1409 1409 - Attorney General of Minnesota

Fluomchemical Steeling Commillee Members

Jeff MandeL M.D., 333-8670 (220-3W-O5)

Papers/Dr. Gilliland

December 7, 1~ ¯

These are two additional papers that a~e being prepered by Dr. Gill=land_ They need ou~ inpul/appmval before being su~itted for publication.

The papers are negative for lhe.mos! part, We’re workirg with him regarding some of the wo~ding. We’re operaSng under the pre~nise Ihat these s0Hs of topics need 3M suppod.

Please send you~ ~,omments back !o me by the end of Ihe monlh so I can communJc~e with IDr_ Gitlilat~l.

JHM/vtk

CONFIDENTIAL - SUBJECT TO A PROTECTIVE ORDER ENTERED IN HENNEPIN COUNTY DISTRICT COURT, NO, 27-CV-10-28862

3MA00323875

Exhibit

1409 State of Minnesota v, 3M Co,,

Court File No, 27-CV-10-28862

3M MN00051484

1409.0001

Peripheral Blood Lymph~T~e Coun¢ in Men Occupationally Exposed to

Perfluomoctanoic Acid

Prank D. GiRiland, M~D., Ph.D.

Jack S. Mandel PKD.

Affiliations: Division of Environmental and Occupational Health, School of

Public Health, University of Mirmesota, Minneapolis (F.D.G); and

Department o~ Internal Medicine, Occupational a~d ]~nvivonmei~tal Medic~e

Section, St_Paul Ramsey Medica~ Center, SuPaRI0 M~nnesota (F.D.G.)

Reprint requests should be addressed to F~-n~ir D. (~il!i!~d, Univers~W of

New Me~xic~ Sclmol of Me~e, New ~ T~or ~, ~ C~mi~ de

Salud ~, ~buque~ue, ~ 87131. Tetep~ (~5) 277-~41, F~

277-7041.

This study was supported in part by NIOSH Grant T150h07098-16 and 31v£

Medical Departmen~

Running title: Lymphocyte count and PFOA "

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CONFIDENTIAL - SUBJECT TO A PROTECTIVE ORDER ENTERED IN HENNEPIN COUNTY DISTRICT COURT, NO. 27-CV-10-28862

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ABSTRACT

Studies in Rhesus monkeys suggest thac perfluoroocta~oic acid (PFOA) has

immua0toxie effects on the coil-mediated immune system in pximates. The

predominant histopathological lesion in PFOA-treated monkeyswas diffuse

atrophy of lymph nodes and splenic germinal centers. Although PFOA

acmunts for the majority of fluorine presen~ in the serum of the general

population and in occupationally exposed worker~0 little information is

available concerning human responses to PFOA exposure. To as~ss whether

PFOA exposure is associated with effects on the human �~ll-mediated

immune system, we examined the cross-sectional associations between

peripheral bk~l lymphocyte courts and PFOA in 115 workers employed at- a

PFOA production planh Total serum fluorine was ur~ as a surrogate

measure for sex~.un PFOA levels. Peripheral blood lymphocyte cotmt was

sig~facanrly ass~L~e~_ with total serum JQ~oride level; however, the

magnitude and direction of the ~elationship was dependent on smoking,

alcohol use, and obesity status. For example, for nolI-smokers and moderate

drinkers, an ~ncrease of 10 ppm in/~! serum fluoride w~s associated with a

decrease in lymphocyte count of 1~40 ce)]s in non-obese (BM]=25 l~g/m2)

worker~ and by 925 cell in obese workers (BMI=35 kg/m2). ]’I~OA is

s~iated with alterations in peripheral blo~ lymphocyte numbers ha PFOA

pro~iuc~ion workers, suggesting ~hat cel].media~i immurd~y may be affected

~ PFOA.

WORDS perlluorooctanoic acid, epidemiology, immune toxicity,


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Perfiuorooctanoie acid (PFOA) is widely used in industrial proce~3es and

consumer produet~ a~ a reset ofit~ unique chemleal properties and potent

surface activity (Ot~fith, 1980). Because PFOA has s long biological half-life,

small ~quent do~es can ac~lmulate to appreciable levels (Ube], 1980). As a

result, PFOA ha~ been found in the serum of all human population studied,

and accounl~ for the majority of fluorine pre~ent in the ~rum ofp~pul~i~n$

in industrialized countries1"7.

Little i~ known abou~ the toxic potential of PFOA in human~; however,

studie~ have suggested that the celIomediated immu~e system may be a site

of tuxicity in primates. Rhesus monkeys treated with oral PFOA developed

histologle ehange~ in ~p~eeu and lymph nodes. The primary histopathologic

lesion was atrophy in lymph node and splezfic gex~ninal centers1. The immune

system o~rodent species bee not been rei~orted to undergo

histvpathologie changes in subacute and chronic feeding studies !. No data

available concerning immunotox~c~y in human~. Becatt~e PFOA ~s present in

the ~erum ofexpo~ed worker~ and in the general population ~-7, it is a matter

of concern whether the finding~ in monkeys indicate the potential rot human

immunotexicity. To assess whether PFOA could affect the cell-media~ed

immune system in humans, we studied the a~ciation of PFOA. as measured

by total serum fluorine, wi~h peripheral b~ lymphocyte count (PBL) in 115

occupationally exposed emp]oyee~ at_ a plant l:hat produces

B~L-kTERL-~L~ AArD ~THODS

;3MA(}0323878


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All current workers employed in PFOA production over the 5 previous years

and a sample of workers in jobs with no apparent PFOA e~posure for the

previous ~ years were invited to participate. Participants had vi£a!

paxameters measured in the.plant medical department by an occupational

health nurse, c~mpleted a medical history questionnaire, and underwent

venipunc~me. Blood was drawn foz assays ofperipheral lymphocyte count. A

fluorine-L,-ev 15 ml va~tainer was used to ~Hect blo~i for total serum

fluorine dete ~r~_ i~Ation. Total serum fluorine was used as the measure of

PFOA in this eccupatieual g~oup, Total sexttm fluorine was determined using

the sodium biphenyl extraction and atomic absorption spectrometry s,

RESULTS

Pax~icipant charac~ezistics are displayed in Table L Total serttm fluorine

values ranged f~m 0 and 26 ppm with a mean of 3.3 ppm. Twenty-three

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(20.0%) par~cipants bad ~erum values less than I ppm. 6 (5.2%} had values

between t0 and 15 ppm, and 5 (4.456) had values greater than 15 ppm_

Table 2 presents the o~rdation coefficients between PIlL and total serum

fluori~e, age, BMI, akohol use, and cigarette co~umption. Peripheral blood

lymphocyte count was significantly correlated with total ~erum fluorine

(r=.19, p=,04). The fin~] regression model for PBL, which adjusted for age,

BMI, cigarette use, and alcohol use. included data for Ill workers; 4 of the

115 workers had miss~g assay values and were excluded from the analysi~

(Table 3). Total serum fluorine was inversely associated with PBL; however,

the relationship was complex, with si~ut interactions bel~ween total

serum fluorine and BMI0 ~iRarette use, mud alcohol use. Table 4 illustrates

t~e rela~onship for a 10 ppm chan~e in total s~rum fluor~e for comb~uations

ofsmokingo alcohol consumption, and BMI st~tu~. In moderate ~h~;-ker~ (1o

3oz ~kohol per day), PBL decrea~d in the categar~es of ~moking an~ of

obesity; ~n light drinkers (<loz per day), PBL decre~d ~n non-obese smokers

only.

Discussion

Serum PFOA level is asaocia~ed with chan~es in the cell-mediat~ imm~me

~’stem, as e~denced by cha~ ~ PB~ ~we~r, t~ rela6o~ depen&

u~n smok~, alcohol use, ~d ob~i~. PFOA ~d ~ ~B ~ by

alte~ng the kn~ effec~ of smo~g, ~co~l ~ump~o~ and a~si~ on

pe~phe~ le~o~e ~ (~). No human sc~es oft~ eff~ oEPFOA on

~he ~mune sys~m a~ av~ble for ~mpa~n: howe~r, a study of th~

~up ofworke~ s~gesm ~hat PFOA may mo~ ~ ~pafic ~pon~ ro


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alcohol and obesity. Modulation of endobiotic and xenobiotic metabolism

could represen~ a common mechanism for ~he observed association o~ PFOA

with alterations in hepatic and immune response.

q’he relationship between the changes in PBoL in humans and the lymph node

and splenic germinal center atrophy observed in monkeys is not clear.

Clothes in PBL may be unrelated to ger_m~al ~nter atrophy. Alterna~icely,

small changes m PBL could reflect larger ¢~__~n~e~ in T cell subset~ In

addition, the modi~ation of the relationship between smoking and PBL by

PFOA could be the ~es~lt of changes in the number of particular T

subset~. Furthermore, PI~)A may be associated with eh~nge~ in

function beyond simple changes in eell number. Cytokine

important in immtme f~m~ion and mu~d be all~l by PFOA exposure ~. The

response to ~u~i~en binding depends upon rearrangement of membrane

pmtein~. Changes in the membrane physical characteristics prod~i by the

potent surfactan~ action of PFOA could alter ~rnm~ r~b-pon~eso

Interpretation of the finding~ requires careful consideration of the study

limitations. Given the occupational study .setting. the voluntary

participation, and the requirements foz blood sample collection, the overall

participation was unexpectedly, high. and non-response bias is likely to be

small Workers not included may have had a different response pattern than

those who were included. Migration our of the high exposure ~obs is ~ml~kely

to be the result of subdinic~! changes in PBL counts. The vast majority of

workers who had sight exposure over the previous 5 years would be

included in the s~udy ~ample as the ~urn-over rate in plant emplo.vees was h)w

{3% per year) and the study included all current employees with appropriate

3MA00323881


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job histories_ Selection bias is not a likelx explanation for the findings in this

study.

Inflammatory and infectious processes, which are major determinants of PBLo

were not assessed in ~his study. Because the~e is no evidence that these

precedes are related to tot~ serum fluorine or serum PFOA, they are

unlikely to confound the estimated ~elationships.

The cha,~ges in PBL count~ a~sociar~-d with PFOA expe~ur~ p~.sent a complex

picture. Alcohol use, cigarette use, and BMI modified the association of cell

count with PFOA. The magnitude of these association~ is not cllnle~lly

significant from an infectious disease perspective: however, ~udgment as to

the clinic~ relevance of such changes mus~ av~it further stedy. More

3MA00323~82


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remrarch is needed in the area ofPFOA immunotoxicity. The findings ofthe

pre~ent study need to be confirmed. Changes in T cell subsets could be

confirmed by immunophenotyping lymphocyr.es using well eetabli~hed flow

~;~metry metho~ls 2s. ~. In acldition, the standard Immunotoxicologic

assessment defined by the National Toxicnl~gy Program ~e ne~s to be

conducted Rsr PFOA.


3M MN00051492

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BMI tk~/mS) z6.~ zs.s.40.5

per day 8 ~uces per d~y 20 17.4

pe~ day 0 non-re~

3MA00323~84


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TABLE2 P~arson �orrelztion coeE~ients between to~l serum fluoride, bo~ly ~ass index (HMI). dai~y akoho! use, d~ily tobacco co~suml~tion, and

peripheral blood lymphocyt~ count

Total Age BMI Alcohol Tobacco

~pm)

LYMPHOCYTF.S .19 p=o04 p=.O02

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TABLE 3 Linoar multivariate ~egr~ssion mod~l of fac~rs predicting the lymphocy~ count among 111 workers.

2205.6 6tL1 .0005 Total ~1~or~ (ppm) ~

-526,6 222.7 .07~

-977.1 35~.7 .007 low X Fl~odae

189.0 52.3 .00~5

247.9 103.9 Cilg~u-ettegday

""~’~- 34.0

~..s .o~ ~ ~)

1.58 19.8

7.t5 4.1

R2= .~

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Table 4 Change in lymphocyte count for 10 ppm increase in total serum fluorine

oz. alcohol/day

Non-smoker Smoker (20 oig~/day)

BM125 mg/Kg2 +7~0 -406

BM135 mg/Kg2 .+965 +306

1-3 oz. of aIcohoYday

Non-smoker Smoker {20 cigs/day}

BM125 mg/Kg2 -1~10 -9.300

BM136 mgiKg~ -924 - 1585

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Tares D. Comparision ot~’ortnmic" fluoride in human and mmhuman serums. J Dent Rea 783.

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CONFIDENTIAL - SUBJECT TO A PROTECTIVE ORDER ENTERED IN HENNEPIN COUNTY DISTRICT COURT, NO. 2~-CV-10-28862

3M MN00051497

1409.0014

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1409 1409 - Attorney General of Minnesota