apoptosis in endocrine glands

Review

Apoptosis in Endocrine Glands

George Kontogeorgos, MD and Kalman Kovacs, MD, PHD

Abstract Apoptosis or programmed cell death, is a phenomenon with ultrastructural and biochemical characteristics, which is thought to be distinctive from ordinary necrosis. Shrinkage of cells associated with crescent clumps of heterochromatin and formation of membrane- bound apoptotic bodies are thought to represent distinguishing morphologic features. Internucleosomal cleavage of DNA strands reveals a characteristic ladder pattern in gel electrophoresis. Apoptosis is mediated by an active regulatory mechanism, constitutively expressed in normal and neoplastic cells, bcl-2, bd-x, bax, and APO-1/Fas (CD 95) genes are specifically involved in the apoptotic process. Rat thymocytes exposed to glucocorticoids represent a useful model to study cell death. Steroids and peptide hormones play a role in the regulation of apoptosis. Although there is a great interest in monitoring apoptotic process in endocrine cells and their tumors, only a few studies address apoptosis in endocrine glands so far. One goal of future investigation should be directed to explore therapeutic applications. Key Words: Apoptosis; bcl-2; electron microscopy; histology; necrosis.

Department of Pathology, General Hospital of Athens, Greece, (GK); and Department of Pathology, St. Michael's Hospital, University of Toronto, Ontario, Canada (GK, KK).

Address all correspondence to Dr. George Kontogeorgos, Department of Pathology, General Hospital of Athens, 154 Messogion Avenue, 115 27 Athens, Greece.

Endocrine Patholog~ vol. 6, no. 4, 257-265, November 1995 �9 Copyright 1995 by Humana Press Inc. All rights of any nature whatsoever reserved. 1046-3976/95/6:257-265/$5.80

Introduction Historical Note

Cell growth, differentiation, and death represent universal processes in the life cycle in any living organism. In contrast to death by necrosis that occurs in response to various harmful or injurious conditions and as a rule affects large numbers of adjacent cells, another form of cell death typically develops spontaneously in scattered single or small groups of cells. The studies of Kerr [1] by electron microscopy on the rat liver provided detailed morphologic description of the evolu- tionary shrinkage necrosis. Kerr and his associates used the term "apoptosis" for this specific, hitherto unrecognized form of cell death. The term comes from the Greek word "0~6~:'r03ots" that describes the "falling off" of petals of flowers and leaves of trees [2]. Since then, scientists obtained deep insights into this poorly understood process of cell death.

General Remarks

Apoptosis occurs under several physiologic and pathologic conditions. Apop- tosis represents a specific spontaneous form of programmed cell death, a phenomenon with distinctive morphologic and biochemical characteristics [3,4]. If the injured cells have time, they can probably elect to commit suicide by apoptosis instead of necrosis [5]. This possibility emphasizes the absence of clear-cut borders between the two types of cell death that may overlap. From the morphologic point of view of the cell pathologist, the area of apoptosis vs necrosis, still remains a conflicting field that creates great debates and controversies [6]. However, for the endocrine pathologist, the fact that glucocorticoids, other steroids, and several peptide hormones regulate the active process of apoptosis [7-12], stimu- lates an amazing interest and motivation to explore and understand the importance of this challenging phenomenon.

257

258 Endocrine Pathology Volume 6, Number 4 November 1995

Morphology

The morphologic features of apoptosis and their distinct characteristics from ordinary necrosis have been described in detailed at the light and electron micro- scopic levels [13,14]. The histologic features of apoptosis are manifested by remarkable decrease in cell volume, accom- panied by striking nuclear shrinkage and chromatin condensation (karyopyknosis). It should be noted, however, that even though karyopyknosis is regarded as a histologic sign of cell death, it is practically indistinguishable from cells in metaphase. Therefore, it could not be used as a marker per se to assess apoptosis. By electron microscop)~ apoptotic cells display several discernible features. At the early stages, the cells lose their cytoplasmic junctions and microvilli. Nuclear changes include condensation of chromatin and formation of heterochromatin, with clumping into cres- centic caps attached to the nuclear membrane. At the later stages, fragmentation of cells give rise to the formation of multiple, small membrane-bound apoptotic bodies that represent the hallmark of this event and contain intact cytoplasmic organelles and nuclear fragments [15,16]. Apoptotic cells show compaction of cytoplasmic organelles, vacuolization, loss of cell to cell contact, cytoplasmic blebbing, and at later stages, loss of membrane integrity. The apoptotic bodies that vary in size and number are eventually phago- cytosed by macrophages and less frequently by neighboring cells before losing the integrity of plasma membrane, and they rapidly disappear. Thus, there is no leak- age of cytoplasmic organelles and no inflammatory reaction. In contrast, necrosis is characterized by cytoplasmic and mitochondrial swelling, early rupture and dis- solution of cytoplasmic membrane, and accumulation of cellular content in the

extracellular space. Nuclear changes include chromatin clumping with formation of ill-defined aggregates. Inflamma- tion, which commonly accompanies necrosis, represents an additional feature in differentiating it from apoptosis [4].

Apoptosis represents an active regulatory mechanism for maintaining homeostasis by controlling the optimal numbers of living cells. Although this mechanism for com- mitted cell death seems to be constitutively expressed in normal and neoplastic cells, it is difficult, even impossible, to identify apoptosis by morphology alone. One of the reasons is that apoptosis is a rapid process; it is usually an asynchronous transient event of cell death, lasting only for a short period of time. Therefore, the cell morphologist, who has to rely on a given instance of this multistep spectrum, is unable to rec- ognize conclusively scattered, often scarce cells, undergoing apoptosis. Glucocorticoid- exposed rat thymocytes represent an excel- lent and weU-established experimental model for studying apoptosis [7,9].

Molecular and Biochemical Basis

Degradation of DNA leading to cleavage into nucleosome-sized fragments of approx 200 bp and multiples (DNA lad- ders), represents the biochemical marker of apoptosis [3]. These changes are caused by breaks in DNA strand by activation of endogenous Ca2+-dependent endonuclease [7,9]. Agarose gel electrophoresis of the fragmented DNA, extracted from apoptotic cells, reveals a characteristic DNA ladder pattern, as initially analyzed and described in glucocorticoid-exposed rat thymocytes [7]. It has been shown that the chromatin is cleaved at the linker DNA sites between nucleosomes, producing oligonucleosomal fragments [7,13]. It has been suggested that in apoptosis, selective

Apoptosis in Endocrine Glands 259

activation of endonuclease activity results in specific chromatin cleavage, which cor- responds to the major nuclear alterations seen by morphology [9]. The internucleosomal DNA cleavage of the apoptotic process has been further studied in several normal and neoplastic tissues, mostly from animals including endocrine glands and hormone target tissues, such as thyroid gland [17], testis [10,18], ovary [19], endometrium [20], and prostate [8].

The internucleosomal cleavage precedes the appearance of morphologic signs of apoptosis and lasts to the end of this event, terminating in cell death. It should be emphasized that this type of DNA fragmentation correlates with cell death and strongly indicates the specificity of this pattern in the apoptotic process [21]. In contrast, when necrosis is induced by injury, DNA fragmentation either does not occur or the DNA is degraded in a non- specific fashion, leading to the formation of a continuous spectrum of various length fragments [22].

Regulatory Genes

bcl-2 is a protein derived from the inner mitochondrial membrane originally dis- covered by its involvement in chromosomal translocations occurring in follicular lymphomas [23]. It is related to programmed cell death and is restricted to cells with long life spans, such as stem cells, neurons, and so on. Overexpression of bcl-2 gene blocks the apoptotic process, thus bcl-2 oncogene may play a significant role in prolongation of cell survival with- out activating mitogenic mechanisms [24,25]. Extracellular signals can promote cell survival by stimulating the expression of bcl-2, whereas others have an opposite effect. In addition to bcl-2, the bcl-x, bax, and APO-1/Fas (CD 95) genes are also

implicated in regulation of apoptosis, bcl-x has a similar action to bcl-2, whereas bax, exerts an opposite effect [26]. APO-1/Fas ligand is a type 1 transmembrane receptor that belongs to the tumor necrosis factor and epidermal growth factor (EGF) family. APO-1/Fas transduces signals within various cells and induces apoptosis [27]. Adenovirus transforming protein (El B) has a similar action to bcl-2 [28] in pro- longating the life span of cells, whereas the transforming gene of Ableson virus (v-abl) leads to immortalization of relevant target cells [29]. In contrast overexpression of p53 tumor suppressor gene can arrest the cell cycle, resulting in induction of apoptosis [30]. bcl-2 and other genes which partici- pate in the regulation ofapoptotic control may play key roles in novel therapeutic approaches acting through the same cell suicide pathways.

Techniques for Demonstration

In addition to morphology and analysis of DNA fragmentation by gel electrophoresis, novel methods have been introduced for detection of apoptotic cells. Immuno- cytochemistry for localization of bd-2 and other antigens related to apoptosis can be performed applying the usual staining pro- cedures [31].

In situ labeling technique can be used according to various protocols. The prin- ciple of this technique is to demonstrate nuclear DNA fragmentation by labeling DNA containing free 3'-OH ends (3'-OH nick end labeling). The enzyme polymerase is used, first to generate 5' overhangs from any 3'-OH ends of DNA strand beaks and then to end-fill the overhangs and incorporate biotinylated deoxyadenosine or deoxyuridine triphosphate. These steps are followed by visualization of the labeled sites by immunoperoxidase detection systems


[32]. This technique is ideal for the demonstration of DNA strands containing breaks, and it is specific only for DNA and not for RNA. Modified protocols utilize residues of digoxigenin-nucleotide to incorporate into the 3 ' -OH ends of fragmented DNA with the aid of terminal deoxynucleotidyl transferase (TdT) [12,33]. Visualization can be done with appropriate systems either for light or fluorescence microscopy [34,35]. Slight modifications permit the application of this technique in fresh cells or in formalin-fixed, paraffin- embedded tissues. Flow cytometric or automated image analysis systems can facilitate the counting of the labeled sites [35,36]. However, it should be stressed that the morphologist must be cautious in the interpretation of the in situ labeling findings. Serial sections should be examined to exclude adjacent necrosis, which may give positive signals. In addition, DNA breaks occurring either during tissue processing or when fixation is delayed may be respon- sible for false-positive results [37], whereas cells containing typical apoptotic bodies may not be labeled [38].

Endocrine Glands

Several studies have confirmed the existence ofapoptosis in endocrine glands, primarily in animal tissues in vivo and also in tissue cultures. Most of the studies were performed on testes, ovaries, and adrenals, whereas information on other endocrine glands is limited.

Testis

Germ cell death occurs spontaneously in the course of the development of germ cell epithelium and during spermatogen- esis [39]. Recent studies have shown that deprivation ofgonadotropins from immature rat testes results in apoptotic DNA

fragmentation [10]. The same effect was reproduced after treatment with a long- acting gonadotropin antagonist, and in situ analysis showed major apoptotic changes in spermatocytes of adult rat testis. In contrast to spermatocytes, Sertoli and Leydig cells were not affected [18]. These results suggest that, in response to lack of hormonal stimulation germ, cells die by apoptosis [ 10,12].

Surgical induction of cryptorchism in the rat causes disruption of spermatogen- esis and infertility. Recent studies revealed apoptotic DNA fragmentation and it was demonstrated by in situ DNA labeling analysis that germ cells, primarily spermatocytes, were affected. In addition, in the rat testes of unilaterally induced cryptorchism by surgery, the percentages of the seminiferous tubules containing labeled cells with DNA breaks were increased, compared to the contralateral intact testis in the same animal [40].

Ovary

It is known that >99% of ovarian follicles, including oocytes, undergo atresia, which is mediated via apoptotic mechanisms. This type ofapoptotic evolution was demonstrated by in situ DNA labeling in immature, estrogen-treated, and adult cycling rats. Granulosa theca cells of preantral and antral atretic follicles and scattered theca cells were positive in all three models. Healthy antral and preantral follicles and stromal cells were consistently negative [41]. The manipulat ion of apoptosis with various hormones and other factors in follicular atresia represents a field of great interest. The importance of sex steroids in regulating follicle cell apoptosis was investigated in immature hypophy- sectomized rats. It was shown that estrogens prevent apoptosis and androgens antagonize the effect of estrogens [19].


The substantial decline in Fas mRNA expression, noted when pregnant mare's gonadotropins are injected into murine atretic follicles, indicates an apoptotic regulation of oocyte death in atretic follicles [11]. In another study using immortalized ovarian cell lines, normal and malignant ovarian epithelia, the inhibitory effects of transforming growth factor beta (TGF-[3) on cell proliferation, were investigated. Although cell proliferation was inhibited in all instances, no evidence of apoptosis in normal epithelia was seen, but it was noted in 3 of 10 cancers [42]. These results suggest that malignant cells are more susceptible to apoptosis than normal cells.

Adrenal Cortex

The human fetal adrenal cortex under- goes dramatic postnatal remodeling. Apoptosis in the adrenals of normal neo- natal rats is prominent during temporary physiologic adrenocorticotrophic hormone (ACTH) deprivation, whereas ACTH administration can prevent this effect [43]. By in situ labeling of cleaved DNA, it was recently found that cells of the human fetal zone die by apoptosis, with the peak incidence occurring I wk after birth when the involution of the fetal zone is confirmed by histology. It was also demonstrated that activin-A and TGF-J3, promote apoptotic DNA cleavage in cells of human fetal zone in vitro [44]. In addition, it was found that ACTH attenuates internucleosomal DNA cleavage in the intact rat adrenals in vitro. It was suggested that ACTH is probably the sole pituitary hormone that regulates the death of adrenocortical cells in vivo [45].

Apoptosis was demonstrated in the rat adrenals by histology. The apoptotic bodies were apparent in the inner parts of the cortex. In situ localization of DNA strand breaks revealed that apoptosis occurred nearly exclusively in zona reticularis cells

abutting the medulla [45]. Recent results obtained in normal human adrenals by in situ end labeling disclosed apoptotic cells in the zona faciculata and infrequently in the inner zona reticularis. Apoptotic tumor cells were also noted in approx 43% of cortical adenomas and carcinomas. How- ever, no differences were observed among normal, adenoma, and carcinoma cells. These findings show that apoptosis plays a significant role in maintenance of orga- nized cell turnover in normal cortex and in tumor cells [33].

Endocrine Pancreas

Development of pancreatic J3-cell tumors can be induced by an insulin-regulating transgene in multiple lines of transgenic mice. It was shown that in insulin-like growth factor (IGF)-II gene knockout mice, the tumor cells appeared to be more benign with smaller nuclei, larger cytoplasm, and a fivefold higher incidence ofapoptotic bodies than in those of appropriate controls. These data signify the role of IGF-II as modulator in the apoptotic process during multistep tumorigenesis [46]. Another study showed that pancreatic cells from RINm5F cell line treated with interleukin-l~ underwent typical DNA fragmentation and morphologic changes with formation ofapoptotic bodies. Inhibition of nitric oxide synthase activity prevented apoptosis [47].

Thyroid In a dog's thyroid, cell deprivation of

thyrotropin and EGF activate apoptotic cell death. Cells collected from the super- natant of tissue-culture medium demonstrated characteristic morphologic changes of apoptosis and internucleosomal DNA fragmentation. These results are in keep- ing with the in vivo thyroid regression in the absence of growth stimulation of the


thyroid gland [17]. Apoptosis was also studied in human medullary carcinoma cell line (TT). Exposure of cells to TGF-I31, in the presence of increased levels of c-myc expression, resulted in inhibition of cell proliferation and acceleration ofapoptosis. Apoptotic cells displayed typical ultrastructural changes and DNA laddering pattern [48]. In another study, immunocytochemi- cal analysis for bcl-2 was carried out in tumors arising in the follicular epithelium of human thyroid gland. The results showed that nearly 80% of well-differentiated and poorly differentiated carcinomas were positive for bcl-2, whereas only 13.5% of undifferentiated carcinomas exhibited immunoreactivity. Mutual exclusion of bcl-2 and p53 expression was noted in the undifferentiated and differentiated compo- nents. In contrast, fetal and adult normal thyroid glands were immunonegative for bcl-2 [31].

Pituitary

The effects of estrogens and bromocriptine on apoptosis have been recently studied in rat anterior pituitary gland. Increased cell counts of apoptosis were noted in rats with estrogen-induced pro- lactin cell hyperplasia after estrogen withdrawal in vivo. Subsequent bromocriptine administration resulted in an approxi- mately twofold greater increase of apoptotic cell counts compared with the nonbromocriptine-treated animals. These results suggest that dopamine agonists induce apoptosis and affect the phagocytic properties of stellate cells [49]. Similarly, bromocriptine induces apoptosis in growth hormone (GH)-producing rat adenoma (GH1) cells and murine ACTH-secreting adenoma (AtT-20) cells [50,51].

Induction of apoptosis by long-acting somatostatin analogs was demonstrated in AtT-20 mouse pituitary tumor cells. It was

suggested that somatostatin-induced increase in c-myc, associated with inactiva- tion of growth factor receptor kinases, may play a role in promoting apoptosis [52].

Perspectives Although substantial progress has been

made since the initial concept ofapoptosis was introduced, it is obvious that a long journey is required to understand this com- plicated phenomenon. Manipulation of cell life span by regulating apoptosis gen- erates amazing interest and perspectives for future pharmaceutical applications, including endocrine therapy. Induced cell death in mammary carcinoma cells by administration ofgonadotropin-releasing hormone analogs and somatostatin or estrogen withdrawal in cell lines in vitro [53,54], represents only a minor example of promising applications for anticancer therapy in the near future.

Acknowledgments This work was supported in part by

Grants E-391 of the National Health Council of Greece (George Kontogeorgos) and MA-6349 of the Medical Research Council of Canada (Kalman Kovacs).

References 1. Kerr JE Shrinkage necrosis: a distinct mode

of cellular death. J Pathol 105:13-20, 1971. 2. Kerr JFR, Wyllie AH, Currie AR. Apoptosis:

a basic biological phenomenon with wide- ranging implications in tissue kinetics. Br J Cancer 26:239-257, 1972.

3. Appleby DW, Modak SE DNA degradation in terminally differentiating lens fiber from chick embryos. Proc Natl Acad Sci USA 74:5579-5583, 1977.

4. Trump BF, Berezesky IK, Cowley RA. The cellular and subsellular characteristics of


acute and chronic injury with emphasis on the role of calcium. In: Cowley RA, Trump BF, eds. Pathology of shock, anoxia and ischemia. Baltimore, MD: Williams and Wilkins, 1982; 6-64.

5. Wyllie AH, Kerr JFR, Currie AR. Cell death: the significance of apoptosis. Int Rev Cytol 68:251-306, 1980.

6. Farber E. Programmed cell death: necrosis ver- sus apoptosis. Mod Pathol 7:605-609, 1995.

7. Wyllie AH. Glucocorticoid-induced thy- mocyte apoptosis in associated with endogenous endonuclease activation. Nature 284:555,556, 1980.

8. Kyprianou N, Isaacs JT. Activation of programmed cell death in the rat ventral prostate after castration. Endocrinology 122:552- 562, 1988.

9. Arends MJ, Morris RG, Wyllie AH. Apop- tosis. The role of the endonuclease. Am J Pathol 136:593-608, 1990.

10. Tapanainen JS, Tilly JL, Vihko KK, Hsueh AJ. Hormonal control of apoptotic cell death in the testis: gonadotropins and androgens as testicular cell survival factors. Mol Endocrinol 7:643-650, 1993.

11. Guo MW, Mori E, Xu JP, Mori T. Identifica- tion of Fas antigen associated with apoptotic cell death in murine ovary. Biochem Biophys Commun 23:1438-1446, 1994.

12. Hikim APS, Wang C, Leung A, Swerdloff RS. Involvement ofapoptosis in the induction of germ cell degeneration in adult rats after gonadotropin-releasing hormone antagonist treatment. Endocrinology 136: 2770-2775, 1995.

13. Wyllie AH. Cell death: a new classification separating apoptosis from necrosis. In: Bowen ID, Lockshin RA, eds. Cell death in biology and pathology. London: Chapman and Hall, 1981; 9-34.

14. Walker NI, Harmon BV, Gob~ GC, Kerr JFR. Patterns of cell death. Methods Achiev Exp Pathol 13:18-54, 1988.

15. Wyllie AH. Apoptosis ISI atlas science. Immunology 1:192-196, 1988.

16. Brusch W, Klein L, Tenniswood M. The bio- chemistry of cell death by apoptosis. Biochem Cell Biol 88:1071-1074, 1990.

17. Dremier S, Golstein J, Mosselmans R, Dumont JE, Galand P, Robaye B. Apoptosis in dog thyroid cells. Biochem Biophys Res Commun 200:52-58, 1994.

18. Billing H, Furuta I, River C, Tapanainen J, Parvinen M, Hsueh AJ. Apoptosis in testis germ cells: developmental changes in gonadotropin dependence and localization to selective tubule stages. Endocrinology 136:5- 12, 1995.

19. Billing H, Furuta I, Hsueh AJ. Estrogens inhibit and androgens enhance ovarian granulosa cell apoptosis. Endocrinology 133:2204- 2214, 1993.

20. Rottelo RJ, Hocker MB, Gerschenson LE. Biochemical evidence for programmed cell death in rabbit uterine epithelium. Am J Pathol 134:491-495, 1989.

21. Compton MM, Cidlowski JA. Rapid in vivo effects of glucocorticoids on the integrity of rat lymphocyte genomic deoxyribonucleic acid. Endocrinology 118:38-45, 1986.

22. Duvall E, Wyllie AH, Currie AR. Death and the cell. Immunol Today 7:115-119, 1986.

23. Tsujimoto Y, Cossman J, Jaffe E, Croe C. Involvement of the bcl-2 gene in human follicular lymphoma. Science 228:1440- 1443, 1985.

24. Hockenbery D, Nufiez G, Milliman C, Schreiber RD, Kosmeyer SJ. Bcl-2 is an inner mitochondrial membrane protein that blocks programmed cell death. Nature 384:334-336, 1990.

25. Hockenbery DM, Zutter M, Hickey W, Nahm M, Kosmeyer SJ. BCL2 protein in topographically restricted in tissues characterized by apoptotic cell death. Proc Natl Acad Sci USA 88:6961-6965, 1991.

26. Grell M, Krammer PH, Scheurich P. Segrega- tion of APO-1/Fas antigen- and tumor necrosis factor receptor-mediated apoptosis. Eur J Immuno124:2563-2566, 1994.

27. Bargou RC, Daniel PT, Mapa MY, Bommet K, Wagner C, Kallinich B, Royer HD, Doken B. Expression of bcl-2 gene family in normal and malignant breast tissue: low bax-alpha expression in tumor cells correlates with resistance towards apoptosis. Int J Cancer 60:854-859, 1995.

28. White E, Sabbatini P, Debbas M, Wold WS, Kusher DI, Gooding LR. The 19-kilodalton adenovirus E1B transforming protein inhib- its programmed cell death and prevents cytolysis by tumor necrosis factor alpha. Mol Cell Biol 12:2570-2780, 1992.

29. Evans CA, Owen-Lynch JP, Whetton AD, Dive C. Activation of the ableson tyrosine


kinase activity is associated with suppression ofapoptosis in hemopoietic cells. Cancer Res 53:1735-1738, 1993.

30. Lane DP. Cancer. p53, guardian of the genome. Nature 358:15,16, 1992.

31. Pilotti S, Collini P, Rilke F, Cattoretti G, Del Bo R, Pierotti MA. Bcl-2 protein expression in carcinomas originating from the follicular epithelium of the thyroid gland. J Pathol 172:337-342, 1994.

32. GavrieliY, SchermanY, Ban-Seasson SA. Iden- tification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol 119:493-501, 1992.

33. Sasano H, Imatani A, Souichirou S, Suzuki T, Naguara H. Cell proliferation and apoptosis in normal and pathologic human adrenals. Mod Pathol 8:11-17, 1995.

34. Schmitz GG, Walter T, Seibl R, Kessler C. Nonradioactive labeling of oligonucleotides in vitro with hapten digoxigenin by tailing with terminal transferase. Anal Biochem 192:222- 231, 1991.

35. Wijsman JH, Jonker RR, Keijzer R, van de Velde CJH, Cornelisse CJ, van Dierendonck JH. A new method to detect apoptosis in par- affln sections: in situ end-labeling of fragmented DNA. J Histochem Cytochem 41:7-12, 1993.

36. Darzynkiewicz Z, Bruno S, Del Bino G, Gorczyca W, Hotz MA, Lassota P, Traganos F. Features of apoptotic cells measured by flow cytometry. Cytometry 13:798-808, 1992.

37. Sasano H. In situ end labeling and its applications to the study of endocrine disease: how can we study programmed cell death in surgical pathology materials? Endocr Pathol 6:87- 89, 1995.

38. Dowsett M, Johnston SRD, Newby J, Golding M, Sacks N, Smith IE. Mechanisms of hormone response: a role for apoptosis. Endocr Relat Cancer 2:3-11, 1995.

39. Wing T-Y, Christensen AK. Morphometric studies on rat seminiferous tubule. Am J Anat 165:13-25, 1982.

40. Shikone T, Billing H, Hsueh AJ. Experimen- tally induced cryptorchidism increases apoptosis in rat testis. Biol Reprod 51:865- 872, 1994.

41. Palumbo A, Yeh J. In situ localization of apoptosis in ovary during follicular atresia. Biol Repr 51:888-895, 1995.

42. Havrilesky LJ, Hureau JA, Whitaker RS, Elbendary A, Wu S, Rodriguez GC, Bast RC Jr, Berchuck A. Regulation of apoptosis in normal and malignant ovarian epithelial cells by transforming growth factor beta. Cancer Res 55:944-948, 1995.

43. Wyllie AH, Kerr JFR, Macaskill IAM, Curie AR. Adrenocortical cell deletion: the role of ACTH. J Pathol 111:85-94, 1973.

44. Spencer SJ, Mesiano S, Jaffe RB. Programmed cell death in the human fetal adrenal cortex: actions of activin, follistatin and TGF-beta, Abstracts 77th Annual Meeting of Endocrine Society, Washington, DC, 1995; 611.

45. Carsia RV, Macdonald GJ, Tilly KI, Tilly JL. Terminally differentiated adrenocortical cells are most susceptible to apoptosis induced by adrenocorticotropic hormone (ACTH) withdrawal, Abstracts 77th Annual Meet- ing of Endocrine Society, Washington, DC, 1995; 615.

46. Naik SP, Christofori G, Hanahan D. Both developmentally-regulated and maternally- imprinted alleles of IGF-II are focally acti- vated to provide a survival factor that attenuates apoptosis during multistep tumorigenesis, Abstracts 77th Annual Meet- ing of Endocrine Society, Washington, DC, 1995; 30.

47. Ankarcrona M, Dypbukt JM, Brune B, Nicotera P. Interleukin-1 beta-induced nitric oxide production activates apoptosis in pancreatic RINm5F cells. Exper Cell Res 213:172-177, 1994.

48. Khosla S, Ousler MJ, Schroeder MJ, Eber- hardt N. Transforming factor-beta 1 induces growth inhibition of human medullary carcinoma cell line despite an increase in steady state c-myc messenger ribonucleic acid levels. Endocrinology 135:1887-1893, 1994.

49. Drewett N, Jacobi JM, W'dlgoss DA, Lloyd HM. Apoptosis in the anterior pituitary gland of the rat: studies with estrogen and bromocriptine. Neuroendocrinology 57:89-95, 1993.

50. Yin D, Kondo S, Takeuchi J, Morimura T. Induction of apoptosis in rat somatotropin- secreting adenoma cells by bromocriptine. Oncol Res 5:383-387, 1993.

51. Yin D, Kondo S, Takeuchi J, Morimura T. Induction of apoptosis in rat ACTH-secreting adenoma cells by bromocriptine. FEBS Lett 339:73-75, 1994.


52. Sharma K, Srikant CB. Somatostatin analogs induce c-myc gene expression in AtT-20 mouse pituitary tumor cells, Abstracts 77th Annual Meeting of Endocrine Society, Wash- ington, DC, 1995; 276.

53. Szende B, Lapis K, Redding T~, Srkalovic G, Schally AV. Growth inhibition of MXT mammary carcinoma by enhancing pro-

grammed cell death (apoptosis) with analogs of LH-RH and somatostatin. Breast Cancer Res Treat 14:307-314, 1989.

54. Kyprianou N, English HF, Isaacs JT. Pro- grammed cell death regression of the MCF-7 human breast cancer following estrogen ablation. Cancer Res 51:162- 166, 1991.

apoptosis in endocrine glands

Documents