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J 26 THE NEW, ENGLAND JOUPUN.111 OF MEDICINE 30 gOl Jan. 6, 1983 N?'T'CE LNs t~. :r,vi r3y be p' `'"j CY ;,Cv~~'~`t . 3. .t~;.. laiciTiiti~ 17 U.S. C3ii;). Fiqarn 2 Reparative Proaadua. The shunt is r.movad iand an anastomosis is made (A) bwwsisrt th.r3ph1 atrium and putrnonary'annria (PA); The int.ratrial IbaMlr!oonrucss thrl.k atrium (LA) to th. rght v.n'trida via thratrial sfpta/ dsfttct (B) (M1rV dhnotrs mitral vWR: and TV tricus)id!va(vw)d simplicity and the potential' fbr growth' and develop- ment of the reconstructed heart and great vessels. Pul- monary blood flow appropriate for normal develop- ment of the pulmonary vasculacure can then be achieved either by a 4-mm central shunt,, as; in, the patient described above, or by a conventional Blalock- TatJUig, shunt„ which we now prefer. Physiologic correction was'complrted in our patient at the age of 16 months,,when the pulmonary vascular' resistance had reached normallow levels. Afterremov- al of the shun't; an interatritallbaffic was constructt:d aoo provide continuity' between the' left atrium, and the tricuspid valve through the atJialiseptal defect, thereby separating the pulmonary and systemic circulations (Fig. 2): The pulmonary arterial circulation was com- pltted by anostomosis!of the right atrium to the pulmo- nary arteries. Postoperative cardiac utheterization re- vealed i'mean right;atrial pressure of'lil to.14 mm Hg,, a well-functioning right ventricle with normal cardiac output, and normal systemic arterial oxygen satura- tion. Obviously, determination of the efficacy of such managemencmust~awaitlong-terrnifollow-up of a svb- statttialigroup of patients. However, this child's gnaa- Eying postoperative course and current well-being serve to documettt the feasibility of surgically altering the'prognosis of newborns with aortic atresiaand other forms of'hypoplastic left heart syttdrome. Wrareindebced to,Dr. Mchad A. IaCorte for his partieipation in the eare of'tAia pa(aent;,and to Dr, Aldo R Castaneda and Dr, Alexander S. Yadaa i for their interest and ~ support. Rkyzxsrt(crs 1. Ler M. PaLolbyc soatomy md 'mrvreladamitiP o( bypoplasia of Or .ornc , aaet mnwkaes.,t.W ta+ee 19S2t 1:61+70.1 2. Noonan JA.,*Ldas A$,. The ttrpophsoc knihaas ryedromr: aa an.l7sb of 101 nrea.,Pedi.v Clis Marth Am. 19!{: S:1t12036. 3. Frier IDC: Report of tdc New EeglaM Rspond )etant t:aediac NoQ.m: Pldiuriat: 19M 63:)76-t71. 4. Norwood Wl. KiriiJk JK. Sanders SP. Hbpoptasdc leA lteart syndrarse: eatpcoea4t rith palliac.e stapry, Am J Cantiot. 1910: aS:17-91. 5. Nonvood i WS. Lans P. Cawoeda AR; C.mpbeJl DN: Eapctxnce W~ah oper.ooor for,bypoplasoc kA:bnn iyndroaoe. J'TlroncCardiorau Sur;. 1961;!tYJ111.9: 6. Mondie IDS. Galln WJ. FriedbaR DL Cooan(ot .atio aawia: report of " arvivd nd wme spavkooen .bar rupeaF appra.ct+o. J Thor.e Grdiovnc Smr. 1973: 61:7T631. 7: CaykrGG.ShaIbN'EA.,MiIkrGElr.,SiepcatpolinioaofAypoPla.oe'leR .ide of t0e Aeut. N En{l I Mad. 1990 2fx7tDa. i. Fre.doo R.N. CulA.m JAG. Rbs RD. Aa+ic avssin wirti isofmai kfc renQiek: dirdoccws .npomdibpapAic 6ediap. Catlkt iCardiovaa Diap. 19Tf5 7:213-95: % , tnwia SR. Van Pr..gh R. Deiet+art VYF. A pdtisdve aperaoos forvervm ietam with aoroc atab iroemupdon. Aen Thnne Surg. t9TS: 1r:369.71. 10. trs.idry S, vru der Hont,Rl. HaroeiQ AR. Eckaer FA. Blaeea E7i Surycal ip.ttiruoe otsmoc aaesis. a TManc Crdiowc~SaR; 1990: 79i43lL 61. tt. Molri H. Horiuebi T. Hao.d. K. st al. Swgicsl ue.emer for trypoplroe;kR lun: sytdwmc:,wu ~epura. J Tbar CaNiovrc St+S; 19'/9: 7i:22S-S. 12. Doty tD9.,Manri6 WJ Jr. SalvaYm RAt:1Wrc R7d. Hypopkrec knhout sysdoma auoeeasfuw ipa117.oaa ndt a.rr openooa. J Tloe.e Catdiornc Sur1. 1990: iP.14i.32: 13. Sade RM. Ca.haodr AR. TU dirpmalir ry6t'.eaeiek. Suryey. 1975; 77:62Y.a1'. 14. Rabmorildi M. HarsiA SO. Caaorrda AR. NaEr AS. Reid {..N. Lung: t+iopry in oaegCninl twtt di.moe: a ewephomeQit,appewes o pWmoaary Yamlmdisesst. r,Rwaan. 1!lM' ssattn.u. STIM 'LJ4ATED HUMAN PHA!GOCYTES PRODUCE CYTOGENETIC CHAMGES' TN CULTITREDMAMM;ALIAhI CELLS At.Ax B. WcrracRO„ MLD:, StcMUrtm A. WtrrzswN, M.D:, MARGARCT DES'rAE7[ltS, B.S.,. SAMUEZ A. LArr, M.D., AND THOtws P. S rossst., M.D.. C H'RONIiC inflammation can, be' associated with N caacer.l Phagocytes in i'n8a2smaltorv lesions en- 0' zymaticalNy reduce oxygen to reactive metabolites, N Fnwn the Httsmointy-ontokw Unit. SVlas..d.una Geaaal Hospni. and Me Ctimca( Genncs Di.uwe, , CAiY4so'1 Hospqut Medical' Ceaoa. Hurad NeEjvsf SctioW: Boum Ad(iresr rspm r.quan m Dc Wealerf ,at tee Coa lldg,. 6d% F)oor. Marsacbwnu Geaer.l Hoipeall 8oaae. MA 0I11r' Supponed Rq p.atr Gom idn fLa U. Pards Found.noa. thr Ladier' A-WMn,v to the Vetenuu of'Forelp WIn.,E"in.W. Hiavm. tAe Cap=a for Tob.cco ReseareD (ps0+t).,dre Ameneae CaoarSocmay ICIlJ6). antitNt Ed no S:. Wetr sva Famdtdon.

b Vo1. 308 No. I MEDICAL IiA'TELLIIGEVCE- WEITBERG ET AL 27 which include superoxide anions; hydrogen peroxide, and hydroxyl radicals. These reactive species kill microorganisms, damage DNA, and lyse mammalian cells.2'a Subletha'l damage to genetic: material in in- damed fo¢i caused by phagocyte-derived oxygen me- tabolites may be one mechanism by which cancer arises in the presence of inflammation. The observa- tion that human ~hagocytes are important in this mu- tagenic~ process~' is consistent with this idea, since there is a stronI 'correlation between mutagenesis and carcinogenesis. o In this paper we report that human phagocytes that are activa2ed to produce oxygen radi- cals also produce cytogenetic damage in cultured mammalian cells - an observation that may help ex- plain the relation between chronic'i'ttflammatory statess and cancer in vivo. Ms'rHODs Chinese hamster ovary (CHO) cells were maintained in modified Ham!1 F10 medium (Gibeo Laboratnrnes„Grand Island, N,Y:);and I per cent penidllin/streptomytut (Gibco Laboratories) in a hu- midi6ed uteubator at 3M. Culture dasks.(25 an=) wereseaded with 2X 105 to 3x 10# CHO cells from a single exponentially `rowin8 culture with a doubling time of 12 to 14 hourn. In 8eneral„tarset CHO celli wenrineubatedlfor30 minute witli, utudmulated human leukorytes or leulcocytes stimulated with a soiubik or partieulate stimulatins,aRenr, After t}iis ineubation,,the kukoeytees andydmulating agents were washed out,and rt+plieed with fresh medium containing 2.SxI0"s'M brtuttodeosyuridine and incubated for 30 hours. The tar=ct Cl1O cells were then atuuyzed for sister-ctiromatid exchanses: Sisuer-ehromatid exchanges nepresettt the interchange of'DNA replication prodleets at,hottwloaotu loci, occurring through a proc- ess ofDNiA:breaka6e and raunion."'This reoombinantevent can be: studied by usia8 the halogenated nudtroside Sbromo-2'-demry- utidine that permits diQerenttial staining oGiister chtomatids. Sister, ehmtoatid exchanges ane seored arpoints otdiseattinuiry in stainin8g and are reported au the mean number of exchanges per metaphase after at least 30 metaphases per data point have been counted. The occurrence of sister.chromatid exchanges in response to diverse phyaicial and chemical agents aornelates strottEly, wwith theinduetion of m+unmaliut mutasenidry or nranogenieicy or both.'2 I.euk'oeytes,wers isolated from anaooasttlated human blood by deatran aedimmtacon under sterile eonditions.'s (Only freshly prr pared Itulkoeyta should be usedTwenty-four hours after tFie, CH0 cell cultures were begun, the medium was rxmoved and leuko- cytes were added in various eoncenttations (see Aesulis). Some 9asks were also treated with 12-0-tetradecanoyiphorboi 13-acetate .(phorboi myristate acetate) (Lot No. 029, Consolidated,Midlared, Brewster, N.Y.) (100 n; per milliliter) or opsoniscd rymoaan (3.6 mg per milliliter) prepared by standard methodl.'aControlf consist- ed of CHO orJls alone and with phorbol myrist:te aeente, 106 laJcocytes, and opsoaised rymosan added individually to,theail- tured cells. In two uperitnents, leukocy,tes i Gvtn a patient with chTonirIrrattulomatousdisease'a were used in plaoeofnotmai leuko- oytes. The flaska wert< incubated for 30 minutes at 3TC, the supem, tant was washed out and replaced with 10 iml lof a medium enriched with 2.Sx1o'° M S+brona»2~'-deoxyuridine (Sigma Chemical, St: I:ouia, Mo:); and incubated for 30 hours as dbcribed by Istt.11 Three hours before harvestin8„ 0:2 ml of' Coloemid (Sigma Chemical (22.5 es8 per milliliter) i was added to each aaak. After hsrvestins, the cells wex hypotonieally swollen with. 0475 M potassium chloride for 12 to 13 minutes,and subsequently washed three times with absolute methanol:Sladal aceuc aeid (3:1). , Of this suspension. 0.2 ml was dropped from i. diatanoe of 8 to l'5 am onto glass slides held at an angle to rupture,the celat. Chromosomes were stained'wiih thefluorescenee-plus-Giemsa method,'a and sis- ter-ehromarid exchanges were scored as dest=ibed above: 'I l avirAw NYI+S"At/EIlJe'J7 111110MtZR4r9w Figure 1. SistervChromatid ExcMarqes in Cultured CHO iCees Ex- ptasetw tolVarious Cottoentratlotts of StinwlatedlL.atlcocytes: Part A showe 1.ukocyNs stlrnu4ted with phorbol myriatete aer talr (PMA) (100 nq per mi1811ter). EacAiihe representti a zieprrue experirnsret Controls consisted of ineubations of CFNC) cells ala» (0). with PM& orwitri 10s urtstimulaled Ieukorytee (WBCs). The upper'Itln. in Part B sttOws pooled results hom the expKiments shown in PartA. The lower kne repressnts tMnumt»r of sister- chromatid exchanges in cultured CHO oells exposed to leuko- cytes fnam aI paient with chronic 9ttinulomatous disease (CG'O). The CGD leukocytes were added in various oor>arraatiores and incuCatrd with PMA (100 ng per milGlner). Rrst7irs Figure 1 shows the results of a series of experi'rnents using vatious concentrations of leukocytes stimulated, with phorbol myriatate acetate. In each experiment there was a coneentration-dependent increase in sts- ter-chromatid' exchanges. Fibroblast monolhvers dued when 10° stimulated leukocytes (not shown) were added. When compared with control values, these m• creases were highly significant (P<0.0005). Phorboi myrisute acetate alone did not cause a reproducible increase in sister-ehromatid exchanges, and there was, a consistently low basal rate of sister-chromatid ex- changes in the untreated CHO cells. Representateve metaphases from these experiments i are shown in Figure 2. To assess thrrole'ofoxygenmetabolites in this proc+ ess, we repeated the same', experiments using leuko-

28 THE NEW EM6LAI+iD:JOUIt.~tAL OF MED[CI!YE cytes from a patient with chronic granulomatous dis- ease: Neutrophils and mononuclear phagocytes from~ this patient were completeiy unable to generate re- duced oxygen meubolites but were normal in terms of other fiinctions:19 As shown in Figure 1, the cells from the patient with chronic granulomatous disease were not able to induce incneased sister-chromatid ex- changes. We also investigated whether, particulate stimulat- ing agents, of leukocyte oxidative metabolism eouldd cause an inczease in sister-chromatid exchanges in cul- tured CHO cells. Zymosan partieles that had been opsonized with fresh serum were incubated1with C'HO cells and' various concentrations of ' leukocyte3: As shown in Figure 3, an increase in sister-chromatid I ex - ehanges in the CHO cells was fbund to be dependent on the concentration of opsonized zymosan•stimulated leukocytes. To verify that oxygen radicals by tliemselves were able to cause these cytogenetic changes and, to investi- gate further the effect of phorbol myristate.acetate in these experiments, we quantitated! sister-c3tromatid exchanges in CHO cells exposed to an ertaytne-mediat- ed superoxide-generating system. Using,the same ex- perirnental design, we:exposed cultured CHO cells to hypoxanthine (7' µg per milliliter) and xanthine oxi-, dase ('1'S µg per milliliter) (Sigma Chemical) withlandh without phorbol myristate acetate. The ! results are shown in Table 1. There was a significant (P<O.00q5) increase in sister-chromatid exchanges when we usedi the hypoxanthine-xanthine oxidase system, and phorbol mytistate acetate had no effect on these re- sulta. DssccSsrorr The discovery that phagocyte-derived oxy,gen, me- tabolites can produce mutations in bacteria is evidence to support the hypothesis that phagoc~tes may have a role in the pathogenesis of cancer.~' ' However, the great difference in the archicecture of mammalian cells and bacteria raises the question whether the observa- tions concerning bacteria can be extrapolated to mam- malian cells. Eukazyptie cells might theoreticalliy, be able to detoxify oxygen metabol6tes by me:aa&of cyto- plasmic scavengers aad enzymes before damage to nu- clear genetic matenizl could occur.9 The resulis,of the present studiy show that srimuLat- ed phagocytes can cause cytogenetic changes in main- malian ceLls by anoxygen-mediated process. Using the sister-chromatid-exchange assay, we demonstrated a concentration-dependent increase in sister-chromatid! exchanges in cultured CHO cells incubated with leu- kocytes activated by phorbol myristate acetate. A par- ticulate activator of oxygen metabolism - opsonized zymosan - was also effective, altbough not to: the same extent. This difference is ascribable to the fact that at the concentrations used in this study, opsonized zymosan is a lesv potent stimulating agent for leuko- cytic oxzidative metabolism than phorbol myristace Jan. s, 1983 A i Fgurs t FB.prssntatlva NLtapMas.a ir,om CHO Calb.ihcvtaa.d witlti Phorbot Myristata Aptata (10n ng p.r mfAilit.r) (N) or Phorbcl Myristat. Aic.taq (100irqp.r miltlBt.r),plus 10' Lsukocytp (B): Etwan sest.r-cfnroma6d .xchangeN ans pns.nt in Part A and 23 •xclrtargp ; in Part B, as ixlwtrac.d iwith artowa. Excnangs ean be i n.ocgnit.d' at pvU+ta of dilsoortinuity in stainirq alonq i.at:n cnromaod& acetate. We realize, however,,that these in vitro condi- tions do not necessarily reflect in vivo eoncentrations of oxygen radiulsa° Our results prove that leukotyte-genented oxygen rnetabolites or cheir, reaction products with target-cellfl components can reach the nucleus of the cultured mammalian cell and' cause eytogenetic damage. al-

vol. 308 No. !. MEDICAL INTELLIi','ENCE- WEITBERG ET, AL. 29 though i some cells may die, others suffer sublethal i ge- nt:tic insults that can be propagated to daughter cells and may be important for tumorigenesis. The results of our experiments using a cell-free su- peroxide-generating system demonstrate that oxygen radinla cause sister•chromatid exchanges in mamma- lian cells„and they del'ineate the role of phorbol myt is- cate acetatrin this system. Phorbol myristatracetatris a potent tumor promoter that has a variety ofeffects on mammalian cells.2'1 Ito our experiments phorbol myris• tatr acetate did not increase the frequency of sister- chromatid exchanges when added to leukocytes from a patient with chronic granulomatous disease or, to, the hypoxanthine-xanthine oxidase superoxide generat- ing syste= Furthermore, phorbol myriatate acetate alone produced' no increase in sister.-chromatid ex- changes in CHO cells, consistent with the results of other •inNestigators.''u In other experiments a small increase in sister-chromatid exchanges was observed' after ex sure of cell cultures to phorbol myristate acetater and, was suspected to be due in part to superoxidrproduction;26 reflecting an indirect effect of ' myristate scetate.2s The present results support this idea and suggest that: variation in such an indirect effect of phorbol myristatracetate may in part underlie variations in the induction, of siater-chromatid es- eliulges obtained with phorbol myristate acetate in different laboratories. Therefore„we can condilde that its major effect in this system is as a potent stimulus of the respiratory burst of phagocytes. These data also suggest that phorbol myristate acetate may participate in tumor production in vivo bystimulating,production of toxic oxygen radicals from leukocytes. Under tbe: eonditions i used in this study, phorbol myr6state ace- tate did not augment the effecr of the oxygen metaboo- lites on target cells. fipun. 3. Rsulb 0tit.in.d wilh LeukocyAes S7ir>xdWd with Op. sorAz.d Zymosan (3;6' nq p.r rtwYiptar). Controb ootasists4 of CHO cells abn. (ft with opeordz.id syrno- san (Oi or with 10' l.ukmtytes (AWBCs). Each i4nm r.qr.a.MS a ssparatr.xpsrim.nt Trbl+ 1. SMar-Chromatid Exdurgss in CulEund CHO C.ds Ex- poslMd to a C.II-Fr.. Supsroxidr-G,naating System with and. wil~out PhorboF 3Nyristat6 Acetate (PMA);• Aoe1}fOM ,iD alo cyLW Cti7L'NaO s.r..{w.oWSm fse.nrm !MM (400 aB/mq, ......... .............._.. ..e sssac 5.l3s0:311 6x_o.4o. Hypon.&ue (7 µWi .............. ..,... 5.8020.431, XaeI aaidm (15'Mtml) ................ 4.t9so.336 Nypo..ndlw.,..aI:..Otine oaianc ........... 1l.r0a0.910 Ny9oa..aume. aaedbiae oa;a..r. .ea Iltru... . 14.67s0.974 w.e..~ .. 10 r... These: results indicate a primary role for oxygen metabolites in the; generation, of sister-chromatid ex- changes produced by leukocytes. Although we cannov yet directly relate our results to carcinogenesis, this observation may help explain the relation between chronic inflammatory states and cancer in vivo. We are iadebmd to Ms. Maei[yn Griroai for barapert technicai aaoanna Rsrzpmqcaa 1. Fe.ommiJFh. finon at liO ussk olKaeorw in Wpro.as Io are>r stiolop . aed laoaad. New YaN= Aodeaoie Itiw: 1975+. 2. BabiQ BM. 0aypo,d.p.dmt siaobi.l l3IIi.g by ppypeyut. N IE* J M.d. 197»: 29tA'54~6{. 721•5. , 3. 10.6ieotr 57. ory=s mmtidi® aed ide b:ie popeeor v(p!.{aeyr. Am laoe M.d. ,19q, 97i46D.9.. 4. Bae.ey, JA. YmsovYry lrC.. AeQS onS.r rp.eir aed Ytie lhr.tion d phgoeyoe leutoiey..: Alem Bs. BroeArs. 191t); Nli69S726. s: Bimbaim HC. DNA m.ed brekaae 's,lim.e MWkoeyr aspaM w a e®ar pomorr. p6oeEni mlrtiwie aeeme: Seimcc. 19it,2; 215d1247-9. 6: , Simoe RH. SmMio Qi. Pn.noa D. Hydope paeucide oo.r ir. 6a.1 lojpry to h umm 64oblaats acpowA ifa aary{en indiaa4: J Biol Cbam. 1992; 296:71s1,6: 7: we=s SA, s+orda. J1luaem amurd by 6minampAqayl.: Sciiea. , 1961: 212316.1:. a: IdeR. Fdi.aa of oaY edioal awveern ud lemzidaoa m peyoep~ imirc.d onio3mi J lavicumot. 1981- 12r177iY2. 9. MtCaN IJM. fi ie0 1. 1Lr biolop aod p.molop of'osyaa ndieall. Aaa 16eee M.d. 19'ik t9:133.7. 10. MeCam J./lmea BNc 7Le SnG~oee!ldmiaaome,ouo~seieiry ~re pei tivs value foi kareiootmieity.1e:1G.a 1Dl. N'.om JD. Rriaom JA. .di. odphW d Mm.o oaoor: Cdtl'Spri.ill, dYi N:Y:: GoW Spr* HsOa tatiaaoory. 1977:1431-50. 11. Stiir.i.hi'Yu Sasdbe5 AA. Sbi : edomrod eub.n{c i. Mrs ahomo- wmead iadhd9og oE..n.dom ie o.oplaoa. GmaiGoee Cy+o~ 19l0: W67d0. 12. Lae sA. A!{m l. Bloam SEL at al. S'uwer<liam.od iwdrew a u.pnt ef .>>s G4er7m AvQam. ,T,'Imt lla. 19111; ,t7:17.6II. 13, , Itibrk JC: Seood'R. Deeeuoosi yr4ogmau: aod permoop d6Yeqs o t®.e Q.wtilocym eaond by meeastiar ah ryloa .roa1 5bw impiio• aroes for 6)naoaa lataptiaeit. J iQm 1ei ,19T1; 60a1[340. 14: 511eCWoi J. aabtra BJ. D.ie.d IAR. Bnm J. Term.r jE. Qs: iHG.,L . veo lleceoa rd po.o-aaedtsias .rviqi of'psealocyrs ooliumd by aonoauao-so: oesQinKaoceaodley6laaemila~k.pemeaia.Bbod 1976;. H:315'`26. 15. Soore!'77: Fid6RJ. Gitl'm JD: A'IIMr G. Ao..o PS. 7Ls op.aairtr.~.m of ine di oompoemt of hi oooaplemes J FsP Mad. 1975i J/11:1M 47. 16. }lOyiprt A1. Colbdey A3t: BichoerRA. Cbmirgrmabmuos dhre of ehilfmocd. J P.d'sv Sorg. 1969; 4itS•M. 17. tWsA 5iwneEram.m eu~ae~e.ao~. deum.a adomo.om. e.mqe .od leep.c daictioa !y 6uwrsomae :aed isdocoar try, minmyei C. !!oc Nad Acd'Sd USA. ,1974; 71:31U+{. li. lary P. Wolfri5. Wm CAema memod for iedCaa.ed oieog ial aiv cheom.oi Nama. 197Y; 351:1l64. 19. Nad= DL'i; BrAee RL. Wdve DK. , Fadis af eiao bine eesdiom wduaooo Ao ube pp.{ocyoe w.nnb a(Iautxyoes in alroeie Srsulomroue diurs. 141ia ls.+iiiii. 19d9: 4i:1s93+901:

THE'NEW ENGLAND JOURNALt OF MEDICINE Jan. 6, 1983 20. McCord JM. Futradioala aedlioEammaoao: parttioaoftryaprial IEuid by tuqrmaoda diamn.e. Soimoe. 1974; US:SM31. 21. Diamood L. O'&isn TG. Baud !Wbt. 7Mlmor pomoma md itie im-haoirm d eama peomodon Adv Caeov Rr. 19t~ 321'•74. 22. Lo..dar KS1,La¢ SA. Tsa sffia ofa mmor pamoee. 12-0-ne.daauoyl- ppaeboi-13+.=ow (TPA). m aqaslyam.dd aelaep foemmom i. au4 oA.d,CEime harmaer eella. Atw.e Rn: 1979: 67il43!1. 23: Popeaeu NC. Amab.uSE SC. DiAlolb JA'. f.8ttaecaeeo[ o! N•meMyl-M. niaoN-®aoatuaoJdioa vamfcr~ of'Sytiaetamoeslb by a ptiorbol ai.ae v,iedepeoaeae of isiae mam.od iueeseSe. aa oemomromr,aea• noooa. Proe Nad Aad Sq USA. 19tb 77:77,32-6. 24: Popacv NC, AmaEwSE SC. Lae.mmdl! lf4. Molb IA'. 12-0,n~ d.camy4phabo1.13-aeraa aed,ia utlaooadip, to SCE lioAcooo iia Sym ®d Chims lmav odh. Enwoe Nfutyen. 1912; 4s7+S•!!1 25. Mi11er RC. O..wd'CR: Omma1c Rb. at ai. Madi6eaeoe ot siw chromaod aemn~a aad aadiaootioduaed ioraeatamaoon m eodem cell! ib., d`e oumor peomoaer t1-0,uevdoaeoylptiaebo413raeesoe aed two teooodc: Caaixr' Rn. 19t1: 41:6SS-9. 26. Napaava H. Liak,18. lem+cdaa a( cheomoaome atiemoos aod aiaer atyomaod nedaeps !y R-era i. dlmityiahiEird cvlo.q of mowe lOSSi aeW. Radiat R+.. 1911t 1933i-Sl. 27. ScEwuR IL. Daada,Ft1. Woitr'S: 1Z40-eeadiomoylpbotbd-l3-asnae (t~A) lisolr r,aiaartheom.od eusan{a aad Idahya iir odl ipeopeµioe ie Clioar.tl.mroe a.ry sod Euoae all lies. Mum Rea. I9l2: 9Y:393-409. 23. Fmnft'I. Caam PA. TWmour pomaer plrorbo! III;®yaataoe.I3-aeenoe iod>co olvamaaosal duap via iedie.ct aeooe. Naous: 19t1L293:1w46. CASE FtECORDS' OF ?= MASSACI3HUSETTS GENERAL HOSPITAL Weekly Clnsticopathologicad Exercises FotJNDW: sY~ atC=AAD C. eA'SI04, Rosns L Scuutt, M.D., FdiJor Evaurs J. MwRSC, M.D., Armeiorr Editor BsrrY U. McNasLY, Ar,ufant Editor ' CASE 1» 198t PivssrMaztoN o. Casr A 37-year-old' woman was admitted to the hospitail because of hypoglycemic ~ episodes. There was a long history of psychiatric problems,, with multiple admissions wmentalihospiiala becausee of suicidal behavior, assaultiveness, and paranoid ideas. Thirteen years beforcadmi+sion shecameto~this hospital, where she was. followed in the psychiatric clinic with a diagnosis of chronic schizophrenia. Var- ious taedicationa were.prescribed, including thiorid- azine, chlorpromazine; 8uphenaaine hydroctiloride,, trifluoperazine hydrochloride, and imipramine hydro- chloride. Three and a half years before entry she re- ported' consuming up to 180 ml of vodka dailiy for relaxation. The urea nitrogen was 12' mg per 100 ml (4.3 mmol' per liter), the glucose' 39 mg per 100 ml, the bilirubin 0.5 mg per 100 ml (9 fcartol per licer)'„ the cholesterol 187'mg,per 100 m1(4.8 mmol per liter), and the protein 7.7 g, (the albumin 4.5 g and the globulin 3.2 g) per 100 m1. A three-hour oral glucose tolerance tesrwas performled'; the fasting glucose was 56 mg;,the glucose at 30 'miitutes was 84 mg, at 60 minutes 80 ing, 'rp coowt to st vai,. mtaIriqy by o.oessl. at 120 minutes~ 101 mg, and'ar 180 minutes 47 mg per 100 ml': After an overnight fast the glucose was 59' mgg per 100 ml, and the insulin less than 3 µU per milli, licer. T7ie patient continued'to be followedl in the psy- chiatric ! clinic. Five months before admission her mother observed her staggering as if drunk and confused; after, eating an orange she impnovedi within 10 minutes. One month later on a visit to the clinic the ~ patient reported that she was wea'k andl sweaty at lunch time. The glucose was 53 mg per 100 mli A 6ve-hour glucose tolerance test showed that the fasting, glucosr was 67' mg; the glucose at 30!minutes was 121 1 mg, at %minutes 117 mg, at 120 minutes 117 rng, at 180:minuces 107 mg, at 240 minutesi 57'mg, and at 300 minutes 54'mg per 1100 ml. The patient had no symptoms during, the test. A hig)1-protein, low-carbohydrate diet was provided; and she felt improved. She discontinued the intake of alcohol at an uncertain point. While she was adhering to the diet her weighr fell fnom 77 to 64' kg. Two weeks before entry she was examined in the clinic, where slight galactorrhea was noted from the right breast. The fasting glueose was 50' mg, the urea nitrogen 14 mg,per 100 ml ('5.0 mmol per liter) j and the thyroxine 7:5 µg per 1100 ml (97 nmol per liter),. The thy,roid'+ stimulating hormone was 1.3 ytU per milliliter. The prolactin was 8.5 ng per milliliter. The serum aspar- tate aminotransferase (SGOT),was 26 U per milliliter (0.21 µrnol - sec' per liter)', and the.alkalinephospha- tase 28 IU (0.47 µtnol - sec') per liter. During the next two weeks she experienced repeated bouts of con* fusion, and disorientation, during which observers thought that she was inebriated; each episode was promptly relieved by the ingestion of orange juice or sugar, and she began to carry a thermos bottle of or- ange juice at all times. She was admitted to the hos- pital. ~ There was a long history of a chronic cough without evident cause. Intermittent incontinence of urine had ~ occurred for many years, with repeated urinar,v=tract N infection; the symptoms improved after urethral dila- cation. She complained of restless sleep and nocturnal j.i sweating: There was a history of diabetes mellitus in QD her maternal: gpandparents and l maternal aunt and of r carcinoma of the bowel' and psychiatric problems in CA her mother. She denied syncope, seizures„reeencinges- W