Ir» J Gynaecol Obstet 17: 219-225, 1979
An Ultrastructural and Clinical Study of Theca and Granulosa Cell Tumors P. J. Klemi 1 and M. Grònroos 2 Departments of Pathological Anatomy1 and Obstetrics and Gynaecology, University Central Hospital, Turku, Finland
ABSTRACT
MATERIALS A N D M E T H O D S
Klemi PJ, Gronroos M (Depts of Pathological Anatomy and Obstetrics and Gynaecology, University Central Hospital, Turku, Finland). An ultrastructural and clinical study of theca and granulosa cell tumors. IntJ Gynaecol Obstet 17: 219-225, 1979 To establish the cell type responsible for the production of estrogens in an ovarian tumor, seven thecomas, three granulosa cell tumors and one fibroma were studied with electron microscopy. The secretion of estrogens by the tumor was determined by measuring the plasma levels of estrogens in the ovarian and cubital veins of some patients and examining the histology of the endometria. Polygonal cells with abundant smooth endoplasmic reticulum (SER) at the ultrastructural level were found in the tumors of patients with clinical signs of hyperestrogenism. It is likely that these cells are the site of steroidogenesis, which is known to be reflected by SER in the cells of the human corpus luteum.
T h e tumors were classified and staged according to the system adopted by the World Health Organization (16) and the Federation Internationale de Ginécologie et Obstétrique (12). T h e criteria for increased estrogen production were a proliferative endometrium with or without vaginal bleeding at postmenopausal age, elevation of the plasma estrogens in the cubital a n d / o r ovarian vein and an anovulatory menstrual cycle. Serum estrone, estradiol-17/8 and estriol were determined by radioimmunoassay. After the samples were extracted with ether, Sephadex LH-20 chromatography (Pharmacia Fine Chemicals, Uppsala, Sweden), as described by Carr et al (2), was used to separate estrone, estradiol and estriol. T h e radioimmunoassay was performed using a modification of the method of Hotchkiss et al (7). Tissue samples were fixed in neutral formalin and embedded in paraffin and the sections were stained according to van Gieson and hematoxylin-eosin methods for a general characterization of the tumor. Additional methods included the Gõmòry for reticulin fibers, the Verhoef for elastic fibers, Sudan black for fat and periodic acid Schiff, with a n d without diastase digestion, for glycogen (11). Small pieces (1 mm 3 ) of the tumors taken during or right after surgery were processed for study with a J E O L J E M 100C electron microscope. The cells found in the tumors were classified according to Amin et al (1) as spindle (S), intermediate (I), polygonal (P) a n d granulosa (G).
INTRODUCTION Theca and granulosa cell tumors of the ovary are fairly rare, representing about 4% of all ovarian tumors (8). Both tumors may periodically be endocrinologically active. Most often patients with these conditions complain of menstrual irregularity or bleeding which is caused by increased estrogen production beyond the reproductive age. O n the other hand, it is fairly difficult to determine on morphologic grounds alone whether the tumor is capable of producing hormones. There have been studies dealing with the ultrastructure of theca cell (1, 17, 18) and of granulosa cell (5, 13, 15, 17) tumors, but not all of them have found the tumor to contain cells with smooth endoplasmic reticulum (SER), which would reflect steroidogenesis. T o establish ultrastructural correlation with the clinically suspected or proven increase in estrogen activity, seven theca, three granulosa cell tumors and one fibroma were investigated with light and electron microscopy.
RESULTS Clinical findings All the patients with theca cell tumors and the one with a fibroma were of postmenopausal age. T h e women with granulosa cell tumors were of reproductive or premenopausal age. Four of the eight who were postmenopausal h a d proliferative
IntJ Gynaecol Obstet 17
220
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endometrium, two of them with vaginal bleeding. O n e of the three patients of reproductive or premenopausal age had dysfunctional uterine bleeding. All tumors were of clinical stage I and could be removed completely. At follow-up periods of one to four years, the patients were asymptomatic. T h e estrogen values of the blood samples taken from the ovarian vein during surgery were estrone, 0.45 nmol/liter; estradiol, 0.13 nmol/liter; and estriol, 0.02 nmol/liter. T h e plasma levels in the cubital vein of the same patient were estrone, 0.23 nmol/liter; estradiol, 0.01 nmol/liter; and estriol, 0.02 nmol/liter. Estradiol values in the cubital vein of two patients were 0.45 nmol/liter and 0.17 nmol/liter. None of the women had been treated with exogenous estrogens. Some clinical and histologic details of the material are summarized in Table I. Histologic and electron microscopic findings T h e distribution of cells in the tumors are presented in Table I. Long or polymorphic S cells were found in every tumor, and the fibroma was composed mainly of these cells (Fig. 1). T h e cell membrane was slightly irregular with some pinocytotic vesicles. T h e sparse cytoplasm contained some elon-
gated mitochondria and ribosomes, few microfibrils and a poorly developed Golgi complex. T h e nucleus was sometimes irregular in shape with chromatin condensed toward the periphery. There were various amounts of collagen fibers and fewer reticulin fibers in the intercellular space. I cells were found in all theca cell tumors a n d in the granulosa cell tumor of a patient with suspected increase in estrogen production. T h e cells were roundish or more irregular in shape than the S cells a n d the nuclear-cytoplasmic ratio varied from cell to cell (Fig. 2). T h e cell membrane was smooth, and there were few desmosomes between the cells. There were more organelles in the cytoplasm of I cells than in that of S cells. Some lipid droplets were found in the cytoplasm but not more than in the cytoplasm of S cells. T h e nuclei were more irregular than the nuclei in S cells. O n e or more nucleolus could be found. Some cells contained microfibrils that were 50 A in diameter and sometimes arranged in whorls (Fig. 3). This material did not stain with the methods used in this study. P cells were found only in the tumors of the patients with signs of elevated estrogen production (Table I). T h e cells were in small nodules or scattered in the tumors and contained a b u n d a n t SER (Figs. 4 and 5). Otherwise they resembled the I cells
Table I. Clinical and histologic findings of 11 cases. Tumor Cell Types8 Age of Patients (years) 51 59 68 61 64 70 70 60
27 32 51
Signs of Increased Estrogen Production None None None Vaginal bleeding, proliferative endometrium, increased level of plasma estradiol Increased plasma estradiol Proliferative endometrium Proliferative endometrium Vaginal bleeding, endometrial carcinoma, proliferative endometrium, increased plasma levels of estrogens None Irregular vaginal bleeding, anovulatory cycles, proliferative endometrium None
Spindle
Intermediate
Thecoma Fibroma Thecoma Thecoma
+ +++ ++ +
++ + + +++
Thecoma Thecoma Thecoma Thecoma
+ + +++ +
+++ +++ + ++
++
+
—
—
+ ++
+
+
+
+++
—
+++
Histologic Type of Tumor
Granulosa cell tumor, microfollicular Granulosa cell tumor, diffuse Granulosa cell tumor, microfollicular
+
Symbols for quantifying cell composition: - none; + some; + + moderate number; +++
IntJ Gynaecol Obstet 17
Polygonal — — —
Granulosa — — —
+ — — —
+ +
—
preponderance.
Theca and granulosa cell tumors
221
^¿JiA'B* Fig. 1. An electron micrograph of an ovarian fibroma. The nuclei (N) of four spindle cells fill the cytoplasm almost completely. Transverse sections of collagen fibers (K) are seen in the intercellular stroma (X7000).
in their ultrastructure. Lipid droplets were numerous in some cells. Numerous mitochondria were elongated with some tubular christae. The G cells were round or somewhat elongated. They also formed Call-Exner bodies whose inner part was composed of lamellated and degenerated cell organelles (Fig. 6). There were many desmosomes between the G cells. In the cytoplasm, there were many round or elongated mitochondria, ribosomes, fibrils of various length, sometimes a welldeveloped Golgi complex and occasionally secondary lysosomes. T h e stroma of the granulosa cell
tumor was composed of collagen bundles and some reticulin fibers within a loose intercellular space.
DISCUSSION Most of our results supported the findings of others concerning the ultrastructure of theca cell tumors (1, 17, 18) and of granulosa cell tumors (13, 15, 17). In addition, we found polygonal cells with abundant SER in the tumors of patients who showed clinical signs of suspected or proven increase
IntJ Gynaecol Obstet 17
222
Klemi and Gronroos
Fig. 2. An electron micrograph of a thecoma with intermediate cells. The intercellular stroma is scant. There are numerous mitochondria (M) in the cytoplasm of the cells (X6600).
Fig. 3. A high magnification electron micrograph of an intermediate cell with abundant microfibrils (m) in the cytoplasm below the nucleus (X27 700).
in estrogen production. Since SER in the cells reflects steroidogenesis, as shown in the human corpus luteum by Crisp et al (6) and Mestwerdt et al (14), we may assume that the P cells in the theca and granulosa cell tumors are the site of steroidogenesis.
IntJ Gynaecol Obstet 17
In the theca cell tumors, there is a gradual change from a spindle cell, via an intermediate cell to a polygonal cell, indicating the stromal origin of polygonal cells. T h e question arises as to the origin of the polyg-
Theca and granulosa cell tumors
223
Fig. 4. An electron micrograph of a polygonal cell with abundant smooth endoplasmic reticulum and some lipid droplets ( x 4700).
V
An Ultrastructural and Clinical Study of Theca and Granulosa Cell Tumors P. J. Klemi 1 and M. Grònroos 2 Departments of Pathological Anatomy1 and Obstetrics and Gynaecology, University Central Hospital, Turku, Finland
ABSTRACT
MATERIALS A N D M E T H O D S
Klemi PJ, Gronroos M (Depts of Pathological Anatomy and Obstetrics and Gynaecology, University Central Hospital, Turku, Finland). An ultrastructural and clinical study of theca and granulosa cell tumors. IntJ Gynaecol Obstet 17: 219-225, 1979 To establish the cell type responsible for the production of estrogens in an ovarian tumor, seven thecomas, three granulosa cell tumors and one fibroma were studied with electron microscopy. The secretion of estrogens by the tumor was determined by measuring the plasma levels of estrogens in the ovarian and cubital veins of some patients and examining the histology of the endometria. Polygonal cells with abundant smooth endoplasmic reticulum (SER) at the ultrastructural level were found in the tumors of patients with clinical signs of hyperestrogenism. It is likely that these cells are the site of steroidogenesis, which is known to be reflected by SER in the cells of the human corpus luteum.
T h e tumors were classified and staged according to the system adopted by the World Health Organization (16) and the Federation Internationale de Ginécologie et Obstétrique (12). T h e criteria for increased estrogen production were a proliferative endometrium with or without vaginal bleeding at postmenopausal age, elevation of the plasma estrogens in the cubital a n d / o r ovarian vein and an anovulatory menstrual cycle. Serum estrone, estradiol-17/8 and estriol were determined by radioimmunoassay. After the samples were extracted with ether, Sephadex LH-20 chromatography (Pharmacia Fine Chemicals, Uppsala, Sweden), as described by Carr et al (2), was used to separate estrone, estradiol and estriol. T h e radioimmunoassay was performed using a modification of the method of Hotchkiss et al (7). Tissue samples were fixed in neutral formalin and embedded in paraffin and the sections were stained according to van Gieson and hematoxylin-eosin methods for a general characterization of the tumor. Additional methods included the Gõmòry for reticulin fibers, the Verhoef for elastic fibers, Sudan black for fat and periodic acid Schiff, with a n d without diastase digestion, for glycogen (11). Small pieces (1 mm 3 ) of the tumors taken during or right after surgery were processed for study with a J E O L J E M 100C electron microscope. The cells found in the tumors were classified according to Amin et al (1) as spindle (S), intermediate (I), polygonal (P) a n d granulosa (G).
INTRODUCTION Theca and granulosa cell tumors of the ovary are fairly rare, representing about 4% of all ovarian tumors (8). Both tumors may periodically be endocrinologically active. Most often patients with these conditions complain of menstrual irregularity or bleeding which is caused by increased estrogen production beyond the reproductive age. O n the other hand, it is fairly difficult to determine on morphologic grounds alone whether the tumor is capable of producing hormones. There have been studies dealing with the ultrastructure of theca cell (1, 17, 18) and of granulosa cell (5, 13, 15, 17) tumors, but not all of them have found the tumor to contain cells with smooth endoplasmic reticulum (SER), which would reflect steroidogenesis. T o establish ultrastructural correlation with the clinically suspected or proven increase in estrogen activity, seven theca, three granulosa cell tumors and one fibroma were investigated with light and electron microscopy.
RESULTS Clinical findings All the patients with theca cell tumors and the one with a fibroma were of postmenopausal age. T h e women with granulosa cell tumors were of reproductive or premenopausal age. Four of the eight who were postmenopausal h a d proliferative
IntJ Gynaecol Obstet 17
220
Klemi and Cromóos
endometrium, two of them with vaginal bleeding. O n e of the three patients of reproductive or premenopausal age had dysfunctional uterine bleeding. All tumors were of clinical stage I and could be removed completely. At follow-up periods of one to four years, the patients were asymptomatic. T h e estrogen values of the blood samples taken from the ovarian vein during surgery were estrone, 0.45 nmol/liter; estradiol, 0.13 nmol/liter; and estriol, 0.02 nmol/liter. T h e plasma levels in the cubital vein of the same patient were estrone, 0.23 nmol/liter; estradiol, 0.01 nmol/liter; and estriol, 0.02 nmol/liter. Estradiol values in the cubital vein of two patients were 0.45 nmol/liter and 0.17 nmol/liter. None of the women had been treated with exogenous estrogens. Some clinical and histologic details of the material are summarized in Table I. Histologic and electron microscopic findings T h e distribution of cells in the tumors are presented in Table I. Long or polymorphic S cells were found in every tumor, and the fibroma was composed mainly of these cells (Fig. 1). T h e cell membrane was slightly irregular with some pinocytotic vesicles. T h e sparse cytoplasm contained some elon-
gated mitochondria and ribosomes, few microfibrils and a poorly developed Golgi complex. T h e nucleus was sometimes irregular in shape with chromatin condensed toward the periphery. There were various amounts of collagen fibers and fewer reticulin fibers in the intercellular space. I cells were found in all theca cell tumors a n d in the granulosa cell tumor of a patient with suspected increase in estrogen production. T h e cells were roundish or more irregular in shape than the S cells a n d the nuclear-cytoplasmic ratio varied from cell to cell (Fig. 2). T h e cell membrane was smooth, and there were few desmosomes between the cells. There were more organelles in the cytoplasm of I cells than in that of S cells. Some lipid droplets were found in the cytoplasm but not more than in the cytoplasm of S cells. T h e nuclei were more irregular than the nuclei in S cells. O n e or more nucleolus could be found. Some cells contained microfibrils that were 50 A in diameter and sometimes arranged in whorls (Fig. 3). This material did not stain with the methods used in this study. P cells were found only in the tumors of the patients with signs of elevated estrogen production (Table I). T h e cells were in small nodules or scattered in the tumors and contained a b u n d a n t SER (Figs. 4 and 5). Otherwise they resembled the I cells
Table I. Clinical and histologic findings of 11 cases. Tumor Cell Types8 Age of Patients (years) 51 59 68 61 64 70 70 60
27 32 51
Signs of Increased Estrogen Production None None None Vaginal bleeding, proliferative endometrium, increased level of plasma estradiol Increased plasma estradiol Proliferative endometrium Proliferative endometrium Vaginal bleeding, endometrial carcinoma, proliferative endometrium, increased plasma levels of estrogens None Irregular vaginal bleeding, anovulatory cycles, proliferative endometrium None
Spindle
Intermediate
Thecoma Fibroma Thecoma Thecoma
+ +++ ++ +
++ + + +++
Thecoma Thecoma Thecoma Thecoma
+ + +++ +
+++ +++ + ++
++
+
—
—
+ ++
+
+
+
+++
—
+++
Histologic Type of Tumor
Granulosa cell tumor, microfollicular Granulosa cell tumor, diffuse Granulosa cell tumor, microfollicular
+
Symbols for quantifying cell composition: - none; + some; + + moderate number; +++
IntJ Gynaecol Obstet 17
Polygonal — — —
Granulosa — — —
+ — — —
+ +
—
preponderance.
Theca and granulosa cell tumors
221
^¿JiA'B* Fig. 1. An electron micrograph of an ovarian fibroma. The nuclei (N) of four spindle cells fill the cytoplasm almost completely. Transverse sections of collagen fibers (K) are seen in the intercellular stroma (X7000).
in their ultrastructure. Lipid droplets were numerous in some cells. Numerous mitochondria were elongated with some tubular christae. The G cells were round or somewhat elongated. They also formed Call-Exner bodies whose inner part was composed of lamellated and degenerated cell organelles (Fig. 6). There were many desmosomes between the G cells. In the cytoplasm, there were many round or elongated mitochondria, ribosomes, fibrils of various length, sometimes a welldeveloped Golgi complex and occasionally secondary lysosomes. T h e stroma of the granulosa cell
tumor was composed of collagen bundles and some reticulin fibers within a loose intercellular space.
DISCUSSION Most of our results supported the findings of others concerning the ultrastructure of theca cell tumors (1, 17, 18) and of granulosa cell tumors (13, 15, 17). In addition, we found polygonal cells with abundant SER in the tumors of patients who showed clinical signs of suspected or proven increase
IntJ Gynaecol Obstet 17
222
Klemi and Gronroos
Fig. 2. An electron micrograph of a thecoma with intermediate cells. The intercellular stroma is scant. There are numerous mitochondria (M) in the cytoplasm of the cells (X6600).
Fig. 3. A high magnification electron micrograph of an intermediate cell with abundant microfibrils (m) in the cytoplasm below the nucleus (X27 700).
in estrogen production. Since SER in the cells reflects steroidogenesis, as shown in the human corpus luteum by Crisp et al (6) and Mestwerdt et al (14), we may assume that the P cells in the theca and granulosa cell tumors are the site of steroidogenesis.
IntJ Gynaecol Obstet 17
In the theca cell tumors, there is a gradual change from a spindle cell, via an intermediate cell to a polygonal cell, indicating the stromal origin of polygonal cells. T h e question arises as to the origin of the polyg-
Theca and granulosa cell tumors
223
Fig. 4. An electron micrograph of a polygonal cell with abundant smooth endoplasmic reticulum and some lipid droplets ( x 4700).
V
