Biomedical subjects
S C Bell
Publications and source records attributed to S C Bell.
Secretory proteins of the endometrium--potential markers for endometrial dysfunction.
In contrast to our detailed knowledge of endometrial structure, the study of endometrial function is still in its infancy. It seems likely that most of the changes that occur in the endometrium during the cycle are directed towards the promotion of implantation, and that the proteins secreted by the tissue have a role to play in the maintenance of pregnancy. One approach to the study of these proteins has been the incubation of endometrium in vitro with labelled amino acid precursors, and this method has revealed that in early pregnancy two proteins are quantitatively more important than the others synthesized and secreted by the endometrium. One of these proteins is a homologue of beta-lactoglobulin, also known as alpha 2-PEG, PP14 or PEP. It is synthesized by endometrial glands and no other source has been identified. Its synthesis is induced in the luteal phase of the menstrual cycle, rises to a peak in the early weeks of pregnancy and then rapidly declines as the decidua spongiosa is replaced by decidua compacta. The direction of its secretion appears to be into the amniotic fluid, and its function may be to transport a vitamin or other essential substance to the early conceptus: another possible function is immunosuppression. The protein can be measured in peripheral blood and may be useful as a marker of the effect of progesterone-or its lack of effect, as in the inadequate luteal phase. The other quantitatively important endometrial protein is IGF binding protein, which is produced mainly by stromal fibroblasts in the late luteal phase of the cycle. During the cycle the contribution of the endometrium to serum levels of IGF-BP is slight, but in pregnancy, as the amount of decidua compacta increases, levels in peripheral blood rise, with peaks at 18-24 weeks and at 36-40 weeks. The function of the protein may be to protect the mother by inhibiting IGF, or to enable IGF to stimulate the growth of endometrial stroma. Links between this protein and disorders of fetal growth have been suggested. The diversity of the possible functions of these proteins, and the continuing uncertainty over their roles, underline the complexity of endometrial functions.(ABSTRACT TRUNCATED AT 400 WORDS)
Decidualization and insulin-like growth factor (IGF) binding protein: implications for its role in stromal cell differentiation and the decidual cell in haemochorial placentation.
The somatomedins or insulin-like growth factors (IGFs) are polypeptides which are involved in proliferation of a number of cell types and have been implicated in fetal growth and development. Cellular responses to IGFs are mediated via binding to two classes of trans-membrane receptors. However, IGFs also bind with high affinity to binding proteins (IGF-BP) in plasma and other body fluids. In plasma the majority of IGF is associated with a GH-dependent 150 kd IGF-BP whereas a second small mol. wt (29-35 kd) IGF-BP is relatively unsaturated. It is this latter class of IGF-BP which has been demonstrated to be synthesized or secreted by a number of cell types and is detected in body fluids and secretions such as amniotic fluid, cerebrospinal fluid and milk. Although the exact relationship between these binding proteins and their role in IGF action has not been clarified, evidence suggests that the 29-35 kd form may function as an inhibitor and promoter of IGF action. During the menstrual cycle IGF-BP synthesis of this latter class appears associated with stromal fibroblast populations and it is proposed that this is involved in specifying proliferation of this cell type which differentiates into decidual cells. This IGF-BP also represents the major soluble secretory protein of the decidualized endometrium and its component decidual cell, during pregnancy and probably is the major source of the protein in pregnancy, contributing to the amniotic fluid and peripheral serum levels. In this article the implications of local decidual production of IGF-BP is discussed with reference to the thesis that decidualization, which is associated with species exhibiting haemochorial placentation, is involved in regulatory mechanisms of feto-placental development and growth.
Serum levels of pregnancy-associated endometrial alpha 2-globulin (alpha 2-PEG) during normal menstrual and combined oral contraceptive cycles and relationship to immunohistological localization.
Serum levels of pregnancy-associated endometrial alpha 2-globulin (alpha 2-PEG), the major secretory protein of the endometrium during the late luteal phase of the menstrual cycle and early pregnancy, were measured serially during normal menstrual cycles and in women taking various combined oral contraceptives. Pill users were also sampled on an individual basis. Endometrium from women taking the combined oral contraceptive was examined immunohistochemically using a monoclonal antibody to alpha 2-PEG. Levels of alpha 2-PEG in the menstrual cycle sera showed consistent changes following the luteinizing hormone peak, with a greater than three-fold increase between basal levels at mid-cycle and those in the late luteal phase. Serum levels of alpha 2-PEG in the pill users remained at basal levels throughout the cycle, regardless of the progestagen in the formulation. This finding was noted in serial and individual samples from both monophasic and triphasic preparations. However, there was evidence of induction of alpha 2-PEG production at the local endometrial level when immunohistochemical staining was employed at various stages of the pill cycle. This suggests that serum levels of alpha 2-PEG may not necessarily reflect local endometrial production of the protein. These observations are discussed with reference to the proposed value of serum measurement of this protein in assessing relative progestagenic activity upon the endometrium.
Anti-type II collagen antibody in naturally occurring canine joint diseases.
Autoimmunity to collagen was investigated in several naturally occurring arthropathies of the dog. Increased levels of serum anti-native collagen type II antibody, as assessed by ELISA, were shown in 72.4% of dogs with rheumatoid arthritis (RA), 88% of dogs with infective arthritis (IA) and 52% of dogs with osteoarthritis (OA) (p less than 0.001). The mean levels of antibody in cruciate disease patients (CR) were also significantly increased compared to control dogs (p less than 0.01). Serum anti-collagen antibody in OA dogs correlated with that in precipitated serum immune complexes. There was also a correlation between anti-collagen antibody level in synovial fluid and in synovial fluid complexes in dogs with rupture of the cranial cruciate ligament. In all patient groups, collagenase digestion of polyethylene glycol (PEG) precipitates from sera and synovial fluids caused a significant rise in specific antibody levels to collagen, indicating the presence of collagen-anti-collagen complexes in all arthropathies. In dogs with RA, the levels of collagen-specific antibody in synovial fluid complexes correlated with the total IgG in these complexes. These findings implicate collagen-anti-collagen complexes in the pathogenesis of naturally occurring joint diseases in the dog, but they are unlikely to be the primary aetiological mechanism.
Characterization of an insulin-like growth factor binding protein, analogous to human pregnancy-associated secreted endometrial alpha 1-globulin, in decidua of the baboon (Papio anubis) placenta.
The major secreted protein of the human decidua (pregnancy-associated endometrial alpha 1-globulin [alpha 1-PEG]), is an insulin-like growth factor-binding protein (IGF-BP) that is immunologically and biochemically similar to placental protein 12 (PP12) extracted from term human placenta. Since previous studies have demonstrated that the baboon and human endometrium synthesize and release a number of biochemically and immunologically related polypeptides in culture, this study was undertaken to further characterize a related IGF-BP in baboon placental tissues. Decidua, chorio-amniotic membranes with adhering decidua (CAM-D), and placental villi were obtained from pregnant baboons between Days 134 and 160 of gestation by Cesarean sections. Portions of tissue were either cultured in the presence of 35S-methionine, fixed for immunocytochemistry, or frozen in liquid nitrogen for cytosol extraction. Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) of tissue culture media (TCM) revealed that the major secretory product of the decidua and CAM-D was an acidic polypeptide (Mr 33,000). Western blot analysis and immunoprecipitation of TCM with murine monoclonal antibody (B2H10) against human alpha 1-PEG demonstrated that this molecule, secreted by the baboon decidua and CAM-D, but not the placental villi, was immunologically identical to the human IGF-BP. Immunocytochemical localization of IGF-BP was intense in the cytoplasm of stromal cells in decidua and CAM-D and absent in the placenta. Gel filtration of TCM and cytosol followed by screening of eluates for 125I-IGF-I binding resolved two peaks (Mr greater than 100,000 and 35,000) of specific IGF-BP in decidua and CAM-D. The 35,000 peak had 100-200 times the binding capacity of the Mr greater than 100,000 peak and a Kd of 1.14-1.83 nM. The eluates contained in the Mr 35,000 peak were also immunoreactive to alpha 1-PEG, as accessed by a polyclonal radioimmunoassay. Affinity cross-linking with 125I-IGF-I followed by sodium dodecyl sulfate-PAGE revealed an immunoreactive complex of Mr 36,000, confirming that the baboon protein represents a high affinity IGF-BP. These studies indicate that the hypertrophied stromal cells of the baboon decidua and CAM-D synthesize and release an IGF-BP as their major secretory product, analogous to the situation in humans. The results of this study suggest that this protein may play a role in the regulation of IGF action during pregnancy.
Monoclonal antibodies to human secretory "pregnancy-associated endometrial alpha 1-globulin," an insulin-like growth factor binding protein: characterization and use in radioimmunoassay, Western blots, and immunohistochemistry.
Monoclonal antibodies were raised against pregnancy-associated endometrial alpha 1-globulin (alpha 1-PEG), a 32 KD insulin-like growth factor binding protein (IGF-BP), which represents a major secretory product of the human decidualized endometrium during pregnancy. This class of IGF-BP has been implicated in the modulation of action, inhibitory and stimulatory, of insulin-like growth factors. Immunization with the protein purified from pregnancy endometrium resulted after myeloma fusion in the isolation of six hybridoma clones and the antibodies produced were characterized. The Ka of the antibodies ranged between 4.75 x 10(9) M-1 and 0.7 x 10(8) M-1. In Western blots all monoclonal antibodies reacted with purified protein of molecular weight 32 KD and specifically detected this IGF-BP species in culture medium and cytosolic extracts of pregnancy endometrium and amniotic fluid. The monoclonal antibodies appear to define three epitope-bearing regions as evidenced by their reactivity to polypeptide fragments of the protein. After synthesis and secretion by tissue explants in vitro the protein is susceptible to cleavage into fragments possessing different monoclonal antibody-defined reactivity. Employing immunohistochemical techniques the protein was principally localized to decidual cells in tissue sections of pregnancy endometrium and solely to these cells after enzymic digestion of the tissue. The implications of these results are discussed with respect to potential role of IGF-BP in the action of IGF upon the IGF-1 receptor-bearing populations, including lymphocytes and trophoblast cells, D in the decidua.
Immune complexes and rheumatoid factors in canine arthritides.
Thirty two domestic dogs with naturally occurring polyarthritis were investigated to determine the contribution of autoimmunity in the pathological mechanisms. Comparisons were made with canine infective arthritis (12 dogs), osteoarthritis (32), and osteoarthritis secondary to rupture of the cranial cruciate ligament (19). Rheumatoid factors, immune complexes, and complement fixation (C1q binding) were measured in sera and synovial fluids. Compared with normal dogs (32), dogs with rheumatoid arthritis (RA) had increased serum and synovial fluid immune complexes and rheumatoid factors. Increases were generally also seen in dogs with other arthropathies, however. Rheumatoid factors were higher in sera than in synovial fluids. Rheumatoid factors correlated with immune complex levels and complexed rheumatoid factor only in the group of dogs with RA. Both rheumatoid factors and immune complexes may contribute to the pathogenesis of canine RA but are considered to arise as a result of non-specific inflammatory mechanisms in the non-rheumatoid groups.
An insulin-like growth factor-binding protein in the baboon (Papio anubis) endometrium: synthesis, immunocytochemical localization, and hormonal regulation.
The major secretory product of the baboon and human decidua during pregnancy is an insulin-like growth factor-binding protein (IGF-BP). This study was designed to determine the site and regulation of synthesis of this protein in the nonpregnant baboon based on our previous findings that this molecule is biochemically and immunologically similar in the two species during pregnancy. Endometrial tissue was obtained from cycling baboons and steroid-treated ovariectomized animals. Portions of the tissue were fixed for immunocytochemical analysis, cultured in the presence or absence of cyclohexamide and actinomycin-D, or snap-frozen in liquid nitrogen for RNA isolation. Immunostaining using monoclonal antibodies to IGF-BP indicated that the protein was localized predominantly in the epithelium of the deep glands and was most intense during the late luteal stage of the cycle. Immunoreactive product was also present in tissue from estrogen-primed progesterone-treated ovariectomized animals. However, the staining pattern was more variable and less intense than that in intact animals. Western blots of explant culture media showed the presence of an immunoreactive product only in those tissues that also demonstrated immunocytochemical localization. The absence of an immunoreactive band in medium obtained from tissue incubated in the presence of cyclohexamide suggested that this protein was synthesized de novo. The mRNA coding for IGF-BP appeared to be stable, as synthesis in explant cultures continued in the presence of actinomycin-D. The cDNA probe hybridized to a single message transcript of 1.65 kilobases. The presence of mRNA in tissues coding for this protein correlates with the immunochemical data relating to the site and hormonal regulation of its synthesis. The presence of this protein in the glandular epithelium of the baboon endometrium may have implications in the autocrine and/or paracrine regulation of trophoblast growth and penetration during implantation.
Immunohistological localization of human pregnancy-associated endometrial alpha 2-globulin (alpha 2-PEG), a glycosylated beta-lactoglobulin homologue, in the decidua and placenta during pregnancy.
Monoclonal and polyclonal antibodies to pregnancy-associated endometrial alpha 2-globulin (alpha 2-PEG), a glycosylated human beta-lactoglobulin homologue, were used in an immunohistological technique to determine the cellular localization of this protein in the decidua and placental tissues during pregnancy. During the first trimester the protein was principally localized to the glandular epithelium of the decidua spongiosa region of the endometrium with only weak staining associated with glands of the decidualized decidua compacta region. No significant cellular staining was detected in the decidua capsularis. At term in the decidua of the amniochorion and the placental bed weak staining for alpha 2-PEG was only associated with the epithelium of attenuated glands. No significant staining was detected in the placenta during pregnancy. These results suggest that the epithelium of glands associated with non-decidualized stroma represents the primary source of alpha 2-PEG during the first trimester and that a function of the decidua spongiosa in early pregnancy may be related to production of alpha 2-PEG. The decline in production of alpha 2-PEG during pregnancy is suggested to result from involution of the decidua spongiosa and at term the attenuated glands of the decidua represents the source of alpha 2-PEG.
Human 'pregnancy-associated endometrial alpha 1-globulin', an insulin-like growth factor-binding protein: immunohistological localization in the decidua and placenta during pregnancy employing monoclonal antibodies.
We have previously shown that pregnancy-associated endometrial alpha 1-globulin, a small molecular weight insulin-like growth factor-binding protein (IGF-BP), is quantitatively the major secretory protein product of the decidualized endometrium during human pregnancy. In the present study, employing monoclonal antibodies raised against this protein in an immunohistological technique, the cellular localization of the protein has been examined in the decidua and placenta during pregnancy. During the first trimester the protein was principally associated with the decidual cell in the decidualized decidua compacta region of the endometrium with both cytoplasmic and extracellular matrix-associated staining patterns being detected. No extensive staining was observed in the placenta. At term the protein was localized in similar cells in the placental bed and endometrium associated with the amniochorion but not in the placenta. These studies suggest that the decidual cell represents the major source of IGF-BP during pregnancy and have relevance to the origin of amniotic fluid IGF-BP and the paracrine role of the decidual cell in the control of trophoblast growth.
Immunohistological localization of pregnancy-associated endometrial alpha 2-globulin (alpha 2-PEG) in endometrial adenocarcinoma and effect of medroxyprogesterone acetate.
Endometrium from postmenopausal women with endometrial adenocarcinoma was examined immunohistochemically using a monoclonal antibody to pregnancy-associated endometrial alpha 2-globulin (alpha 2-PEG), the major secretory protein of the glandular epithelium during the late luteal phase of the menstrual cycle and early pregnancy. Specimens were obtained at initial diagnostic curettage and at hysterectomy after medroxyprogesterone acetate (MPA) therapy. alpha 2-PEG was not detected in any malignant tissue irrespective of histological differentiation. Non-malignant endometrium obtained in association with malignant tissue was negative for alpha 2-PEG before treatment although after MPA therapy all specimens obtained exhibited marked alpha 2-PEG localization in glands. In four specimens endogenous alkaline phosphatase was observed consistently only in the malignant endometrium. Malignant endometrium does not appear to synthesize alpha 2-PEG nor is its synthesis induced by an oral progestogen, so that it does not represent a useful marker for endometrial carcinoma. Non-malignant endometrium in postmenopausal women appears to be fully capable of alpha 2-PEG production after stimulation with an oral progestogen.
N-terminal amino acid sequence of human pregnancy-associated endometrial alpha 1-globulin, an endometrial insulin-like growth factor (IGF) binding protein--evidence for two small molecular weight IGF binding proteins.
We have previously demonstrated that quantitatively the major secretory protein of the human endometrium from the latter half of the first trimester of pregnancy, pregnancy-associated endometrial alpha 1-globulin (alpha 1-PEG), is a small mol wt insulin-like growth factor binding protein (IGF BP) and is detected at high levels in amniotic fluid. N-terminal amino acid sequences have been determined for this protein purified from both these sources and a single N-terminal amino acid sequence Ala Pro Trp Gln Cys Ala Pro Cys Ser Ala Glu Lys Leu Ala Leu Pro Pro was obtained. The first ten residues are identical to those previously reported for the small mol wt IGF BP isolated from amniotic fluid and HEPG2 culture medium. However it was different to that reported for the IGF BP isolated from term placenta, placental protein 12 (PP12), a protein previously suggested to originate in the endometrium. These data suggest that at least two distinct forms of the small mol wt IGF BP exist and supports the hypothesis that the amniotic fluid form originates from the pregnancy endometrium.
Immunohistological localization of the human endometrial secretory protein pregnancy-associated endometrial alpha 1-globulin, an insulin-like growth factor-binding protein, during the menstrual cycle.
Pregnancy-associated endometrial alpha 1-globulin (alpha 1-PEG), a 29,000 mol wt insulin-like growth factor-binding protein, is the major secretory protein of the human endometrium from the latter half of the first trimester to the end of pregnancy. It is also produced and secreted by nonpregnant endometrial tissue, especially during the late secretory phase of the menstrual cycle. We studied the localization of this protein in the endometrium during different phases of the cycle using immunohistochemistry with monoclonal antibodies. Immunostaining was first observed in the midsecretory phase and was most consistently detected during the late secretory phase, during which it was principally associated with stromal cell populations. These cells were localized initially surrounding spiral arteries and subsequently also in the subluminal epithelial region of the endometrium. These results suggest that alpha 1-PEG synthesis is principally associated with the process of stromal cell differentiation. i.e. predecidualization. However, the presence of alpha 1-PEG was not directly correlated with the presence of mature predecidual cells, suggesting that its synthesis is not an inherent feature of this cell.
Immunohistological localization of human endometrial secretory protein, 'pregnancy-associated endometrial alpha 2-globulin' (alpha 2-PEG), during the menstrual cycle.
The distribution of alpha 2-PEG, a human analogue of beta-lactoglobulin, in endometrium at different phases of the cycle was determined using immunohistochemistry with monoclonal and polyclonal antibodies. In the epithelial cells of glands in the functional zone of the endometrium, alpha 2-PEG was first detectable from Days 19 to 21 during the mid-luteal phase and maximal immunostaining was observed during the end of the late luteal phase. Intense staining in the glandular secretions and weaker staining in surface luminal epithelial cells during this period were observed. A minor population of basal glands contained alpha 2-PEG during the follicular phase. These results suggest that alpha 2-PEG synthesis by the glandular epithelium of the regenerated endometrium is hormonally regulated. Maximal staining occurring during the late luteal phase suggests that regulation may be related to the hormonal requirement for pre-decidualization rather than that required for histologically defined glandular epithelial secretion.
Major secretory protein of human decidualized endometrium in pregnancy is an insulin-like growth factor-binding protein.
Pregnancy-associated endometrial alpha 1-globulin (alpha 1-PEG) is quantitatively the major secretory protein product, synthesized and secreted in vitro, of the human decidualized endometrium during pregnancy. This protein has been purified from cytosolic extracts of this tissue and has now been characterized as a 32 kDa somatomedin/insulin-like growth factor (IGF)-binding protein. Immunoreactive alpha 1-PEG isolated from amniotic fluid exhibited identical physiochemical properties and IGF-I-binding characteristics. In cytosolic extracts of pregnancy endometrium, in incubation medium of this tissue and in amniotic fluid, the 32 kDa protein represented the major alpha 1-PEG immunoreactive protein and major IGF-I-binding component. Purified alpha 1-PEG and incubation medium of pregnancy endometrium competed for IGF-I with placental membrane IGF receptors in vitro. The implications of the endometrial source of IGF-I-binding protein are discussed with reference to the origin of the amniotic fluid and serum small Mr IGF-binding protein and to the suggested paracrine effect upon trophoblast proliferation.
Secretory endometrial/decidual proteins and their function in early pregnancy.
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Detection and characterization of human secretory "pregnancy-associated endometrial alpha 2-globulin" in uterine luminal fluid.
Pregnancy-associated endometrial alpha 2-globulin (alpha 2-PEG), the major secretory protein of the human endometrium as assessed by in vitro de novo synthesis and secretion, during the mid- to late-luteal phase of the menstrual cycle and early first trimester pregnancy, has now been detected immunochemically in uterine luminal flushings during the luteal phase of the menstrual cycle. Physicochemical characterization of this immunoreactive alpha 2-PEG demonstrated that it exhibited identical properties to the protein isolated from either the cytosol of pregnancy endometrium or medium from in vitro cultures of pregnancy endometrium, i.e. native Mr 56 K, subunit Mr 28 K, eluted from an anion exchange column at 0.11 M NaCl and bound to concanavalin A. These findings, together with other evidence, suggest that a major route of secretion of alpha 2-PEG during the luteal phase of the menstrual cycle in vivo is into the lumen of the uterus. These results are discussed with reference to the hypothesis that alpha 2-PEG, derived from the endometrial glandular epithelium, may represent the human functional analogue of uteroglobin and to the implications for the human of the hypothesized immunosuppressive role of uteroglobin and transglutaminase (Factor XIII) in masking Class I MHC antigens on gametes and the conceptus.