Histol Histopath (1991) 6: 251 -256 Histology and Histopathology

Amitosis in human adrenal cells

Maria C. Magalhies, D. Pignatelli and M.M. Magalhies Institute of Histology and Embryology of the Faculty of Medicine and Center of Experimental Morphology of the University of Oporto (INIC), Porto. Portugal

Summary. Adrenal pieces obtained from 3 fcmalc ancl 2 (cr!'l~to~i~itosi\)(Elias ancl Hyde. 1983). Yet. for other ~nirlepatients sho\\'cd morpl~ologicirlfigures of umito4is a~lthors.amitosis i4 OIII!, ohscrvecl in lower invertebrates in aclrcnirl zona retic~~l:~riscclls. Such aspects were or ~~nclcrp;rtliological conditions (Yicluan and Hinkung. ohscr\~~~clin both normal and hyperactive aclrcnuls. 1086). This ma!, hc the reason wh! there 112s been :I Nuc,lci appeal-cd constricted. hea\ ill stained. decrease in ~rmitosi4~.csearch (Yinclu~un ancl Bink~rng. \\.it11 coarse chromatin. sometimes scattered among I OSCl) . c!toplasmic organcllcs. hut never niarginating in .l'hc 1lt1r\x)sc oftthe present report i\ to clcscrilw cre4ccntic cirps. mol-p11ologic:ri acll-cnal nuclcur aspects. not !,et ol3ser\,cd C'lea\:~gc 01' the cells originated t\\lo hnl\,cs \\it11 a in li~~rnan:~drenaI. ancl to cliscuss their possillle 1'~1nctionaI nucleolus in each pole. Rin~lcleateclcclls \vcl-c also seen significance. in zona rcticularis. -1~Iie meaning ot' nmitosis in human aclren:rl i4 Materials and methods cIisc11~4ccI. FIurn;r~i aclrCn;ll tissue was obtainctl from t\vo nl:~le Key words: Human adrenal. Aniito5is. Ultra4tr11ct~11-C p;lticnts agecl 32 and 43 ancl three l'cm:rlc patic~its~rgcd IS. I9 ancl 71. 'Thc~male p~~ticntssufI'el.cd from B~~ergc~-'s Introduction dise~~se.tll~. two !.oLlngcr \\omen from ;rcroc!,;~nosis ancl the older OII~from :I virilization a!.nclrome. The patients In recent !'cars little attention has been paid to the received thiopentc~nc (Pentotlial). s~~ccin!Icholinc biological meaning of aclrennl . (Scolinc). di~rll!-nor-toxiferine (Allofcrine). and Latcl!:. \\,c ha\e hacl thc opport~lnityto stutl) the pctliirlinc as ;rn;~t.sthctic drugs. Patients ~~ncler\\~cnt ~~irc~irlcortex fro111 ;I woman \vho untlerwent ~lnilatcral ~~tlrenalcctomyas therap! for the disease. adrenalcctom! as therap!. for virilization 4yntlromt.. We Immetli~rtcl! ~~l'tcrremoval. adrcnals were cut into tia\,e found a special t\.pe of nuclear morphology \vhich small piece4 and irnmersecl in Bouin's solulion ior seemed to be figures of amitc>sis (Prenant et al.. 1004: light mic1-o4cop! ~rnrl 2.5%) glutaraldeliycle in 0.1 M Hueher. 1047. IO(2. 1963: Ohr2 et al.. l96(>: Pessacq. cacc~d!.late IILI~~CI-fol-electl-onmicroscopy: the tr;~gments 1900: Tippit Lrncl Pickctt-Heaps. 1976: Ferg~lsonirntl fixed in gl~lturaldeh!~dcwere postfixcd in 1':6 osmi~lm Palm. 1976: Rargmann. 1977: Tucker et :[I.. 1980: Chrn tctrosidc in \~eronal-;rcetateI~LI~~CI-. After deh!dr:rtion and Wan. IOSO). Such peculiar aspects wcrc further sccn in 21 g~-adedscric4 01' cthanols. the specimens \\.ere in human adl.cn:rl of t\vo male patients who t~nclc~.uent cmbetlcled. rcspecti\,cly. in paraffin ancl Epon 812. unil~rteral aclrcnalcctomy as therap!, for Rucrgcr's Semithin 4cctions. 1 or 2 pm thick were stai~rctlwith disease and in two female patients \\.it11 acrocyanosis. metl~!Icncbluc-~/LII-I1 (Richartlson et al.. 19hO). Somc a~~thorsdeny the occurrence of amitosis in Ultrathin sections were doul3le stained with uranyl nianlm~tls (C;r),nfeltt. 1032: Obcrling and Rcl-nhard. acetate ant1 lead citrate and the specimens were ollscr~ed 1001: Buchcl.. 1003. 1963). whereas others consider that in a Scol 100 L3 electron microscope. it i4 the onl!. t!pe of nuclcar division in adult normill li\.er Results Offprint requests to: Dra. Mar~aC. Magalhaes, lnst~tuteof Histology By light rnicl-oscopy. the adrenal cortex looked and Embryology, Faculty of Med~cine of Oporto. 4200 Oporto, no~.rnalin both n~alcpatients and in the t~oJoungest Portugal women suffering from :rcrocq.:rnosis (Fig. l). In the Amitosis in human adrenal

Fig. 1. Adrenal cortex from a woman aged 18. Adrenal cortex looks normal. A small area of adrenal niedulla (M) is present. (H + E) X 360. ZG - zona glornerulosa: ZF - zona fasciculata: ZR - zona reticulans.

Fig. 2. Adrenal gland from a woman aged 21 with virilization syndrome. Hypertrophy of zona reticularis (ZR) IS evident. (H + E) u 240

Fig. 3. Zona reticularls. Constricted nucleus (arrow). (H + E) u 1,470.

Fig. 4. Apoptos~s. Note heavlly stalned nuclel (arrows). (H + E) 1,470

\vornan with \.irilization \ndrome. thc Iona reticulal-is (Fig. 4). Somc apoptotic boclics appeared to pas4 into scenied larger (Fig. 2). the sinnsoids. In all patients images of binuclcatcd cells ucre By electron microscopy some nuclci cshihitcd observed in thc :~drenal cortex. most of them being coarse chromatin pattern and perinuclc;~r chromatin located in the zone I-cticularis. and the zona fascic~l- appcared heavily stained. The do~~blt:IILICIC;I~ lata: sometimes nuclei were dumb-bell-sliapcd (Fig. 3). membrane was sometimes discontinuous. and denw Though seldom seen. cells undergoing and chromatin masscs lay among cytoplasmic orga~icll~s apoptotic bodies were observed in thc zone reticularis (Fig. 5). Only a few images of karyorrlicsis ucrc Amitosis in human adrenal

Fig. 5. Virilization syndrome. Complex structure of a cell nuc (N). Dense chromatin masses lie among cytoplasm organelles (arrows). x 15,000. nu - nucleolus.

observed. Chromatin never marginated in crexentic On the other hand. for many authors, amitosis only caps, like the morphological feature of apoptosis in occurred in lower invertebrates, aged cells or in electron microscopy. Sometimes a constriction appeared pathological conditions (Prenant et al., 1904: Bucher, in the nuclei and two daughter nuclei appeared to be 1963): for others, it was even considered as the only type formed (Figs. 6-9). The cells showed a cleavage furrow, of nuclear division in normal adult liver (Elias and Hyde, the nucleus being perpendicular to it (Figs. 6-9): the 1982). In addition. amitosis has been studied in living cleavage deepened and widened, the neck decreased in cells cultivated in vitro (Bucher, 1963). and Bargmann diameter becoming *stretched>>along the longitudinal (1977) referred to the fact that it was often seen in highly axis of the cell (Figs. 6. 7). Such a cleavage may lead to differentiated tissues, such as the liver, heart and kidney. the formation of two sometimes unequal halves with one Accordingly, amitosis, in the classical sense. does seem nucleolus in each pole of the nucleus (Fig. 6) and. in the to play a role in the multiplication of cancer cells; a end, nuclear would seem to occur. No chromo- desdifferentiated cell (Oberling and Bernhard. 1961). somes were detectable within the nucleus at any stage. Concerning the adrenal, amitosis has been observed in The nucleolus in cells with a bilobate appearance of interrenal tissue of Rana temporaria (Pehlemann. 1068) amitotic division was well preserved, and contained and in rat adrenal zona glomerulosa after a prolonged fibrilar centres (Fig. 6). Nucleolar material was never low-sodium diet (Palacios et al.. 1976), but it has never scattered through the cytoplasm except in karyopyknosis. been reported in human adrenal cortex. as far as our Cytoplasmic organelles were well preserved and literature suggests. bundles of parallel microtubules were never visible near In this manuscript, morphological aspects of nuclei, the nucleus. which might be considered suggestive of amitosis. were observed in human adrenal zona reticularis. a Discussion differentiated zone of low mitotic activitj (Ford Amitosis or direct division (Prenant et al., 1904; and Young. 1963: Wright, 1971), but which in some Obre et al., 1966) has been postulated as a form of cell circumstances can be stimulated and submitted to an division, and in many cases it appears to be a kind of increased metabolic activity. physiologically-induced tissue proliferation (Yiquan Curiously enough, the zona reticularis is also the and Binkung, 1986). However many questions exist with cortical zone which exhibits more apoptotic bodies; a respect to the significance of amitosis, and even today sign of programmed cellular death (Wyllie et al., 1973, there is no truly satisfactory research concerning this 1980). However, apoptosis has peculiar morphological type of cell division (Bucher, 1963). In fact, large aspects which are easily distinguished from those of numbers of observations described as amitosis were amitosis. such as margination of chromatin in crescentic really not, often merely being artifacts of fixation, caps and cytoplasmic densification. mechanical influences. etc. (Bucher. 1963: Puza. 1969). Molecular mechanisms of amitosis in mammals are Amitosis in human adrenal

Flg. 6. Zona retlc~llarlsfrom the adrenal cortex of the woman wlth v~rll~zatlon syndrome A constrlctlon (arrows) provokes the appearance of two unequal halves of the nucle~(N N') with one nucleolus (nu, nu') In each pole X 10 800

Fig. 7. Buerger's disease. Man aged 23. A cleavage furrow IS observed (arrows), the nuclel [N and N') belng perpendicular to t. 15,000

u~ihnown: in fact. \ve tlo not hno\\ \\li;~t hilitl ol might he intCrprclctl a\ :~mirosiswere scarce*. inductive signal can cause initiation of' arnitosis: We can slxu~latethat amitosis. like apoptozi4. neither do we know if amitotic nl~cleu\i3 cuploid or 0cc111.s in :~tlrcnal~.cticular cclls as part of tlici~~io~'ni:rl suffe1.s subsequent degeneration. In tlic prc\cnt tlc\,clopmcnt ancl i\ ~lrohahly correlatecl \\it11 :In study lrmitosis was ob\ervecl both in nc~rrii:~land in acti\ation ]Truce\\ or nnc Ic;rcling to cell dc:rth. in hyperacti\,c adrenal. and in both ~nirle\and fe~m:~lcs. rclalion to the maturation \t;rtc ol' thc glandular cell. Hence. ~rmitnsisseems to be a form of nuclcar tlivision If this gl;~ndularcell i\ immature. death will occur. in human adrenal. being perhaps the fa\tc\t fol-ni of while aniirosi\ stem\ to occur in niatul-c cclls. Hence. cell division. hut not the usual form siiicc i'igurc\ that the t!,pc of ccllular responses will depend on tlic 3!UOlO3 U! (~~slew!saloal3n~>>)sluau6e~) leal3nu 40 ay3s!doyso~y!w pun a!60lols!H '(~~61)'M uueu6leg pealds pue uo!le~edas '(7,861) 'm'a apA~pue .H 'LLS-LLS 'OS 'IsaAul .qel .Aydo~yadAy 41~016101 xapu! ut, 'lel leleuoau ay$ U! sip3 le!pie3oAw paleal3nu!q 10 uo!leuloj '(9861) 'd's doys!~pue .lr.r'j qqn13 .opodo )o Al!sla~!unaql uo~)palold ~8166a41 Aq pauoddns Alued seM Apnls s!ql ,uoddns le13ueu!)l!aql 40) (le6nuod) ,,lzopues '9L-69 'L11 'leut, ept, ,ado3so~3!wuolpala uo!ss!wsueJl pue ly6!1 sol3npo~d yueql aM .ld!l3snuelu aq] 6u!dAl JO) e~!a!ue~elesalal e!JeN pue sqde~6oloqdaql JO) elso3 ou!lsnej e!lel/y euV 'a3uels!sse ay1 lapun Apnls 'v.1 '1la3 ue!leuwew ay1 10 snapnu ay4 le3!uq3al JO) eJ!alJaj e!laLuV e!leW 'J~!J~SIIU~Uaql )o uo!le~edald 6u!lnp 40 s!sol!we uo Apnls t, '(9861) 'y'g ueM pue '0.h uay3 uolleladoo3 40) he~6luaz.H .Ja yueql sJoqlne aql '5']~3~36p3/~0~y3!j 'LZE-ELE dd 'ssald 3!uape3t, jpa) '3.r'~s!JleH 'I'IOA 'uo!s!~!pIla3 pue ~IMOJ~lla3 :ul 11a3'30s 'lul ay1 40 e!soduAs .asol!ue,l ap aualqold a1 '(896~)'0 ~ay3ng .eie[n3!3~1?jeuoz U! '(~861'uaisa!q ::p861 'PLL-ELL 'QP wut, 'doqs!a put? qqnl~)s!soi!w~? iCy pasne3 iC~q!ssod -sl[a3 el3v 'asol!ue,l ap aualqold ne uo!l3npo~lul'(196 1) '0lay3179 palus[snu!q a~~ssqooi [ensnun jou S! I! a3u!s 'sauoz '~9-09'P xut, evt, ~~1.103IFU~J~~I? lay10 U! id!~ssnuems!qi u! paq!l3sap .au!3!y3103 10 uo!lells!u!wpe salde salA3o~q!jap sa~nlln3 sr? sa!%oloqdlom lealsnu ay1 loj qslcas oi lap10 sap suep sanb!lol!ue sal!e?(x~usuo!s!A!a '(~~61)'0 lay3ng u! ,i.~essasauale sa!pnis laqlln-~ .waisiCs aunww! u! 'E9- L9 dd 7-~e611nis qieap .1t?[nlla3loj (686 1) u!aislot) ,iq pasodold SEM s~qi '6ella~'awa!ayl 61oa9 '~~nt,'L uayx.-ual/y sap a!woleut, su q3ns s!saqlod,iq v ,1133 ay1 jo aleis [r?~uarudola~ap

00L'OZ X .snapnu aql leau paAJasqo ale salnqnlol3!~1ON '(N) snapnu ay1 U! alqel3alap ale (N) sa~~oso~~o~q~ ON .Jaiaure!p a6~ele 4l!M y3aU !(SMOJJ~) MOJJn) a6e~eal3'8 .6!j JO) se aues aql '6 '6!j

leua~peuewny U! s!soj!w~ SSZ 256 Amitosis in human adrenal

neoplasms. Hum. Pathol. 13, 635-639. von Rana temporaria, L Z. Zellforsch. 84, 516-548. Ferguson F.G. and Palm J. (1976). Histologic characteristics of Pessacq T.P. (1969). Special forms of amitotic nuclear division cell cultured from rat placental tissue. Am. J. Obstet. in striated muscle and other insect tissues. Experientia 25, Gynecol. 124, 41 5-420. 977-978. Ford J.K. and Young R.W. (1963). Cell proliferation and Prenant A., Bouin P. and Maillard L. (1904). Division directe ou displacement in the adrenal cortex of young rats injected amitose. In Traite dlHistologie. Tome 1 Cytologie Masson & with tritiated thymidine. Anat. Rec. 146, 125-137. Cie, eds. pp 761-767. Golstein P. (1989). Morts cellulaires et systeme immunitaire. Puza V. (1969). Sources of mistakes in the interpretation of the MedicineISciences 5, 546-553. appearances of amitotic division. Folia Morphol. (Praha) 17, Grynfeltt E. (1932). Sur la valeur histogenetique de I'amitose 66-67. dans les hyperplasies glandulaires de I'endometrite Richardson K.C., Jarret L. and Finke E.H. (1960). Embedding in chronique et des adenofibromes mammaires. Compt. Rend. epoxy resins for ultrathin sectioning in electron microscopy. Assoc. Anat. 27, 343-351. Stain Technol. 35, 313-325. Kriesten K. (1984). Relative incidence of and Tippit D.H. and Pickett-Heaps J.D. (1976). Apparent amitosis in binucleated cells, nuclear volume and nucleolar rate per the binucleate dinoflagellate Peridinium Balticum. J. Cell nucleus in the mammary gland epithelium of the mouse Sci. 21, 273-289. during differentiation in the gestational and lactation phase. Tucker J.B., Beisson J., Roche D.L.J. and Cohen J. (1980). Gegenbaurs Morphol. Jahrb. 130, 307-314. Microtubules and control of macronuclear ~~amitosis~~in Oberling Ch. and Bernhard W. (1961). The morphology of the . J. Cell Sci. 44, 135-151. cancer cells. In: The Cell. Vol. 5. Brachet J. and Mirsky A.E. Wright N.A. (1971). Cell proliferation in the prebubertal male rat (eds). New Academic Press, Inc. N.Y. pp 405-496. adrenal cortex: an autoradiographic study. J. Endocrinol. Obre A., Campan F. and Chanton R. (1966). La multiplication 49, 599-609. cellulaire. In: Biologie Cellulaire. 3a ed Doin (ed). pp Wyllie A.H., Kerr J.F.R. and Currie A.R. (1973). Cell death in the 631 -698. normal neonatal rat adrenal cortex. J. Pathol. 11 1, 255-261. Palacios G., Lafarga M. and Perez R. (1976). Ultrastructural Wyllie A.H., Kerr J.F.R. and Currie A.R. (1980). Cell death: The study of binucleation in cells of the rat adrenal glomerular significance of apoptosis. Int. Rev. Cytol. 68, 251-306. zone after a prolonged low-sodium diet. Experientia 32, Yiquan Ch. and Binkung W. (1986). A study on amitosis of the 909-91 1. nucleus of the mammalian cell. Acta Anat. 127, 69-76. Pehlemann F.-W. (1968). Die amitotisch Zellteilung. Eine elektronenmikroskopische Untersuchung an lnterrenalzellen Accepted December 5, 1990