Biomedical subjects
M R Caudle
Publications and source records attributed to M R Caudle.
Expression of cell cycle regulatory proteins (p53, pRb) in the human female genital tract.
PURPOSE: Recent studies have shown that proliferation and differentiation of various cell types is regulated by cell-cycle-related proteins, such as protein p53 and retinoblastoma protein pRb. METHODS: Three monoclonal antibodies to p53 (PAb240, PAb421, and PAb1801) and 3H9 monoclonal antibody to pRb were utilized for localization of proteins by peroxidase immunohistochemistry in frozen tissue sections. RESULTS: Nuclear and nucleolar p53 expression was detected in nondividing and relatively stable cells, e.g., oocytes in primordial follicles and granulosa lutein cells. On the other hand, strong cytoplasmic p53 expression was detected in proliferating and low differentiated epithelial cells of the ovarian surface epithelium, amnion, endocervix and ectocervix, indicating enhanced p53 synthesis. Not all three p53 antibodies reacted with each tissue, perhaps due to structural and conformational changes in the p53 molecule, accompanying p53 association with other proteins, e.g., tissue specific transcription factor interactions. pRb expression was usually restricted to the cell nuclei and nucleoli. However, glandular cells of the female reproductive tract showed cytoplasmic pRb expression in juxtaluminal (secretory) segments of cells, a feature not previously described in any cell type. p53 and pRb immunoreactivities declined with advanced differentiation of cells. No p53 or pRb was detected in placental syncytiotrophoblast or terminally differentiated squamous epithelial cells. CONCLUSION: Our data indicate that large quantities of p53 are synthesized in cells leaving the cell cycle and entering differentiation. Except in glandular cells, the pRb expression is confined to the cell nuclei and nucleoli. A unique cytoplasmic expression of pRb in juxtaluminal segments of glandular cells suggests a role for pRb in human female fertility and conception.
Quantitative evaluation of the cell cycle-related retinoblastoma protein and localization of Thy-1 differentiation protein and macrophages during follicular development and atresia, and in human corpora lutea.
Ovarian follicular development is dependent on growth and differentiation of the oocyte, as well as the granulosa and theca cell layers. The majority of primary follicles in the adult human ovary are not growing, and most antral follicles undergo atresia. The mechanisms regulating follicular growth and differentiation are poorly understood. Expression of key regulatory proteins in cells of certain follicles may be involved. We have studied the distribution of retinoblastoma protein (pRb), a key cell cycle regulator, in human follicles and CL by quantitative immunohistochemistry. Recent studies suggest that high nuclear concentrations of pRb are associated with the arrest of cell proliferation and the beginning of differentiation; during advanced differentiation of cells pRb is markedly depleted or absent. We also studied follicular distribution of Thy-1 differentiation protein, a morpho-regulatory molecule associated with cell differentiation, and the presence of macrophages. Macrophages have been shown to stimulate steroidogenesis in granulosa cells in vitro, and they are required for release of Thy-1 differentiation protein from vascular pericytes among granulosa cells in vivo. Our results indicate that oocytes in resting follicles exhibit pRb in the nucleoli. During initiation of follicular growth, the pRb expression first extends over the oocyte nuclei and then diminishes from both nuclei and nucleoli in preantral follicles. When the oocytes reach maximum size in small antral follicles, the pRb expression is reestablished in oocyte nucleoli. In differentiating granulosa and theca cell layers of preantral and small antral follicles, pRb expression is high, but it is low in growing large antral follicles. During CL development and regression, pRb expression in the nuclei of granulosa lutein cells first increases and then decreases. Follicular development is accompanied by the presence of Thy-1 differentiation protein and macrophages under the follicular basement membrane. In growing large antral follicles, during the mid-follicular phase, larger macrophages exhibit physical contacts with granulosa cells through the follicular basement membrane, and, during the late follicular phase, small dendritic macrophages can be detected among granulosa cells, but not within the follicular antrum. Large antral follicles undergoing atresia exhibit strong pRb expression in granulosa cells. This is accompanied by a lack of Thy-1 differentiation protein among granulosa cells and the occurrence of large phagocytic macrophages in the follicular antrum. This is the first report of pRb expression in the human ovary.(ABSTRACT TRUNCATED AT 400 WORDS)
Immunohistochemical studies of the adult human ovary: possible contribution of immune and epithelial factors to folliculogenesis.
PROBLEM: Formation of primordial follicles in adult ovaries could be a cryptic process limited to relatively small areas of the ovarian cortex and occurring during a certain stage of the menstrual cycle. Such an event may require a specific milieu provided by factors involved in developmental processes, i.e., morphoregulatory molecules and macrophages. METHOD: Adult human ovaries were investigated by immunohistochemistry for surface epithelium and granulosa cell markers (cytokeratin 18 and MHC class I), immune system-related morphoregulatory molecules (Thy-1 glycoprotein and N-CAM), and macrophage phenotypes (CD14, CD68, and MHC class II). RESULTS: In some ovaries 300-500 microns areas of surface epithelium were overgrown by tunica albuginea, descended into the stroma, and apparently fragmented into individual small (20-40 microns) follicle-like cell nests. Differentiation of the surface epithelium was accompanied by macrophages and Thy-1 glycoprotein. Small segments of surface epithelium showed N-CAM and a lacked MHC class I expression. In such segments, clear spherical germ-like cells migrated into the deeper stroma, associated with the microvasculature, and eventually aggregated with follicle-like cell nests. CONCLUSIONS: Our data suggest that surface epithelium may be involved in the formation of some primordial follicles in adult ovaries. This process, and further follicular fate, may require a precise interplay of immune system related morphoregulatory molecules and macrophages.
Advanced obstetrical training for family physicians: the future hope for rural obstetrical care.
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Ethanol modulates the hormone secretory responses induced by epidermal growth factor in choriocarcinoma cells.
Analysis of clinical data has implicated ethanol (EtOH) as an embryotoxic agent and as an agent that disrupts normal placental structure and function. Because epidermal growth factor (EGF) is an important regulator of placental function, we have studied the effects of EtOH on EGF-induced hormone secretion using JEG-3 choriocarcinoma cells that serve as a model for trophoblast cells. EtOH at physiological (5-100 mM) concentrations modulated effects of EGF in a time and dose-dependent manner. EGF-induced P4 secretion was increased by 20-100 mM EtOH after a 2-day pretreatment of cells with EtOH, but not after a 6-day pretreatment. Preincubation with 50 mM EtOH doubled the P4 responses to 50 and 100 ng/ml EGF. Although a 2- or 4-day preincubation of cells with 10-50 mM EtOH increased the secretion of E2 in response to 20 ng/ml EGF, a 6-day preincubation inhibited the secretory response to EGF. Pretreatment of cells with 10-50 mM, but not 100 mM EtOH for 2 to 6 days enhanced the human chorionic gonadotropin (hCG) secretory response to EGF. At 50 mm EtOH, the secretion of hCG in response to EGF was increased 2-fold. EtOH also increased basal hCG secretion in a dose-dependent manner between 10-50 mM EtOH. These results suggest that EtOH may modulate EGF-stimulated hormone secretion from cells of placental origin. Such alterations, if they occur in vivo, may impact on the function of the placenta and could potentially explain the pathophysiology of alcohol toxicity during pregnancy.
Interferon-gamma (IFN-gamma) and interleukin-6 (IL-6) in peritoneal fluid and macrophage-conditioned media of women with endometriosis.
PROBLEM: The presence of the various cytokines in human peritoneal fluid has been incompletely evaluated. Changes in cytokine levels may be related to activation of peritoneal macrophages, development of endometriosis, and infertility. This study assesses peritoneal fluid levels of interferon gamma (IFN-gamma) and interleukin-6 (IL-6), and peritoneal macrophage production of IL-6, in women with and without endometriosis. METHOD: Peritoneal fluid was obtained from 62 women at the time of diagnostic or operative laparoscopic surgery for benign gynecologic disease. Peritoneal macrophages were isolated, cultured for 24 h, and the culture media collected. IFN-gamma and IL-6 levels in peritoneal fluid samples and macrophage conditioned media were determined by commercial ELISA. RESULTS: IL-6 was significantly higher in the macrophage conditioned media of women with endometriosis as compared with controls. IL-6 levels were fourfold higher in early stage endometriosis (P < 0.05) and eightfold higher in advanced endometriosis. There were no significant differences between groups in the peritoneal fluid levels of IL-6 or IFN-gamma. CONCLUSIONS: Peritoneal macrophage IL-6 secretion is increased in women with endometriosis, and appears to correlate with disease stage. IFN-gamma does not appear to be responsible for the activation of macrophages in women with endometriosis.
The inhibitory action of taxol on granulosa cell steroidogenesis is reversible.
Taxol is a novel anticancer agent extracted from the bark of Pacific yew trees. The drug has been approved by the FDA for the treatment of advanced ovarian cancer and is in clinical trials for other malignancies, including breast cancer. The goals of this study were to determine whether taxol adversely and irreversibly affects ovarian granulosa cell steroidogenesis. Cultured porcine granulosa cells were treated with taxol (0.12-12 microM) or vehicle (0.01-1% ethanol) in the absence or presence of 10(-9) M hCG in a time- and dose-response study. Morphological changes were recorded every 2 h, and media were collected for the measurement of progesterone (P4) and 17 beta-estradiol. Taxol suppressed both basal P4 and 17 beta-estradiol production and hCG-stimulated P4 production in a time- and dose-dependent manner and drastically changed cell shape by causing disorganization of microtubule bundles and other subcellular organelles. hCG partially reversed the steroid inhibition induced by taxol. These changes are not attributed to ethanol used as the vehicle, because ethanol at higher concentrations than that present in taxol did not suppress P4 production. When taxol was removed from the culture, P4 production returned to control levels. The results of this study show that taxol causes a significant, but reversible, inhibition of granulosa cell steroidogenesis. This inhibitory effect can be partially overcome by co-treatment with hCG.
Cellular localization of luteinizing hormone receptor immunoreactivity in the ovaries of immature, gonadotropin-primed and normal cycling rats.
In this study we used two monoclonal antibodies against purified LH receptor (LHR) to localize and quantify LHR in ovarian compartments during follicular development, using gonads from immature, gonadotropin-primed, and normal cycling rats. In early preantral follicles, LHR immunoreactivity (LHRI) was identified in vascular endothelium and subsequently appeared in vascular pericytes. In healthy small antral (200-550-microns) follicles, LHRI continued to be present in thecal pericytes, but not in cells of the theca interna. However, in small antral follicles undergoing atresia, a dramatic decrease in thecal vessel LHRI with a concomitant increase in LHRI in hypertrophied theca was observed. In healthy antral follicles, LHRI of thecal cells was not observed until the cells reached medium (550-microns) size. High LHRI was occasionally observed in macrophage-like cells adjacent to the oocyte of large preantral follicles and among granulosa layers of medium-sized antral follicles. In the membrana granulosa, LHRI first appeared in cumulus cells of medium-sized antral follicles and subsequently spread to the entire granulosa cell population of large (750 microns) antral follicles. Treatment of immature rats with eCG markedly enhanced LHRI in theca and granulosa cells of all antral follicles, while eCG/hCG-treated (pseudopregnant) rats showed lack of LHRI in follicles and interstitial glands, but not in corpora lutea (CL). Within degenerating CL in the cycling ovary, compared to fresh and mature CL, a significant decrease occurred in intracellular LHRI. Our observations indicate that 1) vascular pericytes may play a role in follicular development; 2) LHR expression in granulosa may require an interaction of macrophages, oocytes, and cumulus cells; and 3) thecal hypertrophy accompanied by enhanced LHR expression occurs on follicles undergoing atresia.
The effect of peritoneal macrophage-derived factor(s) on ovarian progesterone secretion and LH receptors: the role of calcium.
Macrophages and their secretory products, cytokines, play an integral role in many reproductive processes. In this study we examined the effect of conditioned media from cultured human peritoneal macrophages on progesterone production by granulosa cells and the role of calcium in this process. Macrophages were pretreated with various concentrations of a calcium channel blocker (verapamil) or a calcium ionophore (A23187). Macrophage-conditioned media (MCM) or cell-free media that contained calcium channel modifiers were added at three dose levels to cultured porcine granulosa cells. Progesterone production and LH receptor content were determined. Macrophage-conditioned media alone elevated basal progesterone production, but significantly attenuated granulosa cell LH receptor content. These effects were neither potentiated nor suppressed by pretreating macrophages with verapamil. However, production of the LH receptor lowering factor(s) appeared to be suppressed by calcium ionophore. We conclude that (1) one or more factors produced by macrophages have a net stimulatory effect on basal progesterone production and these factor(s) may not be calcium-dependent and (2) macrophage-derived secretory products reduce granulosa cell LH receptor content. The production of these factor(s) may be calcium-dependent.
Hypercalcemic crisis in pregnancy associated with excessive ingestion of calcium carbonate antacid (milk-alkali syndrome): successful treatment with hemodialysis.
Severe hypercalcemia, a potentially life-threatening condition, has been reported rarely during pregnancy. A patient with hypercalcemic crisis associated with excessive ingestion of absorbable calcium antacid was treated successfully with hemodialysis as well as other therapeutic measures, such as saline diuresis. This acute therapy resulted in long-term normalization of maternal calcium levels. The fetus, who exhibited a low biophysical score during the initial admission, was delivered a month later and had an uncomplicated neonatal course. Acute hemodialysis can be an effective, rapid, and safe method of lowering serum calcium levels in pregnant patients with severe hypercalcemia unresponsive to other medical therapies.
Vasa previa.
Diagnosis of vasa previa requires a high index of suspicion. Vasa previa must be included in the differential diagnosis of all cases of third trimester bleeding. When pulsatile vessels are palpated preceding the fetal vertex, vasa previa should be considered along with cord prolapse. Early diagnosis and intervention result in a favorable fetal outcome in this rare condition.
Multiple myeloma in pregnancy: a case report.
We report here a 33-year-old patient found at 20 weeks' gestation to be anemic and to have both a serum M-protein (immunoglobulin [Ig]G kappa) and a kappa-type urinary Bence Jones protein. The concentration of these proteins, measured during pregnancy and in the postpartum period, demonstrated the presence of the maternal serum IgG kappa component in the amniotic fluid and its transmission to, and eventual disappearance from, the newborn.
Current status of anti-zona pellucida antibodies.
It is clear that the mammalian zona pellucida contains tissue-specific antigens that cross-react among certain species. Certain of these antigens generate antibodies that inhibit sperm attachment. Polyclonal antibody production may be an important aspect of this inhibition. In certain species there are other effects of anti-zona antibodies, such as direct action on the ovary. It is uncertain whether immunization with zona antigens will ever be a practical method of contraception in humans. Such vaccination might require unacceptable adjuvants or large amounts of antigen. The persistence and effectiveness of the antibody is not yet proven, and pregnancy has occurred in some despite presence of anti-zona autoantibodies. A safe and effective vaccine may still be found, however, given the large variety of zona pellucida antigens available. The cause of naturally occurring anti-zona pellucida antibodies in humans is unknown. The incidence of these antibodies depends on the assay used. The significance of positivity in a given individual is also uncertain. A number of patients will conceive if other concurrent fertility problems are treated. Positive results should be confirmed by a second method, preferably by testing the sera against human ova. Specific treatment by steroids or other immunosuppressive regimens remains controversial.
Landry-Guillain-Barré-Strohl syndrome in pregnancy. A report of two cases.
Landry-Guillain-Barré-Strohl syndrome (LGBS) in pregnancy is a rare event previously reported on only 29 times. Both cases reported on here were associated with cytomegalovirus infection. The fetus did not appear to be affected by the paralyzing agent in LGBS, although sequelae of the cytomegalovirus could occur later. Aggressive, long-term care is necessary to minimize maternal morbidity.
Clinical significance of naturally occurring anti-zona pellucida antibodies in infertile women.
Although implicated in the etiology of unexplained infertility, a primary role for antibodies to zona pellucida antigens remains uncertain. We therefore prospectively studied 21 infertility patients found to have significant anti-zona pellucida antibody titers (1:4 to 1:16). When treated for endometriosis or other concurrent fertility problems, 7 of the 21 (33%) delivered viable infants. Two other patients conceived without treatment. Decreases in anti-zona antibody titers were found in most but not all women with successful outcomes. In a given individual, anti-zona antibodies may coexist with other fertility disorders. Therefore, these antibodies should not preclude treatment of concurrent fertility disorders.
Thrombocytopenia in preeclampsia: associated abnormalities and management principles.
Severe thrombocytopenia, abnormal liver function, and renal dysfunction may occur as manifestations of preeclampsia. Failure to recognize that this cluster of abnormalities represents a form of preeclampsia may result in erroneous initial diagnoses. Management of 13 such patients has shown a direct correlation between the degree of thrombocytopenia and the measures of liver dysfunction. Platelet counts and liver functions improved prior to delivery in five patients treated with corticosteroids. Management should be directed toward investigation and correction of deranged physiology and appropriate monitoring of maternal-feto-placental status. Early delivery is indicated in patients with progressive thrombocytopenia and in those with evidence of fetal maturity or distress. Provided that the disease process remains stable, consideration should be given in cases of fetal immaturity, to the use of betamethasone therapy. The occurrence of severe thrombocytopenia in 20% of neonates should be a consideration in selecting the mode of delivery.
Circulating immune complexes in pregnancy, preeclampsia, and autoimmune diseases: evaluation of Raji cell enzyme-linked immunosorbent assay and polyethylene glycol precipitation methods.
Sera from 86 individuals were tested for circulating immune complexes by the polyethylene glycol precipitation method and a Raji cell enzyme immunoassay (Raji-ELISA). These included normal nonpregnant control subjects, nonpregnant patients with autoimmune diseases, healthy women in the second and third trimesters of pregnancy, patients with preeclampsia, and women with pregnancies complicated by preexisting autoimmune diseases. Diseases such as systemic lupus erythematosus and rheumatoid arthritis were associated with increased levels of immune complexes in both pregnant and nonpregnant individuals. Circulating immune complexes were not observed in normal pregnancies or in preeclampsia. Although pregnancy itself is not an immune complex-associated state, the presence of immune complexes in autoimmune diseases may explain some of the complications observed during pregnancy in these patients.