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tance of additional risk factors for cardiovasculardisease. There is evidence that these factors, suchas cigarette smoking, hyperlipidaemia, diabetes,hypertension, obesity, combine with oral-contra-ceptive medication in a synergistic rather thanadditive manner in certain circumstances. 1 Itwould seem wise to consider other forms of contra-

ception for women with more than one risk factorand also for those in the older age-groups. A studyby VESSEY and DOLL 10 of oral contraceptives, in-trauterine devices (i.u.D.), and the diaphragm sug-gests that the excess-mortality rate for oral contra-ceptive users in the United Kingdom may be two toten fold greater than that for women using theI.u.n./diaphragm, depending on the failure-ratesassumed for the various methods. Entirely differentconclusions, however, may be reached in countrieswith higher maternal-mortality rates and where thepopulation may also be less susceptible to vasculardisease. Further experience from the prospectivestudies will allow firmer estimates of the risks oforal-contraceptive medication.On p. 757 we publish two assessments from some

of those closest to the facts who have so far had achance to examine these new data. Their conclu-sions are that renewed caution, more or less, shouldprevail. Those who have even more difficult judg-ments to form are the doctors who must decidewhether or not to prescribe the pill for a particularpatient and also the women to whom they mustexplain the risks. Little in the week’s Lancet is

going to dismay the many women who regard oralcontraceptives as a blessing which carries a minuterisk of premature death. The doctors whose task itis to interpret this news will have to say that thedanger now appears to be greater than it onceseemed, but it is still, in the absence of other riskfactors, very small indeed for younger women (andthe suggested age for reconsideration of contracep-tive method is 30-35).

Aplastic Anæmia: Seed or Soil?To the clinician, aplastic anaemia is peripheral-

blood cytopenia with chronic fatty atrophy of redmarrow. To the experimental hamiatologist, it isthe clinical expression of reduced output from thehsemopoietic stem-cell pool into the more mature,differentiated compartments of the marrow. Thereis, as yet, no technique of counting the ancestral,pluripotent cells in human marrow, and we must

10. Vessey, M. P., Doll, R. Proc. R. Soc. B. 1976, 195, 69.1. Heimpel, H., Kubanek, B. Br. J. Hœmat. 1975, 31, suppl. p. 57.

rely on assays of their closely related descen-dants-those capable of forming granulocytic anderythroid colonies in culture. Most cases of aplasticanaemia show reduced numbers of such colonies.2-4

Leaving aside irradiation and cytotoxic drugs,exogenous toxins can be identified in some 50% of ’

cases of acquired aplasia, though the manner in which they interfere with stem-cell replication anddifferentiation remains an enigma. Some agentsknown to cause aplastic anaemia, such as chloram-phenicol, benzene, and the hepatitis virus, are cap-able of producing chromosomal damage in culturedmarrow cells;5.6 moreover, the persistence of mor-phological abnormalities in the marrow of patientswith partly recovered hypoplasia,7 and the fact thata small number later proceed to paroxysmal noc-turnal htmoglobinuriall or acute leukaemia," maybe cited as evidence of permanently damaged stem jcells. Nevertheless, there are other, possibilities; forexample, BoGGS and BOGGSIO have postulated adislocation of stem-cell kinetics in chronic aplasia,so that differentiation occurs before the parentstem-cell pool is adequately replenished. Again, I

agents causing aplasia might do so by producing Ichanges in the humoral or cellular microenviron- ’Iment which render it inimical to stem-cell growth. I

We have little knowledge about regulation of the !size of the various stem-cell compartments. No Ihumoral factor is known which regulates the div-ision of pluripotent cells; indeed, recent work haschallenged the concept that the hormone erythro-poietin controls the replication-rate of the "com- Imitted" erythroid stem cell,l1 though it certainlyinfluences maturation of its progeny, and some Icases of red-cell aplasia are due to interference with Iits action.12 The cellular or "stromal" component iof the microenvironment is also poorly understood, i,but there exists an animal model which illustratesits importance. The W;WV mouse and the S1/SI’’mouse both have congenital anxmia. The formerhas defective stem cells while the latter has abnor-

,

mal marrow stroma; transplantation of Sl/Sld, ornormal, mouse marrow will cure the anaemia of theW/W mouse, but neither normal nor W/W mar-row will cure that of the Sl/Sld mouse. However,implantation of W/W‘’ spleen tissue will improvethe anaemia, because the S1!Sld stem cells can thengrow within the matrix it provides.13.14 KNOSPE

2. Greenberg, P. L., Schrier, S. L. Blood, 1973,41,735.3. Kern, P., Heimpel, H., Heit, W., Kubanck, B. Br. J. Hœmat. 1977, 35, 613.4. Hansi, W., Rich, I., Heimpel, H., Heit, W., Kubanek, B. ibid. (in the press5. Mitus, W. J., Coleman, N. Blood, 1970,35,689.6. El-Alfi, O. S., Smith, P. M., Biesele, J. J. Hereditas, 1965, 52, 285.7. Geary, C. G., Dawson, D. W., Sitlani, P. K., Allison, H. A., Leyland, M J

Br. J. Hœtmat. 1974, 27, 337.8. Lewis, S. M., Dacie, J. V. ibid. 1967, 13, 236.9. Brauer, M. J., Dameshek, W. New Engl. J. Med. 1967, 277, 1003.

10. Boggs, D. R., Boggs, S. S. Blood, 1976, 48, 71.11. Iscove, N. N. Cell Tissue Kinetics, 1977, 10, 323.12. Peschle, C. Br. J. Hœmat. 1975, 31, suppl. p. 69.13. McCulloch, E. A., Siminovitch, L., Till, J. E., Russell, E. S., Bernstein.

S. E. Blood, 1965, 26, 399.14. Bernstein, S. E. Am. J. Surg. 1970, 119, 448.

749

and CROSBY also showed the dependence of stemcells (or "seed") on an intact marrow microvascu-lature ("soil") by observing the sequence of eventsafter local marrow irradiation, and in experimentsinvolving transplantation of healthy marrow to

irradiated sites. They suggested that some cases ofclinical aplasiain man might result from immuno-logical damage to the marrow sinusoids rather thanthe stem cell itself. IS

If clinical aplasia is due to deficient or defectivestem cells, it should be correctable by transplant ofhealthy cells from a compatible donor, and the factthat such grafts can be established in at least 75%of patients with aplastic anaemia suggests that thisis indeed the basic defect.16,17 (Although vascularand other stromal cells may be carried over withthe graft, the prompt growth of allogeneic haemo-poietic cells suggests that they are nourished in therecipient’s own stroma, vascular cells having amuch slower-rate of turnover.) There are still for-midable immunological problems to be overcome,and when rejection occurs it seems to be related tohistocompatibility differences; however, in a few aninhospitable environment could explain both theoriginal episode of marrow aplasia and failure ofthe graft. Moreover, in every series, a few cases arefound in which grafts are completely unsuccessful,even between identical twins. Although these maybe related to the recipient’s poor condition, or tofailure to transplant enough cells to establish a self-perpetuating stem-cell population, some of thesepatients may have a defective marrow soil. On page742, the Royal Marsden Hospital group report thecase of a young girl with post-hepatitic aplasia inwhom two attempts at marrow grafting, using agenetically identical twin as donor, were only tran-siently successful. However, when the recipient wasprepared for the procedure with powerful immuno-suppressive drugs, haemopoiesis was re-established.Further studies showed that the patient had a cir-culating inhibitor which interfered with the growthof the twin sister’s cells in vitro; inhibition waspartly or completely abolished by growing donatedmarrow in the presence of antithymocyte globulin.There are good grounds for believing that somecases of post-hepatitic aplasia represent an autoim-mune process, and this may have been true here;for example, it has been found that, unlikemost patients with drug-induced and idiopathicaplasia, those with post-hepatitic aplasia may haveautoantibodies to leucocytes and platelets,19 and

15 Knospe, W. H., Crosby, W. H. Lancet, 1971, ii, 20.16 Thomas, E. D., Storb, R., Clift, R. A., Fefer, A., Johnson, F. L., Neiman,

P. E., Leiner, K. G., Glucksberg, H., Buckner, D. New Engl. J. Med.1975, 292, 895.

17 Cline, M. J., Gale, R. P., Stiehm, E. R., Opelz, G., Young, L. S., Feig,S A , Fahey, J. L. Ann intern. Med. 2000, 83, 691.

18 Pegg, D E., Fleming, W. J. D., Compston, N. Postgrad. med. J. 1964, 40,213.

19 Beickert, A., Siering, I. Acta hœmat. 1958, 18, 52.

glucocorticoid therapy has occasionally been bene-ficial. 20

Rather similar observations have occasionallybeen made after allogeneic marrow transplanta-tion : the patient’s own cells have repopulated hismarrow. TERRIT021 has reported two such casesand reviewed others in which this has happened.Evidently the immunosuppressive treatment givento prepare the patient for the graft has suppressedtoxic or inhibitory cells in the marrow environ-ment ; an alternative explanation might be that thetransient presence of infused normal marrow pro-vides a "hormone" or inducer substance to stimu-late recipient cells, or perhaps supplies lymphocytesor macrophages for recognition of abnormal en-vironmental cells.Some patients with aplastic ansemia have a mar-

row lymphocytosis (although others have a lym-phopenia), and this has been cited as evidence of animmunological reaction.22 The possible role of in-hibitor cells in the pathogenesis of aplastic anaemiawas demonstrated by KAGAN et aI.23.24 who foundthat marrow lymphocytes from patients with apla-sia could suppress proliferation of autologous mar-row cells and the differentiation of granulocytes;when these were removed by velocity sedimen-tation, the capacity of the marrow to form granulo-cytic colonies in agar culture was much enhanced.In another experiment, similar to that performedby the Royal Marsden group, these workers showedthat treatment of the aplastic marrow, in vitro,with a cytotoxic antilymphocyte serum which

spared stem cells, allowed the marrow to form col-onies in agar. It was found that approximately athird of patients with aplastic anaemia had marrowlymphocytes which suppressed the growth of

granulocytic colonies on incubation with normalmarrow. 21 More recently, HOFFMAN and his col-leagues26 have shown that the peripheral-bloodlymphocytes from five out of seven patients withaplastic ansemia suppressed erythroid colony for-mation when added to normal marrow in the pre-sence of erythropoietin; interestingly, similarresults were obtained with blood lymphocytes fromchildren with congenital erythroblastopenia (Black-fan-Diamond syndrome). Further work will showwhether peripheral-blood lymphocytes from theadult patients can also suppress granulopoiesis.Obviously, we need to know more about the preciseidentity of these "suppressor" cells in marrow andblood, and to exclude the possibility that they are

20. Bodenbender, R. H. Am. J. Dis. Child. 1971,122,440.21. Territo, M. C. Br. J. Hœmat. 1977, 36, 305.22. Benestad, H. B. Acta med. scand. 1974, 196, 25523. Kagan, W. A., Ascensao, J. A., Pahwa, R. N., Hansen, J. A., Goldstein, G.,

Valera, E. B., Incefy, G. S., Moore, M. A. S., Good, R. A. Proc. natn.Acad. Sci. U.S.A. 1976, 73, 2890.

24. Ascensao, J., Paliwa, R., Kagan, W., Hansen, J., Moore, M. A.,Good, R. A. Lancet, 1976, i, 669.

25. Good, R. A. New Engl. J. Med. 1977, 296, 41.26. Hoffman, R., Zanjani, E. D., Lutton, J. D., Zalusky, R., Wasserman, L. R.

ibid. 1977, 296, 10.

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a sequel to hsemopoietic-cell damage rather than itscaused Nevertheless, these observations haveevoked considerable interest in the possible im-munopathogenesis of the disorder.

Aplastic anaemia is not one disease, but several.The bulk of evidence suggests that most cases

represent damage to the marrow stem cells by en-vironmental agents-perhaps on the basis of gene-tic susceptibility, or following previous depletion bysome unrecognised insult. The best hope for thesemay well be marrow transplantation, if a suitabledonor can be found,27 although it may be possibleto define a group of patients who have a betterprognosis and who can be managed with supportivetherapy.28,29 If we can now identify another groupin which the disorder is secondary to immunologi-cal disturbance in the marrow soil, directed eitheragainst the stem cell or its immediate progeny, oragainst some component of the supporting stroma,this will have important therapeutic implications.

ENDOCRINOLOGY’77

THE expanding research interests of endocrinologistswere reflected at the sixth in a series of biennial inter-national meetings organised by the endocrine unit of theRoyal Postgraduate Medical Schoo1.I In addition to thecustomary discussions of calcium homoeostasis therewere major sessions devoted to the chemistry, biochemis-try, and secretion of peptide hormones, to the molecularbiology of steroid hormone action, and to neuroendoc-rinology.H. Niall (Howard Florey Institute, Melbourne) gave

an outstanding address featuring his recent work withrelaxin, and L. Orci (University of Geneva MedicalSchool) elegantly integrated his startling electronmicro-graphic and chemical studies of peptide-hormone secre-tion. I. MacIntyre (R.P.M.S., London) discussed currentconcepts on the regulation of vitamin-D metabolism.1,25-dihydroxycholecalciferol, 1,25(OH)2D3, is now

known to be the most potent single hormonal factoraffecting calcium balance. Its production in the kidneysis regulated by dietary calcium and phosphorus, byparathyroid hormone, and through negative feedback by1,25(OH)2D3 itself. Work at Hammersmith and in M. R.Haussler’s laboratory (University of Arizona) now indi-cates that, in states of calcium stress such as growth,pregnancy, and lactation, the most important regulatorsof 1,25(OH)2D3 biosynthesis are prolactin and growthhormone, possibly acting via somatomedin.The keynote of this year’s meeting was molecular

endocrinology, highlighting recent work which aims tointerpret endocrinological phenomena in molecular

27. U. C. L. A. Bone-marrow Transplant Team. Lancet, 1976, ii, 921.28. Lohrmann, H-P., Kern, P., Niethammer, D., Heimpel, H. ibid. p. 674.29. Mathé, G., Schwarzenberg, L. ibid. 1977, i, 1361.1. MacIntyre, I., Szelke, M. (editors) Molecular Endocrinology. Amsterdam (in

the press).

terms. A fine example of this was the plenary lecture byB. W. O’Malley (Baylor College of Medicine, Houston)describing a molecular model for the mechanism ofaction of steroid hormones. The progesterone-receptorprotein of chick-oviduct cytosol comprises two non-identical subunits A and B, each binding one moleculeof progesterone. This hormone/receptor complex istranslocated into the nucleus where the steroid/Bsubunit complex selects, and binds to, an acceptor siteon the non-histone protein component of chromatin. Inso doing, it promotes the binding of the hormone/Asubunit complex to D.N.A. at a regulatory site. This, inturn, facilitates binding of R.N.A. polymerase to an in-itiation site for R.N.A. synthesis, followed by transcrip-tion of the hormone-responsive genome (e.g., thatcoding for ovalbumin).

Neuroendocrinology has been the most rapidlyexpanding branch of endocrinology since the advent ofhypothalamic releasing hormones in the late 1960s. Asecond phase of rapid growth followed with the dis-

covery of the opiate peptides in the brain. J. Hughes(Marischal College, Aberdeen), whose discovery withKosterlitzZ of the enkephalins in 1975 opened up thisnew subject, spoke about their possible physiological sig-nificance. Biosynthetic studies in rat brain and gutshowed that incorporation of the radiolabelled amino-acids methionine and leucine was proportional, respec-tively, to the ratio of endogenous methionine5 andleucine5 enkephalins in both tissues. Hughes postulatedthe presence in the gut of a distinct protein precursor toleucine5-enkephalin.

R. Guillemin (Salk Institute, La Jolla) introduced theendocrinology session with a brilliant plenary review ofthe endorphins. The Salk group, jointly with scientistsfrom Denver (University of Colorado Medical Center),3has just completed the isolation from mouse pituitarytumour cells of a protein (molecular weight approxi-mately 31 000) which may be the common precursor toboth corticotrophin (A.C.T.H.) and p-endorphin. Guille-min drew attention to parallel changes in pituitary levelsof A.C.T.H. and p-endorphin (derived from p-lipotropin)in response to stress, adrenalectomy, and dexamethasoneadministration. Release of A.C.T.H. induced by a purifiedcorticotrophin-releasing-factor preparation (alsoobtained at the Salk) was accompanied by a simul-taneous release of p-endorphin.

D. G. Smyth (National Institute of Medical Research,London) presented new data on the enzymatic stabilityof &bgr;-endorphin (also called the C-fragment 61-91 ofp-lipotropin which was first isolated in his laboratory4 inearly 1975). It w as found to be unusually stable toamino- and carboxy-peptidases and to a membrane-bound enzyme from rat brain, whereas its N-terminalpentapeptide sequence, methionine-enkephalin was

rapidly degraded by these enzymes. This stability, attri-buted to the peculiar native conformation of &bgr;-endor-phin, may explain its long-lasting analgesic effectobserved after intraventricular administration. How-

2. Hughes, J., Smith, T. W., Kosterlitz, H. W., Fothergill, L. A., Morgan,B. A., Morris, H. R. Nature, 1975, 258, 577.

3. Mains, R. E., Eipper, B. A., Ling, N. Proc. natn. Acad. Sci., U.S.A. 1977,74, 3014.

4. Bradbury, A. F., Smyth, D. G., Snell, C. R. in Peptides: chemistry, structureand biology (edited by R. Walter and J. Meienhofer). Ann Arbor, 1975.