important behavioural changes could be subsequently measured; courses were not designed to produce the changes that researchers measured. Refresher courses produce educational and social benefits that may not have any observable influence on the quality of care for many years. Informal communication and "impressionnaire" evaluations indicate that physicians believe refresher courses are of much value. Adequate budgets for evaluation of courses are rarely provided. To carry out a thorough evaluation of short courses would increase the operational costs beyond what practising physicians would be willing to pay.

Recent studies suggest that careful preselection of important achievable behavioural objectives can lead to the accomplishment of appreciable improvements in physician performance. We believe that it is practical to introduce this procedure into the planning of refresher courses. We are impressed that the preselection of both learning and behavioural objectives can greatly enhance the efficiency and effectiveness of all programs in continuing education. The key to success is correct identification of the participants' current learning needs. The major objectives of refresher courses should be confined to a relatively few specific changes that

are achievable with the time and resources available. Both students and teachers must be thoroughly familiar with and agree upon the objectives before the course begins. There is a great need for financial support for model programs in which the development of practical evaluation systems is emphasized. This would be an excellent project for national organizations concerned with education and the quality of medical care. O.E. LAXDAL, MD, FRCP[CJ, FAAP

Director Continuing medical education University of Saskatchewan Saskatoon, Sask.

Individuality in disease and therapy Clinical trials provide the objective basis for selection of optimal treatment. A particular treatment is adopted if a significant difference in results between a treated and an untreated or differently treated group can be observed. The superior therapy is identified in terms of the proportion of cured or improved patients, though this proportion will rarely reach 100%. Consequently, a treatment selected by clinical trial may be inappropriate for some patients who cannot be identified in advance. Such treatment failure is due to the heterogeneity of the disease or of the response of individuals to the disease or the treatment, or is due to both factors. Better definition - that is, identification of homogenous disease entities - is an age-old objective of clinical research. Recently this has involved identification of molecular factors, as in the advances in the understanding of inborn errors in cascade reactions such as blood clotting and complement activation; in these diseases the failure of one component may cause serious disturbance. Yet definition based on identification of the affected component is only the first step to classification that is adequate for decisions on treatment. In hereditary angioedema the Clesterase inhibitor may be absent (eniotypy), or may be present but functionally incompetent (allotypy).16 In allotypic disease there is immunologic tolerance to the major portions common to competent and incompetent

molecules. Consequently, replacement therapy with the normal factor can be used repeatedly in treatment of clinical crises. In the eniotypic disease, where there is no tolerance, repeated replacement therapy may result in resistance due to immunization, thus becoming ineffective and possibly dangerous treatment. Despite refinement in the definition of specific diseases, and development of adequate therapy, there will remain some patients who respond abnormally to treatment. The cause for these variations in response may be complex. The approaches to analysis of this problem may be illustrated by relatively simple situations of variable response to drugs. The muscle relaxant succinylcholine causes prolonged apnea in approximately 1 out of 2000 individuals and is ineffective in another small proportion of patients. Both of these undesirable responses are related to individual molecular differences - the polymorphism of cholinesterase - and can be predicted by determining the polymorphic phenotype of the patient.7'8 Therefore, the drug can be used safely in patients who have the common type of the enzyme, as long as there is no muscle abnormality or electrolyte imbalance,9 but it should be avoided in patients whose enzyme abnormality results in inadequate metabolic breakdown of the drug. Thus, one can expect that the administration of the drug will have the desired effect

if drug and dose are prescribed on the basis of previously determined enzyme polymorphism. The relation between untoward drug effects and genotype can be less direct: some forms of polymorphic enzymes affect membrane stability, so that drugs can damage membranes although there is no action of the enzyme on the damaging drug.7 Environmental agents also interact with genetic material in a way that determines whether or not disease is induced. Aryl hydrocarbon hydroxylase (AHH) is one of the mixed-function oxidases in the microsomal fraction of many mammalian tissues.'0 While its exact physiologic role is not understood, this membrane-bound enzyme is known to hydroxylate a wide variety of exogenous hydrocarbons, including many drugs, insecticides, steroids and chemical carcinogens.""' In the initial step in the microsomal oxidation of polycyclic hydrocarbons, which is catalyzed by AHH, epoxides are formed;'3 they are considered to be the active form of some carcinogens.'4"' Hydrocarbon substrates,'6 barbiturates and other chemicals'7"8 act as inducers of AHH. The extent of induction differs among individuals.'9 In the mouse it is controlled by a single autosomal locus,'9" and the carcinogenic potency of polycyclic aromatic hydrocarbons is related to high AHH inducibility.22'. In short, disease induction is related to polymorphism in the tissue concentration of inducible enzyme, and dis-

CMA JOURNAL/JULY 12, 1975/VOL. 113

11

can be predicted by of induced enzyme. A similar situation may exist in man. Tokuhata24 found excess mortality among both smoking and nonsmoking relatives of probands and concluded that susceptibility to lung cancer is inherited. Possibly induction of lung cancer in smokers depends on the rela¬ tive concentrations of epoxide-forming and -degrading enzymes in the affected tissues. Such genetic variations in hydrocarbon hydroxylase have been ob¬ served in human lymphocytes,25 and in one study the incidence of AHH inducibility in a group of 50 patients with bronchogenic carcinomas was sig¬ nificantly higher than expected.26 Much information remains to be assembled and other explanations cannot be ex¬ cluded,17 but another marker for dis¬ ease susceptibility in man may have been found. One of the indications that variations in response of the individual are con¬ trolled by different alleles at the same locus (i.e. that the response is poly¬ morphic) came from studies of the genetic variations in immune response. The extreme consequences of polymorphism are encountered in genetic deficiencies affecting the development of thymus-derived or bone-marrowderived lymphoid cells, or of both, or as in IgA deficiency affecting the late stages of cell differentiation.28'29 Certain types of agammaglobulinemia appear to be attributable to the effect 8f suppressor cells, which prevent anti¬ body formation by otherwise normal

susceptibility

ease an assay

.

.

lymphoid cells.29j30

The molecular basis for these defects is not known, but a recent develop¬ ment may be the first step towards the molecular definition of one defect of the lymphoid system. Adenosine deaminase deficiency has been discovered in children with combined immunodeficiency.31'32 At first it seemed pos¬ sible that this might be a marker, that is, jthat correlation might be the result of genetic linkage. However, it now appears more likely that a functional connection may have been discovered, for it can be shown that increased cyclic adenosine monophosphate (AMP) inhibits cell-mediated immunity. Thus, adenosine deaminase deficiency might result in immunodeficiency through an increase in the intracellular concentra¬ tions of cyclic AMP.33 When the molecular basis of genetic defects has been determined we shall have markers for a new set of cellular properties and shall be able to investigate polymorphism in the healthy in¬ dividual and whether it is a controlling factor in disease induction or progres¬ sion. Presently the most interesting ge¬ netic variations are not those that them¬ 12 CMA

selves can result in disease, but those that manifest polymorphism within the normal range. There are polymorphic genes that affect the general capacity for response to many different anti¬ gens34 and there is a polymorphic gene region that controls response to indi¬ vidual determinants.3* Genes less well studied that control tolerance and toler¬ ance breakdown may be important in the propensity to autoimmune dis¬ ease.36-38 Many other polymorphic cell properties may be relevant to a con¬ sideration of pathologie processes. To relate onset and progression of disease to genotype would obviously represent a major step toward the ef¬ fective selection of those individuals who are likely to respond to a treat¬ ment known to be beneficial in a ma¬ jority of patients. The type of infor¬ mation required to take this step can be illustrated by an examination of the factors that determine whether a given virus can cause mouse leukemia and whether the leukemia will have a fatal outcome. Mouse leukemia viruses can be divided by their infectivity into Ntropic and B-tropic variants. The alleles of the gene locus Fv-1 determine whether a given virus can infect a

of a particular genotype (Table Because of the genetic linkage between the gene locus Fv-1 and the locus controlling the polymorphism of glucose-6-phosphate dehydrogenase mouse

I).39-41

(Gpd-1)42 one can predict from the polymorphism whether a given virus will be able to multiply in a mouse of a particular strain. Another gene locus, Fv-2f determines whether Friend virus43

will cause erythroleukemia, as meas¬ ured by the spleen-focus assay.39'44 This locus is linked with a locus that de¬ termines the polymorphism in malic de¬ hydrogenase (Mod-1).4* Thus, from knowing the polymorphic form in which malic dehydrogenase appears in

particular

a

mouse,

one can

predict

whether a virus will induce erythroid disease. Whether disease progresses or

depends on polymorphism at locus,46,47 part of the link¬ age groups on the 17th autosome, de¬ signated the H-2 complex (Table II).48 In this region the major transplantation antigens are determined. Therefore, in¬ formation on the polymorphism of glucose-6-phosphate dehydrogenase, malic dehydrogenase and H-2 specificity per¬ mits a reasonable prediction as to whether a given virus will infect a par¬ ticular mouse and whether it will cause progressive disease. Because of crossovers it is likely that linkage groups will occur in outbred animals in many possible combinations. Only very close linkage with a very low crossover probability will be de¬ tected and be useful for predicting pro¬ gression through disease processes. Linkage disequilibrium is an important factor in maintaining associations. It becomes difficult to distinguish be¬ tween a gene locus that affects disease directly, a gene locus that affects dis¬ ease indirectly and a gene product that has no functional connection with the disease process but is the product of a closely linked gene locus. Nevertheless, it appears possible that a constellation of polymorphic markers could serve to regresses

a

different

Table I.Genetic controls of murine leukemia

*See Table II Table II.Genetic map of the H-2

Complex Ends

Regions Subregions Loci

JOURNAL/JULY 12, 1975/VOL. 113

complex H-2

K

D

K Ir-IA H-2K

Ir-IA

H-2I

Ir-IB Ir-IB

Ss, Slp

H-2D

circumscribe elements of response that are relevant to progressive disease or would identify individuals who have an inherently high risk of contracting a disease. The process by which this might be done is like the process by which paternity is established - the use of markers that do not play a direct role in the biologic process. The larger the number of genetic systems, the smaller the chance of erroneous identification of the father.49 None of the markers is responsible for fertilization, yet considered along with other evidence they can contribute to the identification of the origin of the fertilizing sperm. The first correlations between disease incidence and cell markers were observed in studies of the ABO blood groups.50 A second set of correlations has been obtained from a study of the products of the major transplantation locus, HL-A,51 which indicates that not only incidence but also progression through disease is linked to the HL-A region and that, in at least one instance, this correlation is so high as to make the determination of the HL-A product diagnostically valuable. Advancement in this field will depend on studies that relate a variety of gene products to disease. Some of the tasks in this area are already discernible from animal experiments. Foremost is a study of the normal lifespan of different cells of different individuals. Indications of independent controls for different cell lines have come from studies of tetraparental (allophenic) mice.52'53 Such animals are obtained by aggregating two genetically dissimilar blastomeres and culturing them to form a single embryo. When the genetic composite has reached the morula or blastocyst stage it is transferred to the uterus of a foster mother. Each of the resulting animals originates from two embryos and thus from four parents. Clonal histories can be detected by means of appropriate genetic markers. For example, all pigment cells in the adult coat were found to be donally derived from only 34 genetically determined primordial melanoblasts.55 The relative predominance of cells of different genotypes can be studied as a function of age. In general, within a given cell type, cells of one genotype may predominate at one time in the life of the animal and at a later stage cells of another genotype may predominate. This has been observed with spermatogonia and erythroid cells.53'54 It appears as if each cell type was programmed genetically in terms of replication and in terms of cell survival. This programming is unique for each differentiated cell line of the same individual; that is, the controls of the

different cell types appear to be independent of one another. We have all observed this phenomenon among our colleagues who have run out of adequately functioning hair follicles while retaining functional cells and regenerative capacity in most other parts of their body. Indeed, there appears to be a programmed quota of cell divisions for each tissue, and external provocation of increased doublings seems to result in faster exhaustion of this quota, that is, in premature ageing. This may well happen in grafts and may be responsible for the severe atherosclerosis found in renal and coronary arteries of some kidney and heart transplant patients.55'56 Indeed, the reaction between tissues that are not histocompatible results in increased cell divisions, as is known from the experimental model of the graft-v.-host reaction. In addition to cell line controls, there may also be genetic controls of senescence that are common to the entire organism. Normal diploid fibroblasts have a finite ability to replicate in vitro, which is related to the age of the donor.57 As cultured cells approach cessation of mitotic capacity, the heatlabile fraction of cell enzymes increases and the capacity of fibroblasts to increase the rate of clotting and the rate of clot retraction decreases. In all these respects, diseases of premature ageing, such as progeria, Werner's syndrome and diabetes mellitus, result in cell changes that are similar to those found in cells that have been aged in culture.57'58 Thus, a variety of genetic controls appears to exist and to determine the rate of differentiation, replicative phase and progression to senescence. All these properties may affect initiation of a pathologic process and progression through disease. The discovery of cell markers for these genetic controls, possibly differentiation antigens of cell membranes, is a challenging task. The way is open for an orientation of our science policy towards a study of genetic markers for response, disease susceptibility and progression of a cell through normal life and through disease processes. The results of such studies would provide the scientific basis for therapy based on molecular individuality and would increase the probability of benefit to the individual when the results of clinical trials are applied to him. It might also assist in the identification of populations "at risk" and in the development of a new dimension in the practice of preventive medicine. B. CINADER, D SC, FRSC

Director Institute of immunology University of Toronto Toronto, Ont.

References 1. CINADER B, DUBIsKI 5, WARDLAW AC: A complement and antigen defect in certain inbred strains of mice; an instance of eniotypy, in Studies of Rheumatoid Disease: Proceedings of the Third Canadian Conference on Research in the Rheumatic Diseases, Toronto, Feb 25-27, 1965, U of Toronto Pr 1966, pp 202-19 2. Idem: Allotypy and eniotypy. Nature (Lond) 210: 1291, 1966 3. CINADER B: Immunochemistry of enzymes, in Methods of immunology and immunochemistry, vol 4, edited by CHASE M and WsiLIAMS C, New York, Acad Pr, 1975 4. RATNOFF 0: Progress in Haemostasis, edited by SPAET JH, New York, Grune, 1972, p 39 5. DONALDSON VH, EVANS RR: Biochemical abnormality in hereditary angioneurotic edema. Am .1 Med 35: 37, 1963 6. ALPER CA, ASRAMsON N, JOHNSTON RB, et al: Studies in vivo and in vitro on an abnormality in the metabolism of C3 in a patient with increased susceptibility to infection. J Clin invest 49: 1975, 1970 7. KALOW W: Pharmacogenetics: Heredity and the response to drugs. Philadelphia, London, Saunders, 1962 8. Pharmacogenetics: Report of a WHO scientific group. WHO Tech Rep Ser #524, Geneva, 1973 9. international Symposium on Malignant Hyperthermia, edited by GORDON RA, BRrrr BA, KALOW W, Springfield, IL, CC Thomas, 1973 10. MASON HS: Mechanism of oxygen metabolism. Adv Enzymol 19: 79, 1957 11. CONNEY AH: Pharmacological implications of microsomal enzyme induction. Pharmacol

Rev 19: 317, 1967 12. TENHUNEN R, MARVER HS, SCHMID R: The

enzymatic conversion of heme to bilirubin by microsomal heme oxygenase. Proc Nail Acad Sci USA 61: 748, 1968 13. GROVER PL, HEWER A, SIMS P: Formation of K-region epoxides as microsomal metabolites of pyrene and benzo(Ajpyrene. Biochem Pharmacol 21: 2713, 1972 14. GROVER PL, SIMS P, HUBERMAN E, et al: In vitro transformation of rodent cells by K-region derivatives of polycyclic hydrocarbons. Proc Nati Acad Sci USA 68: 1098, 1971 15. MARQUARDT H, KuRoKI T, HUBERMAN E, et al: Malignant transformation of cells derived from mouse prostate by epoxides and other derivatives of polycyclic hydrocarbons. Cancer Res 32: 716, 1972 16. NEBERT DW, GELBOIN HV: Substrate-inducible microsomal aryl hydroxylase in mammalian cell culture. J Biol Chem 243: 6250, 1968 17. GIELEN JE, NEBERT DW: Microsomal hydroxylase induction in liver cell culture by phenobarbital, polycyclic hydrocarbons and p,p'-DDT. Science 172: 167, 1971 18. Lu AYH, SOMOGYs A, WEST 5, et al: Pregnenolone-16d carbonitrile: a new type of inducer of drug-metabolising enzymes. Arch Biochem 152: 457, 1972 19. THOMAS PE, KouRI RE, Hurro JJ: The genetics of aryl hydrocarbon hydroxylase induction in mice: a singie gene difference between C57BL/6J and DBA/2J. Biochem Genet 6: 157, 1972 20. NEBERT DW, GIELEN JE: Genetic regulation of aryl hydrocarbon hydroxylase induction in the mouse. Fed Proc 31: 1315, 1972 21. KELLERMANN G, LUYTEN-KELLERMANN M, SHAW CR: Genetic variation of aryl hydrocarbon hydroxylase in human lymphocytes. Am J Hum Genet 25: 327, 1973 22. NETrESHEIM P, HAMMOS AS: Induction of squamous cell carcinoma in the respiratory tract of mice. J Nati Cancer inst 47: 697, 1971 23. SCHREIBER H, NETTESHEIM P, MARTIN DH: Rapid development of bronchiolo-alveolar squamous cell tumors in rats after intratracheal injection of 3-methylcholanthrene.

J Natl Cancer inst 49: 541. 1972 24. TOKUHATA GK: Familial factors in human lung cancer and smoking. Am I Public Health 54: 24, 1964 25. KoURI RE, RATRIE H in, AnAs SA, Ct al: Aryl hydrocarbon hydroxylase induction in human lymphocyte cultures by 2,3,7,8-tetrachloro-dibenzo-p-dioxin. Life Sci lii] 15: 1585, 1974

26. KELLERMANN G, SHAW CR, LUYTEN-KELLERMANN M: Aryl hydrocarbon hydroxylase in-

ducibility and bronchogenic carcinoma. N Engl I Med 289: 934, 1973 27. Aryl hydrocarbon hydroxylase inducibility and lung cancer (E). Lancet 1: 910, 1974 28. LAWTON AR, KINcADE PW, COOPER MD: Sequential expression of germ line genes in

development of immunoglobulin class diversity. Fed Proc 34: 33, 1975

29. WALDMANN TA, BRODER 5, BLAISE RM, Ct al: Role of suppressor T cells in pathogenesis of common variable hypogammaglobulinaemia.

Lancet 1: 609, 1974 30. Suppressor Cells in

immunity, edited by

SINOHAL SK and SINCLAIR NR, London, Ont, U of Western Ontario Pr, 1975

CMA JOURNAL/JULY 12, 1975/VOL. 113

13

31. GIBLETr ER, ANDERSON JE, COHEN F, et al: Adenosine-deaminase deficiency in two patients with severely impaired cellular immunity. Lancet 2: 1067, 1972 32. YOUNT J, NICHOLS P, OCHS D, et al: Absence of erythrocyte adenosine deaxmnase associated with severe combined immunodeficiency. I Ped,atr 84: 173, 1974 33. WOLSERG G, ZIMMERMAN TP, HIEMSTRAK WM, et al: Adenosine inhibition of lymphocyte-mediated cytolysis: possible role of cyclic adenosine monophosphate. Science 187: 957, 1975 34. Biozzi G, STIPPEL C, MOuTON D, et al: Cytodynamics of the immune response in two lines of mice geneticaliy selected for "high" and "low" antibody synthesis. I Exp Med 135: 1071, 1972 35. GASSER DL, SILVERS WK: Genetic determinants of immunological responsiveness, in

Advances in Immunology, vol 18, edited by

DIXON FJ and KUNKEL HG, New York, Acad Pr, 1974, pp 1-66 36. ST ROSE JEM, CINADER B: Genetic control of the tendency to tolerance circumvention.

Eur I Immunol 3: 409, 1973

37. CINADER B, FUJIWARA M: The role of accessorv and thymus-derived cells in resistance to tolerance induction, in Immunological Tol-

erance: Mechanisms and potential therapeutic

applications, edited by KATZ DH and BENACERRAP B, New York, Aced Pr, 1974, pp 67-85 38. FUJIWARA M, CINADER B: Cellular aspects of tolerance. VII. Inheritance of the resistance to tolerance induction. Cell immunol 12: 214, 1974 39. LILLY F: Fv-2: identification and location of a second gene governing the spleen focus response to Friend leukemia virus in mice.

I Natl Cancer Inst 45: 163, 1970

40. ROWE WP, HUMPHREY JB, LILLY F: A major

genetic locus affecting resistance to infection with murine leukemia viruses. III. Assignment of the Fv-1 locus to linkage group VIII of the mouse. I Exp Med 137: 850, 1973 41. AXELRAD AA, WARE M, VANDER GAAG HC:

RNA Viruses and Host Genome in Oncogenesis, edited by EMMELOT P and BENTVEL-

The otitis drop designed to penetrate wax.

bolymycin 0110 The only otic with thonzonium bromide to help the active ingredients get right to the site of infection - even through cerumen and debris. DESTROYS COMMON PATH 0GENS - Provides the broad-spec-

with perforated eardrum or in long standing otitis media because of the possibility

trum bactericidal actions of colistin sulphate and neomycin sulphate.

of ototoxicity caused by neomycin. Contraindications: cOLY-MYcIN oTlc is contraindicated if there is a history of sen-

REDUCES SWELLING, RELIEVES ITCHING -Also contains

the proven

sitivity to any of its components, or in especially herpes simplex, vaccinia and

tubercular,

fungal,

and

benefits of hydrocortisone acetate.

varicella.

Composition: Each ml. contains - colistin base activity 3.0 mg. (as the sulphate); neomycin base activity 3.3 mg. (as the sulphate); hydrocortisone acetate 10.0 mg.; thonzonium bromide 0.5 mg. Dosage: Instil 4 drops bid. Precautions: If sensitivity or irritation occurs, medication should be discontinued promptly. Overgrowth of resistant organisms is possible. Use with care in cases

Supplied: 5 ml. bottles. Full information is avaliable on request.

14 CMA JOURNAL/JULY 12, 1975/VOL. 113

most

viral

lesions,

ZEN P, Amsterdam, North Holland, 1972, pp 239-54 42. ROWE WP, SATO H: Genetic mapping of the Fv-1 locus of the mouse. Science 180: 640, 1973 43. FRIEND C: Cell-free transmission in adult Swiss mice of a disease having the character of a leukemia. I Exp Med 105: 307, 1957 44. STEPHENSON R, AXELRAD AA, MCLEOD DL: Erythroid nature of the response to Friend leukemia virus infection in mice. J Nati Cancer Inst 48: 531, 1972

45. CLARKE BJ, AXELRAD AA: Evidence that the

congeneic mouse strains B6 and B6.S differ at Fv-2, a locus that determines susceptibility of resistance to Friend leukemia virus (in press) 46. LILLY F: H-2 membranes and viral leukemo-

genesis, in Cellular interactions in the immune response: 2nd International Convocation

on lmmunology, Buffalo, NY, June 22-25, 1970, edited by COHEN 5, CUDKOWICZ G, MCCLUSKEY RT, New York, Karger, 1971, pp 103-8

47. CHESEBRO B, WEHRLY K, STIMPPLING I: Host

genetic control of recovery from Friend leukemia virus-induced splenomegaly. Mapping of a gene within the major histocompatability complex. I Exp Med 140: 1457, 1974 48. KLEin J, BACH FH, FESTENSTEIN F, et al: Genetic nomenclature for the H-2 complex

of the mouse. Immunogenetics 1: 184, 1974 49. RACE RR, SANGER R: Blood Groups in Man.

Oxford, Blackwell, 1968 50. MOURANT AE: Associations between hereditary blood factors and diseases. Bull WHO 49: 93, 1973 51. OH JH, MACLEAN LD: Diseases associated with specific HL-A antigens. Can Med Assoc 1 112: 1315, 1975 52. MINTZ B: Gene cohtrol of mammalian pigmentary differentiation. I. Clonal origin of

melanocytes. Proc Natl Acad Sci USA 58:

344, 1967 53. MINTZ B: Hermaphroditism, sex chromosomal mosaicism and germ cell selection in allophenic mice. I Anim Sci 27 (suppl I): 51, 1968 54. Mn.lTz B, PALM 3: Gene control of hematopoiesis. I. Erythrocyte mosaicism and permanent immunological tolerance in allophenic mice. I Exp Med 129: 1013, 1969 55. GRAHAM AF, SCHROEDER JS, GRIEPP RB, et

QuID WARNERICHILCOTT Laboratories Co. Limited Toronto, Canada

al: Does cardiac transplantation significantly prolong life and improve its quality? Circulation 47 (Iuppl III): 116, 1973 56. TAYLOR HE: Pathology of organ transplantation in man. Pathol Annu 7: 173, 1972

57. HAYPLICK L: Current theories of biological aging. Fed Proc 34: 9, 1975 58. GOLDSTEIN 5, NIEwIAROWsKI 5, SINGAL DP: Pathological implications of cell aging in vitro. Ibid, p .