NeuroToxicology 30 (2009) Contents lists available at ScienceDirect. NeuroToxicology

Transcription

1 NeuroToxicology 30 (2009) Contents lists vilble t ScienceDirect NeuroToxicology Anlysis of PCB congeners relted to cognitive functioning in dolescents Jon Newmn, *, Mi V. Gllo b, Lwrence M. Schell b,c, Anthony P. DeCprio d, Melind Denhm e, Glenn D. Dene f, the Akwessne Tsk Force on the Environment g University t Albny, Deprtment of Eductionl nd Counseling Psychology, Eduction 236, 1400 Wshington Avenue, Albny, NY 12222, United Sttes b University t Albny, Deprtment of Anthropology, A&S 237, 1400 Wshington Avenue, Albny, NY, United Sttes c University t Albny, Deprtment of Epidemiology nd Biosttistics, School of Public Helth One University Plce, Room 131, Rensseler, NY, United Sttes d Florid Interntionl University, Deprtment of Chemistry nd Biochemistry, Mimi, FL 33199, United Sttes e Professionl Development Progrm, Rockefeller College, University t Albny, United Sttes f University t Albny, Deprtment of Sociology, A&S 339, 1400 Wshington Avenue, Albny, NY 12222, United Sttes g Akwessne Mohwk Ntion ARTICLE INFO ABSTRACT Article history: Received 2 June 2008 Accepted 8 My 2009 Avilble online 22 My 2009 Keywords: Polychlorinted biphenyls Congeners Cognition Memory Adolescents Mohwk To investigte the chrcteristics of PCBs tht re linked to cognitive functioning, those congeners tht were concurrently found in 271 Mohwk dolescents were grouped ccording to structure (dioxin-like or non-dioxin-like) nd persistence (persistent or low-persistent). After the effects of the congener groups were orthogonlized, regression nlyses (controlling for number of vribles found to be relted to the cognitive outcomes) exmined the reltionship of ech congener group to scores on three cognitive tests (the non-verbl Rvens Progressive Mtrices, the Test of Memory nd Lerning, nd the Woodcock Johnson-Revised). Five subtests from these cognitive tests were found to be ssocited with one or more PCB congener groups, most often t moderte level. Two mesures of long-term memory (Delyed Recll nd Long Term Retrievl) were ssocited with ll four congener groups. Nevertheless, exmintion of the role of individul congeners in the significntly relted congener groups reveled tht lmost ll congeners ssocited with cognitive outcomes were non-dioxin-like nd ortho-substituted. A notble exception ws the Rvens test where scores were ssocited only with dioxin-like congeners. This finding dds to the limited evidence of neurotoxic effects of dioxin-like congeners. Auditory Processing ws relted only to the persistent congener group. The ssocition of the non-persistent congener group with three cognitive test scores (Delyed Recll, Long Term Retrievl nd Comprehension-Knowledge) suggests tht the Mohwk dolescents hve experienced continuing or recent environmentl exposure to PCBs tht is sufficient to result in detectble cognitive decrements. Comprison of our findings with those of other humn studies ws limited by the reltive lck of specificity of both PCB mesures nd cognitive outcome mesures in much previous work. Published by Elsevier Inc. 1. Introduction Numerous studies hve reported negtive effects ssocited with prentl or perintl exposure to polychlorinted biphenyls (PCBs) on spects of cognitive functioning in children (Jcobson nd Jcobson, 1996; Ptndin et l., 1999; Vreugdenhil et l., 2002, 2004; Wlkowik et l., 2001). Concurrent PCB body burden hs been negtively ssocited with memory in dolescents (Newmn et l., 2006), memory nd lerning in older dults (Schntz et l., 2001), nd ttention performnce in dults (Peper et l., 2005). * Corresponding uthor. Tel.: ; fx: E-mil ddresses: jonewmn@ol.com, jnewmn@umil.lbny.edu (J. Newmn). However there is considerble vrition in findings of different studies tht my hve severl sources. Vrition could be due to the prticulr congeners comprising the different exposures in studies of PCBs nd cognition. PCBs include 209 chemicl species (congeners) tht differ in the extent nd pttern of chlorintion of the biphenyl structure (Agency for Toxic Substnces nd Disese Registry, 2000). At ny single time point, PCB body burden is function of: (1) the subject s pst nd recent exposure history nd (2) the subject s individul determinnts of uptke, metbolism, nd clernce, i.e., toxicokinetics (Frme, 1997; Glden et l., 2003; Glynn et l., 2003; Hnsen, 1998; Jmes et l., 2002; Krmus et l., 2001). Biologicl persistence of individul congeners nd, consequently, their reltive contribution to the body burden, vries with structure (Brown, 1994; Sfe, 1994). Higher chlorinted congeners (i.e., those with six or more chlorines) nd those with pr- (i.e., 4,4 0 -) chlorine ring substitution re, in generl, metbolized slowly in X/$ see front mtter. Published by Elsevier Inc. doi: /j.neuro

2 J. Newmn et l. / NeuroToxicology 30 (2009) humns nd typiclly mke up the bulk of body burden. However, recent PCB exposure will lso result in the presence of less persistent congeners in tissue. Pst exposure to lbile congeners my lso result in effects tht continue to be demonstrted (Hnsen, 1998). Typiclly in humn studies, PCB congeners re grouped, either s totl PCBs or the sum of severl commonly found congeners, or re indexed by single congener, usully PCB 153 (Longnecker et l., 2003). However, studies using summtive PCB mesure do not revel which types of congeners re influentil, nd wht fetures of these congeners re linked to effects. Despite the nlytic cpbility of identifying prticulr PCB congeners, humn studies hve not studied groups of congeners defined by structure or toxicologicl ction in prt becuse the humn neurotoxic mechnisms of PCB ction re not well understood t this time. In one of the few studies to consider specific PCB congeners, no pttern could be detected in the congeners found to be relted to neurologicl optimlity deficits t dys in humns (Huismn et l., 1995). Unlike the predominnt neurochemicl findings (Seegl et l., 1990; Tilson nd Kodvnti, 1998; Wong et l., 2001) behviorl evidence in experimentl nimls hs demonstrted physicl nd lerning effects for both ortho-substituted nd dioxin-like PCB congeners, with no cler pttern (Holene et l., 1998; Tilson et l., 1979). Studying the effects of specific PCB congeners of known structure or hypothesized toxicologicl effects would help clrify sources of inconsistency in the findings of the humn studies. In some cohorts, negtive ssocitions hve not been found (Dniels et l., 2003; Grndjen et l., 2001; Gry et l., 2005; Longnecker et l., 2004; Vermeir et l., 2005; Vreugdenhil et l., 2002), nd in others pper to be trnsient (Stewrt et l., 2003; Winneke et l., 2005) or delyed (Wlkowik et l., 2001). Further inconsistency between study findings my rise from differences in the cognitive outcome mesures employed. Study results would vry not only ccording to the PCB congeners evident in the smple of prticipnts, but lso the prticulr neuropsychologicl processes involved in the outcome mesures employed. Humn studies hve included vrious mesures of developmentl or psychologicl functioning, ccording to the ge of the prticipnts, the tests vilble in the yers testing ws conducted, nd experimenter focus. While mny of the tests provide globl mesure such s IQ, the tests used re not functionlly equivlent nd do not comprise the sme number, type or specificity of psychologicl processes. Moreover, the processes involved in psychologicl functioning my chnge with ge nd my be t risk of disruption by PCBs t different ges (Wlkowik et l., 2001). For ll of these resons, reserchers hve clled for more investigtion of specific congener effects (Rice, 2005; Schntz et l., 2003). Such congener specific nlyses would not only increse understnding of reltionships nd the processes by which observed effects occur they would inform recommendtions of public helth gencies concerning the trgeting nd urgency of remedition, ccording the known effects of the congeners prevlent in n re (Rice, 2005). In the present pper we reexmine the cognitive outcomes found to be ssocited with the summtive mesure of PCBs used in our erlier nlysis (Newmn et l., 2006). In tht nlysis we summed ll 16 PCB congeners with rte of detection in the dolescent smple of 50% or greter into single mesure, nd found it to be relted to two seprte mesures of long-term memory, s well s Comprehension-Knowledge. In the current pper we test hypotheses bout the effects of congener chrcteristics on cognitive outcomes by grouping congeners for re-nlysis ccording to their structurl similrity (dioxin-like nd nondioxin-like), nd their known persistence. Becuse of the high correltion between the congener groups creted we trnsformed the grouped PCB vribles to orthogonlize their effects. Mny of the published findings (e.g. Vreugdenhil et l., 2004; Winneke et l., 2005) hve included commonly found congeners known to be of high-persistence (138, 153, 180 nd sometimes 118). We lso investigte the role of subgroup of low-persistence, non-dioxinlike congeners, to determine if either recent episodic or chronic continuing exposure negtively impcts cognitive functioning in the dolescents (Brown, 1994; Hnsen, 1998). If these congener groups show ssocitions with cognitive outcomes, further nlysis of the ssocitions of specific congeners comprising the groups is wrrnted. The bttery of tests employed provided more specificity bout cognitive outcomes thn do most other humn studies, yet ddresses cognitive domins tht hve been ssocited with exposure in these studies. 2. Methods 2.1. Smple nd loction The Mohwk Adolescent Well-Being Study (MAWBs) ws conducted in the yers in prtnership with residents of the Akwessne Mohwk Ntion, sovereign territory lying on both sides of the St. Lwrence River nd spnning the boundries of New York Stte, nd Ontrio nd Quebec, Cnd. Severl industril complexes re in close proximity to Akwessne, including Ntionl Priority Superfund Site (Generl Motors Centrl Foundry Division), nd two New York Stte Superfund Sites (Reynolds Metl Compny nd Aluminum Compny of Americ). Study protocols nd methods hve previously been described in detil (Newmn et l., 2006; Schell et l., 2003), nd re briefly reviewed here. All study protocols were pproved by the Institutionl Review Bord t the University t Albny, Stte University of New York, nd informed consent ws obtined from ll prticipnts. Two hundred nd seventy-one mother dolescent pirs, who met eligibility criteri, enrolled in nd completed the study. To be eligible, the dolescent must hve: (1) been yers 11 months old, (2) been singleton birth, (3) never been hospitlized with brin injury, (4) hd no history of serious orgnic or psychologicl pthology s determined by physicin or psychologist, nor (5) dignosis of Fetl Alcohol Syndrome or Fetl Alcohol Effects. Forty-eight percent of the dolescents were mles Procedures Trined dt collectors, who were members of the Akwessne community nd hd no prior knowledge of prticipnts toxicnt levels, conducted ll dt collection, including cognitive testing nd blood drws, nd mternl interviewing. Dt collectors received extensive trining t the University t Albny nd on-site, both before the study begn nd regulrly throughout the project. Videotpes of the cognitive testing were periodiclly reviewed in order to mintin stndrdized procedures. Fsting blood drws (15 ml) were tken in the dolescent s home upon his/her first rising in the morning. Assessment of cholesterol nd triglycerides ws performed t the Clinicl Chemistry nd Hemtology Lbortory, Wdsworth Center for Lbortories nd Reserch, New York Stte Deprtment of Helth. The fcility is CLIA-pproved nd member of the CDC reference lbortory network for lipid mesurements (Myers et l., 2000). Serum lipid concentrtions were mesured on Hitchi 911 nlyzer (Roche Dignostics, Indinpolis, IN) using cholesterol esterse nd oxidse/peroxidse method for totl cholesterol (Allin et l., 1974) nd glycerol kinse-bsed procedure tht corrects for free glycerol in the specimen (Kohlmeier, 1986) for triglycerides.

3 688 J. Newmn et l. / NeuroToxicology 30 (2009) Toxicnts Anlyses of PCBs nd orgnochlorine pesticides (p,p 0 -DDE, HCB), nd mirex were conducted t the University t Albny s Exposure Assessment Lbortory. High resolution, ultrtrce, congenerspecific nlysis ws performed by prllel dul-column (splitless injection) gs chromtogrphy with electron cpture detection (DeCprio et l., 2000). This method quntittes up to 83 individul PCB congeners nd 18 PCB congeners s pirs or triplets. Led nd mercury nlyses were conducted by Le Centre de Toxicologie due Quebec in Sinte-Foy, Quebec Cnd. Led ws nlyzed by Zeemn-corrected grphite furnce tomic bsorption spectrometry (Prsons nd Slvin, 1993), nd mercury by cold-vpor tomic bsorption spectrometry. The method recommended by the U.S. Environmentl Protection Agency (U.S. Environmentl Protection Agency, 1998) ws used to impute vlues below the method detection limits (mdl) for ll toxicnts with rtes of detection of 50% or greter (see Schell et l., 2003). The lgorithm clcultes vlue depending on the mdl nd the rte of detection of ech toxicnt (Cohen, 1950; Gupt, 1952). This method ws lso used for ll imputtions of nondetected led, mercury, HCB, nd mirex levels. It ws not used for p,p 0 -DDE s ll smples hd detectble levels. For those congeners detected in less thn 50% of the smple, the most common prctice for imputtion ws pplied by replcing ny individul dtum tht ws below the mdl with the midpoint vlue between zero nd the mdl of ech nlyte or congener Congener groups PCBs re indicted by IUPAC numbers throughout. Sixteen PCB congeners with rtes of detection of 50% or greter re included in the present nlysis ( P PCBs50% = PCBs 118, 101[+90], 153, 110, 99, 87, 138 [ ], 180, 95, 52, 74, 105, 149 [+123], 187, 70, 84). Specific congeners were lso grouped ccording to structure: (1) PCB congeners re defined s dioxin-like or non-dioxin-like bsed upon the criteri used by the World Helth Orgniztion nd other regultory gencies nd (2) high- nd low-persistence groupings. This resulted in (1) dioxin-like (PCBs 105, 118, 149[+123]); (2) non-dioxin-like (PCBs 52, 70, 84, 74, 87, 95, 99, 101[+90], 110, 138[ ], 153, 180, 187); (3) highly persistent grouping (PCBs 74, 99, 105, 118, 138[ ], 153, 180, 187); nd (4) low-persistence grouping which includes tetr- or pent-pcbs tht re ortho-substituted nd tht hve t most one pr-chlorine (PCBs 52, 70, 84, 95, 110, 101[+90], 87). The group of dioxin-like congeners does not contin the more potent dioxin-like congeners (Vn den Berg et l., 2006) s they were not mesurble with the nlytic method employed. The lowpersistence grouping of congeners (ll of which re non-dioxinlike) ws found to include the most potent reducers of dopmine concentrtions in cell cultures (Shin et l., 1991). The congeners within specific group were combined quntittively by simple summtion for this nlysis. Similr groupings were creted with PCB congeners detected in t lest 10% of the smple, nd these were lso tested for ny reltionships with the outcomes of interest Cognitive tests The Woodcock Johnson-Revised Tests of Cognitive Ability, Stndrd nd Supplementl Btteries (Woodcock nd Johnson, 1990), or WJ-R, yields mesure of overll cognitive functioning (Brod Cognitive Ability Extended, BCAE), s well s seven cluster scores tht provide informtion regrding n individul s bilities in specific domins of cognition. These domins re Long Term Retrievl (LTR), Short Term Memory (STM), Processing Speed (PSD), Auditory Processing (APG), Visul Processing (VPG), Comprehension-Knowledge (CKN), nd Fluid Resoning (FRG). The WJ-R hs good psychometric properties, with stted vlidity of , nd relibility of Adolescents were dministered both the Stndrd nd Supplementry Btteries, wheres mothers were dministered only the Stndrd Bttery. The Rvens Progressive Mtrices (RPM), Stndrd Form (Rven et l., 1992) is mesure of intellectul bility nd resoning skills. The Rvens ssesses the individul s bility to discern ptterns nd derive mening from complex dt. The Rvens is thought to be culturlly fir becuse it is non-verbl mesure, i.e., lnguge is not utilized in the testing mterils or dministrtion. Furthermore, the Rvens hs been normed on individuls of different culturl bckgrounds. The Test of Memory nd Lerning (TOMAL) (Reynolds nd Bigler, 1994) is mesure of generl nd specific memory functioning. The TOMAL yields Verbl Memory Index (VMIS), Non-verbl Memory Index (NMIS), Delyed Recll Index (DRIS), nd Composite Memory Index (CMIS) Sttisticl nlysis All substntive sttisticl nlyses were performed using SPSS version (SPSS, 2001). For the multivrite nlysis, list of potentil confounders ws generted from mesured fctors tht previous reserch suggests re likely to be relted to cognitive functioning. These were sex, ge (yers), the dolescent s body mss index (BMI), cholesterol nd triglyceride levels, totl cloric, ft nd protein intke, mternl BMI, mternl Brod Cognitive Ability, weighted index of socioeconomic sttus (which includes mternl eduction, mternl employment (current, full/prt time), mternl mritl sttus, size nd condition of the house, number of motor vehicles nd ge of newest vehicle within the household), the number of socil problems (i.e., helth, reltive, legl, mritl, drinking, drug use, etc.) within the household, grvidity, number of cigrettes the mother smoked per dy during pregnncy, durtion of brestfeeding in weeks, nd levels of other toxicnts (HCB, DDE, mirex, blood led, nd mercury). Those selected from this list for inclusion in the multiple regression models were selected empiriclly, bsed on the inclusive decision rule of bivrite correltion of p < 0.2 with one or more cognitive scores, s hs previously been described (Newmn et l., 2006). Vribles with skewed distributions were log trnsformed (led, mercury, PCBs, p,p 0 -DDE, HCB, brestfeeding durtion (weeks), nd number of cigrettes mothers smoked per dy during pregnncy). Mirex levels were ctegorized into three groups becuse of the high rte of nondetectble levels: nondetects (below the minimum detection limit of 0.02 ppb; 54.2%), low detects ( ppb; 18.1% of the smple), nd high detects ( ppb; 27.7% of the smple). Our objective of simultneously ssessing the effect of dioxinlike congener group nd non-dioxin-like congener group, nd highly persistent grouping simultneously with low-persistence congener group, invokes the common but vexing dt nlytic condition of multicollinerity. The problems nd tretments of multicollinerity hve been investigted by mny reserchers nd it is stndrd topic in ll introductions to multiple regression. Strong bivrite correltions between PCB groupings (shown in Tble 3) nd much weker correltions between ech cognitive score nd PCB groupings (reported in Tble 6) substntite our concern nd indicte ll of the usul conditions ssocited with multicollinerity nd prtilling fllcy (Gordon, 1968). Mnging the intercorreltion of risk fctors presents formidble chllenge becuse it is difficult to ccurtely disentngle nd rnk the contributions of individul fctors deemed importnt to the subject of study. Our solution is to orthogonlize

4 J. Newmn et l. / NeuroToxicology 30 (2009) our congener groupings, creting new sets of orthogonl vribles, using modified Grm-Schmidt procedure (Golub nd Vn Lon, 1996) vilble s the orthog procedure in the Stt softwre pckge (SttCorp, 2007). Given tht the order (tht is, the conceptul priority) of the vribles determines the orthogonliztion, we generte four orthogonl trnsformtions: one in which our dioxin-like congener group precedes our non-dioxinlike group, then vice vers; third in which our highly persistent grouping precedes our low-persistence congener group, then vice vers. Orthogonl vribles produce numericl ccurcy for highly colliner vribles nd they re esy to interpret. In essence, the trnsformtions residulize the vribles ccording to their specified order. With two vribles, the first vrible retins ll of its vrition (nd therefore ll of its covrition with the dependent vrible), while the second vrible retins only its residul vrition tht remins fter removing its covrition with the first vrible. The orthogonl vribles re then scled s stndrd norml vribles with zero mens nd unit vrinces (nd stndrd devitions) so tht their metric in substntive multiple regression is in stndrd devition units. 3. Results Tble 1 provides description of the prticipting dolescents, including cognitive test scores. Relevnt mternl scores re lso shown. Prticipting youth rnged in ge from 10 through 16 yers with medin of 13.2 yers, nd on verge showed cognitive functioning similr to comprison dolescents on whom the WJ-R, TOMAL nd Rvens were normed. Forty-eight percent were mles. Grouped PCB nd specific congener levels re shown in Tble 2. Akwessne dolescents PCB levels rnged between 0.05 nd 3.32 ppb, dependent on the congener grouping. p,p 0 -DDE levels were found to be highest of the ll the persistent orgnic pollutnts, with mximum level of 4.80 ppb. Hevy metl levels were consistently low, neither exceeding the CDC threshold. Correltion mong the PCB groupings is shown in Tble 3. All groupings re positively correlted t r = 0.6 or bove. Of the potentil covrites exmined for their reltionship with the cognitive outcomes of interest, bivrite correltions (Tble 4) determined tht the following were relted to t lest one cognitive score t p < 0.2 level, nd so were included in the finl model: sex; ge in yers; dolescent nd mternl BMI (kg/m 2 ); dolescent triglyceride nd cholesterol levels; mternl Brod Cognitive Ability Stndrd score; SES index; number of reported socil problems; grvidity; cigrette use during pregnncy; brestfeeding durtion (in weeks); p,p 0 -DDE; HCB; blood led; mercury; nd mirex. Ech congener group s converse (e.g. nondioxin-like for dioxin-like; low-persistent for highly persistent) ws lso included s covrite in the multivrite nlysis. Multivrite regression models predicting ech cognitive domin were clculted for the four orthogonlly trnsformed PCB congener groups mde up of congeners found in more thn 50% of the smple (Tble 5). The sum of dioxin-like PCBs ws significnt, inverse, predictor of Rvens scores. All congener groupings were consistently nd negtively ssocited with both Long Term Retrievl nd Delyed Recll. The sum of highly persistent PCBs ws inversely nd significntly ssocited with Auditory Processing. The sum of low-persistent PCBs ws significnt, inverse predictor of Comprehension-Knowledge. Verbl Memory, Non-verbl Memory, Composite Memory, Brod Cognitive Ability Extended, Short Term Memory, Processing Speed, Visul Processing, nd Fluid Resoning were not significntly ssocited with ny congener group. The reported results did not differ when clcultions included those less prevlent congeners found in 10% of the smple or more. The effect of n increse of one stndrd devition in the PCB congener group my be expressed s proportion of one stndrd devition of the relevnt cognitive test. Following Cohen s (1988) guidelines for effect sizes tht my be considered smll, moderte or lrge, we found tht most of the significnt reltionships described bove indicte moderte effects of 0.40 or greter. A one Tble 1 Chrcteristics of the smple. Adolescents N Men Medin SD Minimum Mximum Age (yers) BMI (kg/m 2 ) Triglyceride level (mg/dl) Cholesterol level (mg/dl) Rvens (RPM) TOMAL Verbl Memory (VMIS) Non-verbl Memory (NMIS) Delyed Recll (DRIS) Composite Memory (CMIS) Woodcock Johnson-Revised (WJ-R) Brod Cognitive Ability Extended (BCAE) Long Term Retrievl (LTR) Short Term Memory (STM) Processing Speed (PSD) Auditory Processing (APG) Visul Processing (VPG) Comprehension-Knowledge (CKN) Fluid Resoning (FRG) Mothers Mternl BMI (kg/m 2 ) Mternl Brod Cognitive Ability Stndrd Socioeconomic Sttus Index Number of Socil Problems Reported Grvidity Cigrette use during pregnncy (#/dy) Brestfeeding durtion (weeks)

5 690 J. Newmn et l. / NeuroToxicology 30 (2009) Tble 2 PCB nd other toxicnt levels mong Akwessne Mohwk youth. PCBs (ppb) N Structure ROD Dioxin-like (Y/N) Persistence (H/M/L) GM Medin SD Minimum Mximum P Dioxin-like PCBs50% b,c P Dioxin-like PCBs10% d,e P Non-dioxin-like PCBs50% b,f P Non-dioxin-like PCBs10% d,g P Persistent PCBs50% b,h P Persistent PCBs10% d,i P Low-persistent PCBs50% b,j P Low-persistent PCBs10% d,k PCBs >50% IUPAC #b ,3 0,4,4 0,5 99.6% Y H [+90] l 271 2,2 0,3,4 0,5 + 2,2 0 4,5, % N L,M ,2 0,4,4 0,5, % N H ,3,3 0,4 0,6 95.2% N L ,2 0,4,4 0,5 94.5% N H ,2 0,3,4, % N L [ ] l 271 2,2 0,3,4,4 0, ,3,3 0, 90.0% N H,M,L ,5 0,6 + 2,3,3 0,4 0,5, ,2 0,3,4,4 0,5, % N H ,2 0,3,5 0,6 79.3% N L ,2 0,5, % N L ,4,4 0,5 73.4% N M ,3,3 0,4, % Y H [+123] l 271 2,2 0,3,4 0,5 0,6 + 2,3 0,4,4 0, % Y L,H ,2 0,3,4 0,5,5 0,6 57.6% N M ,3 0,4 0,5 55.4% N L ,2 0,3,3 0,6 53.1% N L PCBs >10% <50% IUPAC #d ,2 0,3,3 0,4,4 0,5 45.0% N H ,3 0,4, % N M ,4, % N M ,3,3 0,4,4 0,6 31.7% N M ,4 0,5 31.7% N L ,2 0,3,5, % N L ,2 0,3,3 0,4,5, % N L ,2 0,3,3 0,4,5 0, % N H ,4,5 24.7% N L ,2 0,3,4 0, % N L ,3 0,4 0,6 16.6% N L ,3 0,4, % Y M ,3,3 0,4,4 0,5 14.4% Y H , % N M Other toxicnts b HCB (ppb) % p,p 0 -DDE (ppb) % Mirex (ppb) % Blood Led (mg/dl) % Totl Mercury (mg/dl) % Abbrevitions: PCB, polychlorinted biphenyls; PPB, prts per billion; IUPAC, Interntionl Union of Pure nd Applied Chemistry; GM, geometric men; SD, stndrd devition; persistence: H, high; M, moderte; L, low. Only those congeners tht were found to be significntly correlted with the outcome vribles re shown. b Vlues below the mdl were clculted following the EPA recommended method for estimting non-detected vlues s described in Schell et l. (2003). c P Dioxin-like congeners in >50% of the smple = IUPAC#s 105, 118, 149[+123]. d Vlues below the detection limit hve been replced by the vlue midwy between the detection limit nd zero. e P Dioxin-like congeners in >10% of the smple = IUPAC#s 77, 105, 118, 149[+123], 156. f P Non-dioxin-like congeners in >50% of the smple = IUPAC#s 52, 70, 84, 74, 87, 95, 99, 101[+90], 110, 138[ ], 153, 180, 187. g P Non-dioxin-like congeners in >10% of the smple = IUPAC#s 52, 70, 84, 74, 87, 95, 99, 101[+90], 110, 138[ ], 153, 180, 187, 132, 146, 158, 170, 183, 199, 28, 31, 44, 47[+59], 66, 130, 141, 151, 174, 177, 18, 196, 29, 33, 8, 92, 97, 128, 144, 15, 172, 176, 190, 203, 24[+27], 40, 42, 49, 53, 56, 71. h P Persistent PCBs detected in >50% of the smple = IUPAC#s 74, 99, 105, 118, 138[ ], 153, 180, 187. i P Persistent PCBs detected in >10% of the smple = IUPAC#s 74, 99, 105, 118, 138[ ], 153, 180, 187, 47[+59], 28, 66, 77, 130, 158, 199, 128, 146, 156, 170, 172, 177, 183, 190, 196, 203. j P Low-persistent PCBs detected in >50% of the smple = IUPAC#s 52, 70, 84, 95, 110, 101[+90], 87. k P Low-persistent PCBs detected in >10% of the smple = IUPAC#s 52, 70, 84, 95, 110, 101[+90], 87, 8, 18, 29, 31, 33, 40, 42, 44, 49, 53, 56, 71, 92, 97, 132, 141, 144, 151, 174, 176, 24[+27], 15. l Brcket indictes minor congener bsed on roclor concentrtion (Frme, 1997). stndrd devition increse in the dioxin-like congener group ws ssocited with decreses in Rvens, Delyed Recll nd Long Term Retrievl of 0.32, 0.33 nd 0.35 stndrd devition units respectively. A one stndrd devition increse in the nondioxin-like congener group ws ssocited with decreses of 0.41 nd 0.52 stndrd devition units in Delyed Recll nd Long Term Retrievl respectively. A one stndrd devition increse in the persistent congener group ws ssocited with decreses in Delyed Recll, Long Term Retrievl, nd Auditory Processing of 0.48, 0.49, nd 0.43 stndrd devition units respectively. Finlly, one stndrd devition increse in the low-persistent congener group ws ssocited with decreses of 0.28, 0.41 nd 0.27

6 J. Newmn et l. / NeuroToxicology 30 (2009) Tble 3 Correltion between PCB groupings. P Dioxin-like PCBs50% P Non-dioxin-like PCBs50% P Persistent PCBs50% P Low-persistent PCBs50% P Dioxin-like PCBs50% P Non-dioxin-like PCBs50% P Persistent PCBs50% P Low-persistent PCBs50% Vlues below the mdl were clculted following the EPA recommended method for estimting non-detected vlues s described in Schell et l. (2003). stndrd devition units of Delyed Recll, Long Term Retrievl nd Comprehension-Knowledge respectively. Becuse the regression nlyses showed tht hypothesized congener groups were ssocited with some cognitive outcomes (Rvens, Long Term Retrievl, Delyed Recll, Auditory Processing, Comprehension-Knowledge), bivrite correltions between the individul PCB congeners comprising these groups nd those five cognitive outcomes were lso exmined (Tble 6). (Eight cognitive outcomes were not correlted significntly with ny of the 16 congeners.) Long Term Retrievl ws significntly correlted (p <.05) with seven PCB congeners, more thn ny other cognitive test. Six of these seven were non-dioxin-like (di-ortho-substituted congeners PCBs 87, 99, 101[+90], 110, nd 138[ ], nd triortho-substituted PCB 84). Delyed Recll, similr mesure to Long Term Retrievl, ws significntly correlted with only two congeners (PCBs 153 nd 187), both non-dioxin-like (di-orthosubstituted). Auditory Processing ws significntly correlted with the non-dioxin-like (di-ortho-substituted) PCB 187. The Rvens test ws significntly correlted with only the dioxin-like congener PCB 105. Our multivrite nlysis (Tble 5) showed tht the congeners grouped by persistence ctegory were relted to some cognitive outcomes lso. Exmintion of the bivrite correltions of cognitive outcomes (Tble 6) with congeners in the persistence groups found to be significnt showed tht for those congeners ctegorized s persistent, Delyed Recll ws significntly correlted with PCBs 153 nd 187, Long Term Retrievl ws significntly correlted with PCBs 99 nd 138[ ], nd Auditory Processing with PCB 187. Severl low-persistent congeners were lso correlted with Long Term Retrievl (PCBs 101[+90], 110, 87, nd 84). 4. Discussion In this pper we tested whether congener structure (dioxinlike, nd non-dioxin-like) nd reltive persistence of the congeners detected in concurrent body burden would be relted to cognitive functioning in our dolescent prticipnts. We found tht, with very few exceptions, ll reltionships were negtive. All four congener groups creted for the nlyses were significntly ssocited with some cognitive scores, in most cses to moderte degree. Our sttisticl trnsformtion of the PCB group vribles llowed us to determine their seprte effects. The bttery of tests we used llowed for the investigtion of specific cognitive functions nd the scores on these specific functions reveled ssocitions with PCB congener types. By contrst, the globl scores of memory nd cognition (respectively the Composite Memory score nd the Brod Cognitive Ability Extended score) did not revel these ssocitions. The two mesures of long-term memory (Long Term Retrievl nd Delyed Recll) were ssocited with ll four congener groupings, suggesting tht this ssocition is robust, nd tht the memory function is susceptible to toxic exposure t more thn one developmentl period nd by more thn one congener type. (The vrious mesures of shorter term memory showed different pttern, nd in fct the Short Term Memory score showed only non-significnt reltionships with congener groups.) In contrst to the pervsive ssocitions between long-term memory nd congener group, some of our observed cognitive outcomes were ssocited with only prticulr congener types. Rvens ( nonverbl problem solving test) ws ssocited only with the dioxinlike congener group (nd in fct only PCB 105), Auditory Processing ws ssocited only with the persistent congener group (nd in fct only PCB 187), nd Comprehension-Knowledge ws ssocited only with the less-persistent congener group (nd no individul PCB congeners). The importnce of congener type on neurobehviorl outcomes hs not received much systemtic ttention in previous studies with humn popultions (Schntz et l., 2003). Lck of experimenter control of exposure, limited nd idiosyncrtic exposure ptterns in vrious humn environments, nd correltion between exposure to vrious congeners, hve hindered the conclusions bout congener specific effects tht cn be drwn from humn studies. Reserch crried out in the lbortory nd with nimls hs led to the conclusion tht ortho-substituted, non-plnr PCBs with non-dioxin-like ctivity ffect brin neurochemistry. Seegl et l. (1990) conclude from their in vitro studies tht such congeners reduce brin dopmine, but tht plnr, dioxin-like congeners do not lter dopmine. Tilson nd Kodvnti (1998) conclude from review of the literture nd their own work tht non-dioxin-like ortho-substituted congeners re the most neurotoxic PCBs. They suggest tht PCB-induced chnges in clcium dependent enzymes my hve synptic effects which could be relted to the storge of informtion nd hence memory functioning. This suggestion would be consistent with our own finding of the ssocition of the non-dioxin-like congener group with our two mesures of long-term memory, nd the fct tht lmost ll of the individul congeners ssocited with the longterm memory scores were ortho-substituted non-dioxin-like PCB congeners (but would not explin why Long Term Retrievl ws lso relted to one dioxin-like congener). Animl lerning hs been linked to non-dioxin-like PCB exposure. Holene et l. (1998) reported tht the non-dioxin-like di-ortho-substituted congener (PCB 153) hd negtive effect on rts lerning (but so did coplnr PCB 126). Commenting on previous reserch in 1998, Hnsen reported tht reserchers generlly considered tht ortho-substituted congeners were most potent, nd tht coplnr congeners were not potent. A review of niml studies by Schntz nd Widholm (2001) lso pointed to the role of ortho-substituted non-dioxin-like congeners in deficits in lerning nd memory. Our results show tht both non-dioxin-like nd dioxin-like congener groups relte to long-term memory functioning in dolescents. Nevertheless, lmost ll of the individul congeners ssocited with Long Term Retrievl were ortho-substituted nondioxin-like, s were both of the congeners ssocited with Delyed Recll. Moreover, the non-dioxin-like congener group hd moderte effect sizes on these two memory scores wheres the dioxin-like congener group hd smller effects. The ssocitions of the memory scores nd non-dioxin-like congener group body burden re consistent with the lbortory nd niml studies lredy mentioned. On the other hnd, our

7 692 Tble 4 Correltion of covrites with ech cognitive domin. RPM (n = 268) VMIS NMIS DRIS CMIS BCAE (n = 236) LTR STM (n = 266) PSD (n = 268) APG (n = 263) VPG CKN (n = 250) FRG (n = 258) r p r p r p r p r p r p r p r p r p r p r p r p r p Adolescent Sex (M/F) < < < < Age (yers) < BMI (kg/m 2 ) Triglyceride level (mg/dl) Cholesterol level (mg/dl) Mternl Mternl BMI (kg/m 2 ) < < Mternl Brod Cognitive Ability 0.25 < < < < < < < < < <0.01 Socioeconomic Sttus Index Number of Socil Problems Reported Pregnncy Order Cigrette use during pregnncy (#/dy) 0.19 < Brestfeeding durtion (weeks) Other toxicnts HCB (ppb) p,p 0 -DDE (ppb) Mirex (ppb) Blood led (mg/dl) <0.01 Totl mercury (mg/dl) J. Newmn et l. / NeuroToxicology 30 (2009) Abbrevitions: RPM = Rvens Progressive Mtrices; VMIS = TOMAL Verbl Memory Index; NMIS = TOMAL Non-verbl Memory Index; DRIS = TOMAL Delyed Recll Index; CMIS = TOMAL Composite Memory Index; BCAE = WJ-R Brod Cognitive Ability Extended; LTR = WJ-R Long Term Retrievl; STM = WJ-R Short Term Memory; PSD = WJ-R Processing Speed; APG = WJ-R Auditory Processing; VPG = WJ-R Visul Processing; CKN = WJ-R Comprehension- Knowledge; FRG = WJ-R Fluid Resoning. Bold denotes p < 0.2.

8 J. Newmn et l. / NeuroToxicology 30 (2009) Tble 5 Results of multivrite nlysis of the reltionship of PCB groupings to cognitive outcomes (n = ). P Dioxin-like PCBs50% b,c P Non-dioxin-like PCBs50% b,c P Persistent PCBs50% b,c P Low-persistent P PCBs50% b,c Std. b p r 2 Std. b p r 2 Std. b p r 2 Std. b p r 2 Rvens (RPM) % % % % TOMAL Verbl Memory (VMIS) % % % % Non-verbl Memory (NMIS) % % % % Delyed Recll (DRIS) % % % % Composite Memory (CMIS) % % % % Woodcock Johnson (WJ-R) Brod Cognitive Ability Extended (BCAE) % % % % Long Term Retrievl (LTR) % % % % Short Term Memory (STM) % % % % Processing Speed (PSD) % % % % Auditory Processing (APG) % % % % Visul Processing (VPG) % % % % Comprehension-Knowledge (CKN) % % % % Fluid Resoning (FRG) % % % % Bold denotes p < Covrites included in the nlysis re: sex; ge (yers); dolescent nd mternl BMI (kg/m 2 ); dolescent triglyceride nd cholesterol levels; mternl Brod Cognitive Ability Stndrd score; SES index; # reported socil problems; grvidity; cigrette use during pregnncy; brestfeeding durtion (weeks); p,p 0 -DDE; HCB; Pb; Hg; mirex, nd the converse PCB congener group. b Vlues below the mdl were clculted following the EPA recommended method for estimting non-detected vlues s described in Schell et l. (2003). c Orthogonl trnsformtion ws pplied to ll PCB grouping vribles. finding bout dioxin-like congeners is worthy of note, especilly the finding tht Rvens scores were negtively ssocited with the dioxin-like congeners only. Previous reserch hs provided little evidence tht dioxin-like congeners re neurotoxic. Ptndin et l. (1999) found tht in the brest fed members of their Dutch study, mesures of dioxin exposure were not relted to the intellectul or lnguge performnce of the children. Indeed, Schntz nd Widholm (2001) report positive ssocition between dioxin exposure nd some mesures of sptil lerning in nimls. The Rvens is problem solving tsk involving visul-perceptul resoning nd requiring no spoken lnguge in either dministrtion or response. This mkes the test unique mong those in the btteries dministered to the children in the vrious humn cohorts. The test uthor (Rven et l., 1992) reviews limited evidence tht prticulr res nd types of brin ctivity re involved in the performnce of the test; these my be different from those ssocited with the other cognitive subtests used in the current study nd in previous cohorts, nd so be ffected by different PCB generted processes. Thus our findings in regrd to congener structure re in prt consistent with previous reports of the effects of ortho-substituted non-dioxin-like congener structure on cognitive functioning, specificlly long-term memory scores. The findings, moreover, dd to this literture in tht they show tht exposure to dioxin-like congeners is lso ssocited with deficits in memory nd nonverbl resoning scores. However, it should be noted tht the dioxin-like congener most strongly ssocited with this finding (long with other dioxin-like congeners mesured in this study), hs very low dioxin-like ctivity s compred to the highly potent congeners 126 nd 169. This suggests tht the neurobehviorl effects we observed my involve mechnism other thn Ah receptor ctivtion (Vn den Berg et l., 2006). Differentil persistence of PCB congeners will influence the degree to which their effects re evident in individuls of different ges. Developmentl risk posed by exposure to PCB congeners is likely to vry ccording to the extent those congeners ccumulte in the body nd thus hve persistent influence (e.g. Brown, 1994). Becuse of chronic ccumultion, PCB body burden increses with ge (Brown, 1994; DeCprio et l., 2005), but the burden of prticulr congeners will vry ccording to the rte they re metbolized nd clered from humn tissue, s well s their ongoing existence in externl exposure sources (Hnsen, 1998). The prticipnts in the current study were predolescents nd dolescents, unlike those in the other humn cohort studies. Most studies of PCB effects on development hve tken infnt or erly childhood endpoints. Any effects tht re evident in dolescence my hve resulted from episodic exposure tht occurred prentlly, erlier in childhood nd/or currently. Effects my lso result from chronic exposure. Cognitive outcomes my lso be ffected by previous episodic exposure to lbile congeners tht re no longer detectble. We relted our prticipnts cognitive scores to their concurrent PCB body burdens, wheres other studies hve employed mesures of prentl or perintl PCB exposure. The effects of prentl exposure re most estblished with humns (Schntz et l., 2004); niml studies (Holene et l., 1998; Rice nd Hywrd, 1997; Schntz et l., 2003), nd limited humn reserch (Wlkowik et l., 2001) provide evidence of post-ntl exposure effects lso. As reported bove, our results do show tht tht concurrently mesured PCB levels re relted to mesures of cognitive functioning. We cnnot specify the timing of the exposure tht contributed to the current PCB body burden, but by compring congeners of different degrees of known persistence, we cn get some indiction of the likely timing of exposure tht resulted in cognitive deficits. Exposure to low-persistent congeners could not hve occurred prentlly or in erly childhood; there must be source of exposure tht is more recent nd within the physiologicl clernce time of the congener. The presence of low-persistent congeners in current body burden reflects recent episodic or ongoing exposure (Hnsen, 1998). Effects ssocited with these congeners must hve occurred reltively recently. On the other hnd, cognitive effects ssocited with persistent congeners could hve occurred t ny time from conception to the present. We found tht the two long-term memory outcomes (Delyed Recll nd Long Term Retrievl) were relted to both the persistent nd the low-persistent congener groups, mostly to n extent tht would be considered moderte. The impliction of low-persistence congeners in these outcomes, s well s the Comprehension- Knowledge outcome, indictes tht there hs been exposure to

9 Tble 6 Results of bivrite correltions between ech cognitive score nd PCB groupings nd PCB congeners found in 50% or more of the smple. 694 PCBs (ppb) RPM (n = 268) VMIS NMIS DRIS CMIS BCAE (n = 236) LTR STM (n = 266) PSD (n = 268) APG (n = 263) VPG CKN (n = 250) FRG (n = 258) r p r p r p r p r p r p r p r p r p r p r p r p r p P Dioxin-like PCBs50% b,d P Dioxin-like PCRs10% c,d,e P Non-dioxin-like PCBs50% b,f P Non-dioxin-like PCBs10% c,g P Persistent PCBs50% b,h P Persistent PCBs10% c,i P Low-persistent PCBs50% b,j P Low-persistent PCBs10% c,k PCB IUPAC# [+90] l [ ] l [+123] l J. Newmn et l. / NeuroToxicology 30 (2009) Abbrevitions: RPM = Rvens Progressive Mtrices; VMIS = TOMAL Verbl Memory Index; NMIS = TOMAL Non-verbl Memory Index; DRIS = TOMAL Delyed Recll Index; CMIS = TOMAL Composite Memory Index; BCAE = WJ-R Brod Cognitive Ability Extended; LTR = WJ-R Long Term Retrievl; STM = WJ-R Short Term Memory; PSD = WJ-R Processing Speed; APG = WJ-R Auditory Processing; VPG = WJ-R Visul Processing; CKN = WJ-R Comprehension- Knowledge; FRG = WJ-R Fluid. Resoning. Bold denotes p < Only those congeners tht were found to be significntly correlted with the outcome vribles re shown. b Vlues below the mdl were clculted following the EPA recommended method for estimting non-detected vlues s described in Schell et l. (2003). c P Vlues below the detection limit hve been replced by the vlue midwy between the detection limit nd zero. d P Dioxin-like congeners in >50% of the smple = IUPAC#s 105, 118, 149[+123]. e Dioxin-like congeners in >10% of the smple = IUPAC#s 77, 105, 118, 149[+123], 156. f P Non-dioxin-like congeners in >50% of the smple = IUPAC#s 52, 70, 84, 74, 87, 95, 99, 101[+90], 110, 138[ ], 153, 180, 187. g P Non-dioxin-like congeners in >10% of the smple = IUPAC#s 52, 70, 84, 74, 87, 95, 99, 101[+90], 110, 138[ ], 153, 180, 187, 132, 146, 158, 170, 183, 199, 28, 31, 44, 47[+59], 66, 130, 141, 151, 174, 177, 18, 196, 29, 33, 8, 92, 97, 128, 144, 15, 172, 176, 190, 203, 24[+27], 40, 42, 49, 53, 56, 71. h Persistent PCBs detected in >50% of the smple = IUPAC#s 74, 99, 105, 118, 138[ ], 153, 180, 187. i P Persistent PCBs detected in >10% of the smple = IUPAC#s 74, 99, 105, 118, 138[ ], 153, 180, 187, 47[+59], 28, 66, 77, 130, 158, 199, 128, 146, 156, 170, 172, 177, 183, 190, 196, 203. j P Low-persistent PCBs detected in >50% of the smple = IUPAC#s 52, 70, 84, 95, 110, 101[+90], 87. k P Low-persistent PCBs detected in >10% of the smple = IUPAC#s 52, 70, 84, 95, 110, 101[+90], 87, 8, 18, 29, 31, 33, 40, 42, 44, 49, 53, 56, 71, 92, 97, 132, 141, 144, 151, 174, 176, 24[+27], 15. l Brcket indictes minor congener bsed on roclor concentrtion (Frme, 1997).