Document b2dVzE877YBvJj76n6Drpxg0

UJi4s 0>mo a c > INTERNATIONAL LEAD ZINC RESEARCH ORCANIZATION, INC. * V--* 4* H hUQ*ON AVln>l. NEW VO**. H V, TIlMMMI ETE-EOEO Uu 6001 CAEtE AOO*>EEi NV*V* NEW *0*4 Fsbrusry 23# 197 Tot All Motors of tto Lasd Enrlrccasotsl Bssltb Ccoslttss A*t Progress Report oo La-117 "Effect* of Ls*d oo Tissue Culture of the Csatrsl fcrtcui Brstsa* -1 Attached for your Information ( * copy of tto report adleb cover* tto work dooe for us by Dr. Sobkovlct at Coluabl*. I think you will flad :>5 t&Xs quits interesting sad that It opens up teas taportaat SYwaies for futurs investigation. Dr. Boblevies will prepare sa article for pAlicet lea la a nedlcal Journal for sow futurs data. This artlcla will, of courss. bs ssat to us for review before It Is paMletod. .1/I Tsry truly yours. DOfisw enclosure ;1 Don 0. fowler Project Manager & ? ' zf&m&jppvs Principal Investigator: W`n Project Director: -i "Effsct# of Ltad on Tlasus Culture of th* Central Bervoua Oystea* Dr. Margaret 8. Hurray Professor of Anatomy Laboratory for 0*11 Fhytiology Coluabla University College of Ryalclana and Surgeons Dr. Leonard J. Goldvatsr Prof***or of Occupational Hedlcln# School of Public Health and Administrative Medicine Coluabla University Dr. Manna M. Sofckewlet Assistant Profttior of Bsurology n*u* Culture Laboratory of th* Dept. of Beurology University of Vlaconalo Medical School An Investigation of th* direct action of lead compounds upon Bervous Tissues in Culture wea performed. The attbod of tlaau* culture was choaen for the study becauae of two Data advantage*: 1. The portion of the Dervous tyitn under atudy can be *irpoeed directly to th* experimental agent without th* agent havlrg been mstabollsed by other organa jf ` L i 2. Th* blood-braln-tarrlsr with Ita filtration function can be by-passed. Thus through the Isolation of cultured tissue frem the whole organism, secondary effect* on the nervous system fro** polaonlrg of other organ* can be eliminated. The lorg-term culture* from teveral area* of the nervous ayitea* eaiataln their cellular components with eaaentlally noreal etructur* and function for considerable periods (weeks, *cnth). The cultures of central and peripheral nervous system were obtained from new born sous* and rat ctrebellta til cerebral cortex, and fren to&rycnlc mouse apinal cord and sensory garglls. The etudy on the Influence of lead upon the nervous system la concerned Jfth three related but different problems coral Jerri itjar ately as fart* i I, II, and III, "Influence of lead Acetate on th* rwl ;eur.t an.) Ha|ntrac-'e_cf lUrvau* Tiue Fo <3T; t ire.1' [h .M. jotkcrvlct F rf.Ii. Hirray, 1. j7~ivulive*.*r, .8. *Tteron and KTnTlh rnalaln) for thl* atudy 1,17 culture* of newborn rat *M nous* esrstellua and apinal cord and tplnal ganglia were exposed to the heavy matal for peri'da of frrm 1 to J week*. The influence 1**4 acetate In concentration* of frm 10*'M (0.0)7) y/l) to 10'*M {J 7.<00 v/ml) of feedtrg aolutton vu studied for Its effect co myelin Mlntenanc* In feature culture* and on the pattern of development of yourg culture* compared with thoa* where no heavy metal wu present. H. ;~'4 wrm?? rvorn a1 N 477.01 V* found no evidence of prLeary Morphological change* In nerve cell bodies, axons or syelln heath in cultures exposed to lead acetate In vitro, even la high concentration (37000 y/aX of nutrient oediuss up to one week or erpoeure). There vu however a conatant and barked degenerative reaction of Macrophages followed by hypertrophy of glial call*. Slnllar resuita were obtained during preliminary erpoaurea of culture* to triethyl tin sulfate and gold chloride. Cultures exposed to Mercuric chloride (and thalllua chloride and acetate - K.R. Peterson, M.R. Hurray 1965) ahoved evidence of direct toxic action by these aetale on the nerve cell bodies or their axons, with secondary charges In the syelln sheath* The above reactions of cultured nervous tissue see* to divide these Metals Into two group#: 1.* Those which cause a direct Morphological Injury to the nerve cell bodies or axons (aercurlc chloride, thalllua chloride and acetate) and, 2.- Thoae which provoke Morphological change# primarily In eells of ateoderaal origin and only secondarily in neuroglia (lead acetate, triethyl tin sulfate and gold chloride). Of the nrtale which were directly toxic to the nerve cell, the degree of toxicity was dependent on the concentration. The action of aetale which did not produce direct acrptoolcgleal response in the nerve cell was dependent Mostly on tbs duration of exposure. Also the degeneration of ecto*and Meoleraal support leg tissue was c c g bo o to this group of betals. teed, which was tested Most thoroughly, sven facilitated the development and differentiation of nerrwl tissue In young cultures, although the archanisa and Meaning of this observation la not clear to us. II."Absorption of tead Acetate by terrors Tissue In Culture" (H.H. Sobkovics, E.R. wSe Petersen, X.R. Hurray, L.J. OoIdwater) The lack of primary Morphological change# In nerve cells, their axon* and myelin sheathe In cultures expose 1 to lead acetate, as well as the normal pattern cf the development and sqrellnatioa of exposed young culture* (** - 6 deys In Vitro) vae rather lntrlgulrg to us. Using the hlstochenlcal staining technique of rhodlacnlc acid for the pretence of lead w* started a systematic study of the up* take of lead acetate frees the feeding solution by nervous tissue* In Vitro. The nature Myelinated cultures of new born rat and Mouse cerebellum and esbryo bout spinal cord and spinal ganglia were exposed to 50 *v/l of lead acetate In the feeding solution. As the results of our study * showed that there 1* no evidence of the absorptlcm of lead by the nerve cell In Vitro fron lead-containing Medium. A heavy uptake of lead 1* however obeerved In Macrophage#, aentgeal cell* end to a lee* extent by the glial cell#. Therefor* It Is etrorgly (</*ted that the toxicological resfoc.ee* of nervous tlseue culture# exposed to lead .'el*,.e correspond to the hl*toeh*tcel pattern of absorption of lead by the*# cCul lure#, at we observe It. The lack of evidence of lead uptake freer the nutrIrM*'Medium by the reur^a* ccull explain In fact the rather eurprlelng phenrxaonon of the lack of toxicity of thl# oelel to the nerve cell and fibre and the ayelln sheath. C* the other hand tie cells which were Involved In the proves* of lead ekeorptlcn fron live nutrient Eellix shove.| buth toxic reaction and m-.rp>hnlogical damage during the Its* of exposure. The appearance of hypertrophied glial cell* as - observed during repealed experiment* Might be explained a* a secordary effect resulting fr<* the degeneration of racrophege* a/vl their possible liberation of more C'oplex, organic C'xspvxjvte. The rather Insignificant degree of lead absorption ty the glial cell*, a* ehovn hlet<*"hemtcally, doe* not seem t > Juetlfy their narked Involvement in the pathological proc*## as It Is observed w> vitro. * -*-* " W-W awjgr^ yrn1 -v.\ .vjUy'jjAi III. "Influence of gerua froa Laboratory Anlaala ar.d Patient* with I*ad Poisoning on Nervous Ttiu In Culturem~. (H.M. Sobkovic*, h.R. Harrey, L.J. Goldvmier t.ft. Peterson and M.fi. Bernstein). Lo c VIj -^j for * different approach to th* proxies of l*s4 poisoning In vitro tested itra obtained fraa laboratory anlaala and patient* with lead poisoning, against nature nyallnated culture* of th* nervous *y*tea. Th* exposure of th* culture* to the** sera resulted In partial or coeplet* degeneration and breakdown of th* ejy* 1 In *heath, which In sw>*t Instances wma acccsspanled by eccplet* preservation of th* nerve cell bodies and axons, but not. of th* glial cell* (which are equally Involved In ayelln oalntenance). V* also deaonstrated that tb lysis and degeneration of central and peripheral ^rclln In response to the presence of the above sera la not correlated to th* actual concentration of lead In these sera. Therefor* it la poealbl# that th* observed Ryelln disturbance was not du* to th* actual presence of lead but rather to oth*r aubstancea seconder 1 ly pr*snt In ,vit* sera. Our conclusions therefor* ar* turnerlt*d as follower 1. Th*r* la evidence In vitro of cntral and peripheral ayelln degeneration and lysis In response to th* presence of t*rua frea lead poisoned patients, and laboratory anlaala. ?. Then* Is evidence that th* degeneration of th* ayelln sheath observed during exposure to lead poisoned sera lji vitro vu not related quantitatively to the actual concentrations of lead Is these *era. Thus It It possible that th* observed ayelln disturbance* wer* not du* to the presence of th* lead but rather to other dletuxbance* secondarily operatic. (cf. Tart I). ci 3. Lead Itself has no direct toxic effect on the nerve cell and th* Myelin i shesth Wi vitro. This aay be du* to th* deacna (rated lack of uptake of th* cetal by th* nerv* cell frest it# envlronaent. (cf. Tart II) h. tt la alto suggested that th# exeesslv* absorption of lead by assenchyaal tissue results In th* degeneratlcn of thl* tissue, (cf. fart II; -i ). A **condary response of th* glia du* to the degeneration of atsenchymal tissue, a* well as Its noderate ability to absorb leal Ui vltrn suy/etta th* Involveaent of this tissue lo th* patholcgy of lead poieonlrg in vitro. (cf. fart* 1 and II) ft, Th* hypcrlheelt ! proposed that th* direct toxicity of leal to^h* nervous tystea Ui vitro 1* correlablt with aeslallatlon of the aetal by Ite v%rlou# ccapcnent tissue!. ..=c I 'j , Jj $sc&. ygyaagiw sx u ml u h .j .imk `emar*