14] T O Keku A Vidal S Oliver et al ldquoGenetic variants in IGF-IIGF-II IGFBP-3 and adiponectin genes and colon cancer riskin African Americans and Whitesrdquo Cancer Causes amp Controlvol 23 no 7 pp 1127ndash1138 2012

[15] E Feik A Baierl B Hieger et al ldquoAssociation of IGF1 andIGFBP3 polymorphisms with colorectal polyps and colorectalcancer riskrdquo Cancer Causes amp Control vol 21 no 1 pp 91ndash972010

[16] M Harrela H Koistinen J Kaprio et al ldquoGenetic and environ-mental components of interindividual variation in circulatinglevels of IGF-I IGF-II IGFBP-1 and IGFBP-3rdquo The Journal ofClinical Investigation vol 98 no 11 pp 2612ndash2615 1996

[17] I Rietveld J A M J L Janssen E F C Van Rossum et al ldquoApolymorphic CA in the IGF-I gene is associated with gender-specific differences in body height but has no effect on thesecular trend in body heightrdquoClinical Endocrinology vol 61 no2 pp 195ndash203 2004

[18] E-M Rutanen ldquoInsulin-like growth factors and insulin-likegrowth factor binding proteins in the endometrium Effect ofintrauterine levonorgestrel deliveryrdquoHuman Reproduction vol15 no 3 pp 173ndash181 2000

[19] M Nagamani C A Stuart P A Dunhardt and M G DohertyldquoSpecific binding sites for insulin and insulin-like growth factorI in human endometrial cancerrdquo American Journal of Obstetricsamp Gynecology vol 165 no 6 pp 1865ndash1871 1991

[20] T Zou A S Fleisher D Kong et al ldquoSequence alterations ofinsulin-like growth factor binding protein 3 in neoplastic andnormal gastrointestinal tissuesrdquoCancer Research vol 58 no 21pp 4802ndash4804 1998

[21] L M Morimoto P A Newcomb E White J Bigler and J DPotter ldquoVariation in plasma insulin-like growth factor-1 andinsulin-like growth factor binding protein-3 Genetic factorsrdquoCancer Epidemiology Biomarkers amp Prevention vol 14 no 6pp 1394ndash1401 2005

6 BioMed Research International

Melo Pompei Milton Wajman and Sergio Haimovich alsocontributed equally to this work

Acknowledgments

We would like to thank all the academic colleagues andpartners of the laboratory who collaborated directly in thedevelopment and creation of this scientific work

References

[1] M TMazur andR J KurmanDiagnosis of Endometrial Biopsiesand Curettings Springer Publishing Company NY USA 1995

[2] EDreisler S Stampe Sorensen PH Ibsen andG Lose ldquoPreva-lence of endometrial polyps and abnormal uterine bleeding in aDanish population aged 20ndash74 yearsrdquo Ultrasound in Obstetricsamp Gynecology vol 33 no 1 pp 102ndash108 2009

[3] M J Arends ldquoApoptosis in the endometriumrdquo Histopathologyvol 35 no 2 pp 174ndash178 1999

[4] H Yu and T Rohan ldquoRole of the insulin-like growth factorfamily in cancer development and progressionrdquo Journal of theNational Cancer Institute vol 92 no 18 pp 1472ndash1489 2000

[5] M Pollak ldquoInsulin-like growth factor physiology and cancerriskrdquo European Journal of Cancer vol 36 no 10 pp 1224ndash12282000

[6] N Vaessen P Heutink J A Janssen et al ldquoA polymorphismin the gene for IGF-I functional properties and risk for type 2diabetes and myocardial infarctionrdquo Diabetes vol 50 no 3 pp637ndash642 2001

[7] M McGrath I-M Lee J Buring and I De Vivo ldquoCommongenetic variationwithin IGFI IGFII IGFBP-1 and IGFBP-3 andendometrial cancer riskrdquo Gynecologic Oncology vol 120 no 2pp 174ndash178 2011

[8] J Pavelic B Radakovic and K Pavelic ldquoInsulin-like growthfactor 2 and its receptors (IGF 1R and IGF 2Rmannose6-phosphate) in endometrial adenocarcinomardquo GynecologicOncology vol 105 no 3 pp 727ndash735 2007

[9] E Marshman and C H Streuli ldquoInsulin-like growth factorsand insulin-like growth factor binding proteins in mammarygland functionrdquo Breast Cancer Research vol 4 no 6 pp 231ndash239 2002

[10] C Deal J Ma F Wilkin et al ldquoNovel promoter polymorphismin insulin-like growth factor-binding protein-3 Correlationwith serum levels and interaction with known regulatorsrdquoTheJournal of Clinical Endocrinology amp Metabolism vol 86 no 3pp 1274ndash1280 2001

[11] J L Weber and P E May ldquoAbundant class of human DNApolymorphisms which can be typed using the polymerase chainreactionrdquo American Journal of Human Genetics vol 44 no 3pp 388ndash396 1989

[12] J Ma M N Pollak and E Giovannucci ldquoProspective study of356 colorectal cancer risk in men and plasma levels of insulin-like growth factos IGF-1 and IGF binding protein-3rdquo JNCIJournal of the National Cancer Institute vol 91 no 23 pp 620ndash625 1999

[13] Embrapa Fundamentos teorico-praticos e protocolos deextracao e de amplificacao de DNA por meio da tecnicade reacao em cadeia da polimerase Sao Carlos Jun 2007Avaliable from httpwwwcppseembrapabrservicospublic-acaogratuitae-booksLVFundDNACited 18 January 2018

[14] T O Keku A Vidal S Oliver et al ldquoGenetic variants in IGF-IIGF-II IGFBP-3 and adiponectin genes and colon cancer riskin African Americans and Whitesrdquo Cancer Causes amp Controlvol 23 no 7 pp 1127ndash1138 2012

[15] E Feik A Baierl B Hieger et al ldquoAssociation of IGF1 andIGFBP3 polymorphisms with colorectal polyps and colorectalcancer riskrdquo Cancer Causes amp Control vol 21 no 1 pp 91ndash972010

[16] M Harrela H Koistinen J Kaprio et al ldquoGenetic and environ-mental components of interindividual variation in circulatinglevels of IGF-I IGF-II IGFBP-1 and IGFBP-3rdquo The Journal ofClinical Investigation vol 98 no 11 pp 2612ndash2615 1996

[17] I Rietveld J A M J L Janssen E F C Van Rossum et al ldquoApolymorphic CA in the IGF-I gene is associated with gender-specific differences in body height but has no effect on thesecular trend in body heightrdquoClinical Endocrinology vol 61 no2 pp 195ndash203 2004

[18] E-M Rutanen ldquoInsulin-like growth factors and insulin-likegrowth factor binding proteins in the endometrium Effect ofintrauterine levonorgestrel deliveryrdquoHuman Reproduction vol15 no 3 pp 173ndash181 2000

[19] M Nagamani C A Stuart P A Dunhardt and M G DohertyldquoSpecific binding sites for insulin and insulin-like growth factorI in human endometrial cancerrdquo American Journal of Obstetricsamp Gynecology vol 165 no 6 pp 1865ndash1871 1991

[20] T Zou A S Fleisher D Kong et al ldquoSequence alterations ofinsulin-like growth factor binding protein 3 in neoplastic andnormal gastrointestinal tissuesrdquoCancer Research vol 58 no 21pp 4802ndash4804 1998

[21] L M Morimoto P A Newcomb E White J Bigler and J DPotter ldquoVariation in plasma insulin-like growth factor-1 andinsulin-like growth factor binding protein-3 Genetic factorsrdquoCancer Epidemiology Biomarkers amp Prevention vol 14 no 6pp 1394ndash1401 2005

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