Biomedical subjects

N Chegini

Publications and source records attributed to N Chegini.

At least 55 records · Page 3Linked to original sources

PubMed1996

Gonadotropin-releasing hormone (GnRH) and GnRH receptor gene expression in human myometrium and leiomyomata and the direct action of GnRH analogs on myometrial smooth muscle cells and interaction with ovarian steroids in vitro.

The objective of the present study was to determine whether GnRH and GnRH receptor are expressed in myometrium and leiomyomata, and if GnRH analogs alone or in the presence of ovarian steroids can modulate the rate of DNA synthesis, proliferation, and transforming growth factor-beta 1 (TGF beta 1) production in myometrial smooth muscle cells in vitro. Reverse transcription-PCR revealed that leiomyomata, unaffected myometrium, and isolated myometrial smooth muscle cells express GnRH and GnRH receptor messenger ribonucleic acid. Furthermore, in a dose-dependent manner, GnRH agonist (leuprolide acetate) inhibited, but GnRH antagonist [D-pGlu1,D-Phe2,D-Trp3.6] (GnRH-Ant1) stimulated, the rate of [3H]thymidine incorporation into myometrial smooth muscle cells (P < 0.05), whereas GnRH-Ant2 (Ac-D-P-Cl-Phe1.2,D-Trp3,D-Arg6,D-Ala10) had no effect. 17 beta-Estradiol (E2) medroxyprogesterone acetate (MPA), and E2 plus MPA (1 micromol/L) stimulated the rate of DNA synthesis by smooth muscle cells (P < 0.05), which was inhibited by GnRH analogs used at 5 micromol/L (P < 0.05). GnRH analogs had no significant effect on myometrial smooth muscle cell proliferation, with the exception of GnRH-Ant1; however, they inhibited the stimulatory action of E2, MPA, and E2 plus MPA in a time-dependent manner (P < 0.05). These cells also synthesized and released approximately 1.32 +/- 0.02 ng/mL total (active plus latent) TGF beta 1, of which 0.73 +/- 0.02 ng/mL was in an active form. E2, MPA, E2 plus MPA, and GnRH analog treatments resulted in an increase in total TGF beta 1 production, whereas GnRH agonist and GnRH-Ant2, but not GnRH-An1, inhibited active TGF beta 1 (P < 0.05). GnRH analogs also inhibited the action of E2 plus MPA on total and active TGF beta 1 production, whereas GnRH-Ant1 further stimulated E2, MPA, or E2 plus MPA action on active TGF beta 1 production (P < 0.05). The data demonstrate for the first time that GnRH and GnRH receptor messenger ribonucleic acid are expressed in myometrium, leiomyomata, and myometrial smooth muscle cells. The local expression of GnRH and receptor along with the direct action of GnRH analogs on the smooth muscle cell DNA synthesis and TGF beta 1 production suggest an autocrine/paracrine role for GnRH in these tissues, a mechanism that may be involved in leiomyomata regression in women receiving GnRH agonist therapy.

Adult↗

PubMed1996

Suppression of transforming growth factor-beta (TGF beta) and TGF beta receptor messenger ribonucleic acid and protein expression in leiomyomata in women receiving gonadotropin-releasing hormone agonist therapy.

The expression and cellular distribution of transforming growth factor-1 (TGF beta 1) through TGF beta 3 and TGF beta type I-III receptor messenger ribonucleic acid (mRNA) and protein were analyzed in leiomyomata from patients receiving GnRH agonist (GnRHa; leuprolide acetate) compared to those in untreated controls. Standard reverse transcription-PCR revealed that the unaffected myometrium and leiomyomata from leuprolide-treated and untreated patients express TGF beta 1-3 and TGF beta type I-III receptor mRNA. The myometrial and leiomyomata smooth muscle cells were the primary site of TGF beta 1-3 and TGF beta type I and II receptor mRNA and protein expression, as determined by in situ hybridization and immunohistochemical localization. These observations indicate that leiomyomata express a higher of level of TGF beta and TGF beta receptor mRNA and protein than unaffected myometrium during the secretory phase of the menstrual cycle, and women who received leuprolide acetate therapy had a substantially lower level of expression than untreated controls. Furthermore, competition-based quantitative reverse transcription-PCR using synthetic internal standards revealed that leiomyomata express a significantly higher number (copies per cell) of TGF beta type II receptor mRNA, followed by TGF beta 1, TGF beta type I receptor, TGF beta 2, and TGF beta 3 (P < 0.05). However, there was a significant decrease in the levels (copies per cell) of TGF beta 1, TGF beta 3, and TGF beta type I and type II receptor mRNA expression in leiomyomata from leuprolide-treated compared to untreated patients (P < 0.05). The data provide further evidence that leiomyomata express mRNA and protein for all components of the TGF beta system, and GnRHa therapy results in down-regulation of their expression. More specifically, these data suggest that TGF beta 1 and TGF beta 3 may play a more important role in leiomyomata growth than TGF beta 2, which leads us to propose that lowering TGF beta and receptor expression may have a direct effect on leiomyomata regression.

Adult↗

PubMed1996

Detection of transforming growth factor-alpha mRNA and protein in rat lacrimal glands and characterization of transforming growth factor-alpha in human tears.

PURPOSE: To assess whether the lacrimal gland is a possible site of synthesis of transforming growth factor-alpha (TGF-alpha) and to characterize TGF-alpha biochemically in human tears. METHODS: Reverse transcription-polymerase chain reaction (RT-PCR) amplification was used to analyze rat lacrimal glands for the presence of TGF-alpha mRNA. Specific monoclonal antibodies were used to localize TGF-alpha immunohistochemically in lacrimal gland tissue of rats. Human tears were analyzed for immunoreactive TGF-alpha protein using a specific radioimmunoassay, and the molecular weight of TGF-alpha in tears was characterized by Western blot analysis. RESULTS: RT-PCR amplification of rat lacrimal gland RNA generated a band of the predicted 492 base pairs for TGF-alpha mRNA. Immunohistochemical staining of rat lacrimal gland localized TGF-alpha protein to lacrymocytes constituting acini but not to interacinar and intraacinar ducts of lacrimal glands. Western blot analysis of human tears detected a single band at MWt 16,000. Logit transformation of radioimmunoassay data for tears and TGF-alpha standard generated parallel displacement lines, indicating the presence of immunoreactive TGF-alpha levels in human tears with an average concentration of 100 +/- 20 pg/ml (mean +/- SEM). CONCLUSIONS: Rat lacrymocytes synthesize TGF-alpha mRNA and protein, and human tears contain immunoreactive TGF-alpha, suggesting that the lacrimal gland may be an exocrine source for TGF-alpha in tears. The single MWt 16,000 form of TGF-alpha in human tears appears to be generated by an unusual proteolytically processing of the pro-TGF-alpha transmembrane precursor protein.

Adult↗

PubMed1995

Modulation by benzo[a]pyrene of epidermal growth factor receptors, cell proliferation, and secretion of human chorionic gonadotropin in human placental cell lines.

Clinical observations indicate that maternal cigarette smoking has significant detrimental effects on fetoplacental development. The present study used human trophoblastic choriocarcinoma cell lines of placental origin to investigate the effects of benz[a]pyrene (BaP) on epidermal growth factor (EGF) receptors, cell proliferation and human chorionic gonadotropin (hCG) secretion. BaP decreased 125I-EGF binding and EGF receptor protein in a concentration-related manner in both BeWo and JEG-3 cell lines. The steady-state level of EGF receptor mRNA, however, was not changed significantly by BaP in either cell line. Cell proliferation was unchanged or slightly increased following exposure to 10 and 50 microM BaP in the presence of serum, whereas proliferation progressively decreased in cells exposed under serum-free conditions. The mitogenic effect of EGF was inhibited by cotreatment with BaP in both cell lines. Further study of trophoblast endocrine function showed that both basal and EGF-stimulated secretion of hCG was reduced significantly by BaP exposure in BeWo cells, whereas no adverse effect was seen in JEG-3 cells. Finally, cytochrome P450 1A1 (CYP1A1) was induced in a concentration-dependent manner by BaP in both cell lines. Thus, data indicate that the BaP-mediated loss of EGF receptors alters trophoblast proliferation and endocrine function, and that different mechanisms may be involved in the regulation of hCG secretion in BeWo and JEG-3 cells. In addition, this study supports the feasibility of using the BeWo and JEG-3 trophoblastic choriocarcinoma cell lines to investigate biomarkers and mechanisms of placental toxicity.

Benzo(a)pyrene↗

PubMed1995

Detection of insulin and insulin-like growth factors I and II in saliva and potential synthesis in the salivary glands of mice. Effects of type 1 diabetes mellitus.

The salivary glands of mammals synthesize and secrete a number of peptide growth factors that play important roles in cell/tissue homeostasis and embryonic development. Using a radioimmunoassay, insulin, insulin-like growth factor-I (IGF-I) and insulin-like growth factor-II (IGF-II) were detected in saliva from mice. Unlike epidermal growth factor (EGF), there was no sexual dimorphism in the concentrations of the insulin growth factor family. Immunohistochemical localization of IGF-I and IGF-II was confined to the duct cells of both the parotid and the submandibular glands. Reverse transcriptase-polymerase chain reaction amplification of total RNA from parotid and submandibular glands confirmed the presence of all three hormone/growth factor mRNAs in both glands. The levels of insulin and IGF-I were higher in saliva from an animal model for autoimmune type 1 diabetes, the non-obese diabetic (NOD) mouse, than in a second inbred strain, BALB/c. In contrast, the IGF-II levels were decreased relative to the BALB/c strain. With the onset of diabetes in NOD mice, insulin levels declined, while IGF-I and IGF-II levels showed trends toward lower levels of these growth factors when compared with non-diabetic animals. These changes were reflected in the concentrations from parotid and submandibular gland cell lysates.

Animals↗

PubMed1995

Human fallopian tube as an extraovarian source of relaxin: messenger ribonucleic acid expression and cellular localization of immunoreactive protein and 125I-relaxin binding sites.

By means of specific human relaxin primers that originated from relaxin A and B chains, a monoclonal antibody, and 125I-relaxin, the expression of mRNA and immunoreactive protein and the presence of binding sites for relaxin were investigated in human fallopian tubes. Reverse transcription polymerase chain reaction (RT-PCR) analysis of total RNA isolated from tubal tissues revealed the predicted 434-bp fragments originating from both the H1 and H2 relaxin genes. Restriction enzyme digestion of the RT-PCR products with Msp I (Hpa II), present only in the relaxin H1 sequence, resulted in the anticipated 175- and 259-bp fragments, whereas digestion with Hpa I, which is present in the relaxin H2 sequence and should have resulted in 188- and 246-bp fragments, induced a limited and partial digestion of the product. Codigestion of the RT-PCR product with Msp I+Hpa I also resulted in 175- and 259-bp fragments. The immunoreactive relaxin protein was present primarily in the tubal epithelial cells of the ampullary and isthmus regions, and with weaker intensity in tubal smooth muscle cells. Immunoreactive relaxin either was barely present or was absent in other tubal cell types. The intensity of immunostaining for relaxin in the epithelial cells appeared not to be cycle-dependent; however, these cells showed a lower immunostaining at the late secretory phase of the menstrual cycle than at other reproductive stages and an absence of immunostaining during the postmenopausal period.(ABSTRACT TRUNCATED AT 250 WORDS)

Autoradiography↗

PubMed1995

Expression and selective cellular localization of granulocyte-macrophage colony-stimulating factor (GM-CSF) and GM-CSF alpha and beta receptor messenger ribonucleic acid and protein in human ovarian tissue.

Reverse transcription-polymerase chain reaction (RT-PCR), in situ hybridization, and immunohistochemical observations revealed that human ovarian tissue expressed granulocyte-macrophage colony-stimulating factor (GM-CSF) as well as GM-CSF alpha and beta receptor (R) mRNA and protein. The RT-PCR products revealed the predicted 286-, 546-, and 380-bp fragments for GM-CSF, GM-CSF alpha R, and GM-CSF beta R, respectively, which were further verified by restriction enzyme digestion analysis. In situ hybridization revealed that the theca interna of the large follicles and luteal cells are the exclusive site of GM-CSF mRNA expression. Furthermore, immunohistochemical studies using specific monoclonal antibodies indicated that the theca interna of the large follicles are the exclusive site of GM-CSF protein, whereas theca externa and to a lesser extent the granulosa cells are the major sites of GM-CSF alpha R and beta R proteins. Atretic follicles and follicular cysts showed very low or no detectable levels of GM-CSF and GM-CSF alpha R and beta R. In the luteal tissue, both the small and large luteal cells of early luteal (Days 14-19) and midluteal (Days 20-25) phase expressed GM-CSF mRNA and protein as well as GM-CSF alpha R and beta R proteins, and their immunostaining intensity was similar to that seen with theca cells. Luteal cells from late luteal phase (Days 26-28), corpus albicans, and ectopic pregnancy expressed a low level of GM-CSF, GM-CSF alpha R, and GM-CSF beta R mRNA and protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗

PubMed1994

Characterization of the synthesis and expression of the GTA-kinase from transformed and normal rodent cells.

The murine transformed cell line YC-8 and beta-adrenergic receptor agonist (isoproternol) treated rat and mouse parotid gland acinar cells ectopically express cell surface beta 1-4 galactosyltransferase during active proliferation. This activity is dependent upon the expression of the GTA-kinase (p58) in these cells. Using total RNA, cDNA clones for the protein coding region of the kinase were isolated by reverse transcriptase-PCR cloning. DNA sequence analysis failed to show sequence differences with the normal homolog from mouse cells although Southern blot analysis of YC-8, and a second cell line KI81, indicated changes in the restriction enzyme digestion profile relative to murine cell lines which do not express cell surface galactosyltransferase. The rat cDNA clone from isoproterenol-treated salivary glands showed a high degree of protein and nucleic acid sequence homology to the GTA-kinase from both murine and human sources. Northern blot analysis of YC-8 and a control cell line LSTRA revealed the synthesis of a major 3.0 kb mRNA from both cell lines plus the unique expression of a 4.5 kb mRNA in the YC-8 cells. Reverse transcriptase-PCR of LSTRA and YC-8 confirmed the increased steady state levels of the GTA-kinase mRNA in YC-8. In the mouse, induction of cell proliferation by isoproterenol resulted in a 50-fold increase in steady state mRNA levels for the kinase over the low level of expression in quiescent cells. Expression of the rat 3' untranslated region in rat parotid cells in vitro led to an increased rate of DNA synthesis, cell number an ectopic expression of cell surface galactosyltransferase in the sense orientation. Antisense expression or vector alone did not alter growth characteristics of acinar cells. A polyclonal antibody monospecific to a murine amino terminal peptide sequence revealed a uniform distribution of GTA-kinase over the cytoplasm of acinar and duct cells of control mouse parotid glands. However, upon growth stimulation, kinase was detected primarily in a perinuclear and nuclear immunostaining pattern. Western blot analysis confirmed a translocation from a cytoplasmic localization in both LSTRA and quiescent salivary cells to a membrane-associated localization in YC-8 and proliferating salivary cells.

Amino Acid Sequence↗

PubMed1994

Epidermal growth factor, transforming growth factor alpha, transforming growth factor beta, acidic fibroblast growth factor, basic fibroblast growth factor, and interleukin-1 proteins in the cornea.

The purpose of this study was to determine whether epidermal growth factor (EGF), EGF receptor, transforming growth factor alpha (TGF-alpha), transforming growth factor beta (TGF-beta), acidic fibroblast growth factor (acidic-FGF), basic fibroblast growth factor (basic-FGF), and interleukin-1-alpha (IL-1-alpha) proteins were present in cultures of human corneal cells and/or in sections of human corneal tissue. Immunohistochemistry was performed on human corneal sections. Immunofluorescent cell staining was used to evaluate corneal epithelial, stromal fibroblast, and endothelial cells in primary culture. Basic-FGF production was evaluated in culture cells using immunoprecipitation. EGF, TGF-alpha, TGF-beta-1, and IL-1-alpha were detected by immunohistochemistry in cells in all three layers of the cornea. EGF receptor and acidic FGF were detected by immunohistochemistry in epithelial and endothelial cells, but not in stromal fibroblast cells. Differences in distribution of the growth factors were noted within individual layers of the cornea. EGF and basic-FGF proteins were detected in all three predominant cell types of the cornea using immunocytology. IL-1-alpha protein was detected by immunocytology in corneal epithelial and endothelial cells, but not stromal fibroblasts. Immunoprecipitation confirmed the production of basic-FGF in all three cell types. IL-1-alpha protein detection in the corneal stroma by immunohistology, but not by immunocytology in first passage stromal fibroblasts, suggests that IL-1-alpha may localize to the corneal stroma after production by corneal epithelial and/or endothelial cells.

Adult↗

PubMed1994

Neem oil inhibits two-cell embryo development and trophectoderm attachment and proliferation in vitro.

PURPOSE: The in vitro effect of neem oil was studied on the development of mouse two-cell embryos and trophectodermal cell attachment and proliferation. METHOD: Female mice were primed with gonadotropins for superovulation and caged with male mice. Early embryos, at the two-cell and the blastocyst stages, were recovered at 40 and 88 hr post-hCG from the oviducts and the uteri, respectively. In the first experiment, two-cell embryos were exposed to culture medium containing different concentrations of neem oil for 1, 12, and 24 hr and then grown in neem oil-free culture medium and assessed for the formation of total and hatching blastocysts at 96 hr. In the second experiment, partially hatching blastocysts were cocultured with human endometrial stromal cell monolayers in culture medium containing different concentrations of neem oil and assessed for the attachment and proliferation of trophectodermal cells at 96 hr. RESULTS: Exposure of two-cell embryos to neem oil concentrations of 0.050-0.500% for 1 hr, 0.010-0.250% for 12 hr, and 0.005-0.100% for 24 hr caused significant inhibition of the formation of total and hatching blastocysts, in a dose-dependent manner. Neem oil at 0.050-0.100% concentrations inhibited, in a dose-dependent manner, the in vitro attachment and proliferation of trophectodermal cells of partially hatching blastocysts cocultured with human endometrial stromal cells monolayers. CONCLUSION: Neem oil inhibits the development of two-cell embryos and attachment and proliferation of the trophectodermal cells of partially hatching blastocysts in vitro. The study encourages the use of this herbal product as a postcoital contraceptive that warrants further research.

Animals↗

PubMed1994

Identification of epidermal growth factor, transforming growth factor-alpha, and epidermal growth factor receptor in surgically induced pelvic adhesions in the rat and intraperitoneal adhesions in the human.

OBJECTIVE: Our purpose was to determine the presence and cellular distribution of epidermal growth factor, transforming growth factor-alpha, and epidermal growth factor receptor in surgically induced pelvic fibrous adhesions in rat uterine horns subjected to burn, crush, and debridement injury and intraperitoneal fibrous adhesions formed to various organs in the human. STUDY DESIGN: A total of 15 injured and five uninjured rats were used in this study, and fibrous adhesions and intact peritoneum were removed for processing 2 weeks after surgery. Fibrous adhesions formed to uterine, ovarian, and oviductal tissues and the peritoneal wall from eight patients who had gynecologic surgery were also collected. The tissues were processed for immunohistochemical localization of epidermal growth factor, transforming growth factor-alpha, and epidermal growth factor receptor with specific antibodies to human and rat epidermal growth factor and transforming growth factor-alpha and the extracellular binding domain of the epidermal growth factor receptor. RESULTS: All the cell types in the rat fibrous adhesion immunostained for epidermal growth factor, transforming growth factor-alpha, and epidermal growth factor receptor. The highest immunostaining intensity for epidermal growth factor was associated with inflammatory cells infiltrated into the fibrous adhesion, followed by arteriole endothelial and smooth muscle cells, fascial striated muscle, and fibroblasts of the fibrous adhesion. In the uterine tissue at the site of injuries myometrial smooth muscle cells, in addition to inflammatory cells that migrated among stromal cells, also immunostained for epidermal growth factor. Fibrous adhesions also immunostained for transforming growth factor-alpha with three separate polyclonal antibodies to the amino and carboxy termini of transforming growth factor-alpha precursor and the mature transforming growth factor-alpha, with no substantial differences in their intensity and pattern compared with epidermal growth factor. The pattern and cellular distribution of epidermal growth factor receptor was similar to that seen for epidermal growth factor and transforming growth factor-alpha. Fibrous adhesions from patients with intraperitoneal adhesions immunostained for epidermal growth factor, transforming growth factor-alpha, and epidermal growth factor receptor with a pattern and intensity similar to that observed in fibrous adhesions in the rats. CONCLUSIONS: The data suggest that epidermal growth factor and transforming growth factor-alpha may play a key role both in normal mechanism of peritoneal repair after injury and formation and maintenance of fibrous adhesions.

Animals↗

PubMed1994

Growth factors and ocular wound healing.

Protein growth factors regulate many of the processes in vitro that are essential for the process of normal ocular wound healing, including migration, mitosis and differentiation of cells. This has led to the hypothesis that peptide growth factors play key roles in regulating normal ocular wound healing in vivo. A corollary to this concept is that insufficient action of growth factors causes impaired healing, and prolonged action of growth factors produces excessive scarring. If both of these concepts are correct, then the addition of exogenous protein growth factors should enhance healing of chronic ocular wounds and reducing prolonged actions of growth factors should limit excessive scarring. Although much remains to be understood about the role of growth factors in ocular development and wound healing, results of a substantial number of laboratory and clinical experiments indicate that these hypotheses are generally correct. This article reviews the results of pre-clinical experiments and clinical trials investigating the roles of protein growth factors in ocular development and wound healing.

Animals↗

PubMed1994

The effect of surgical glove powder on proliferation of human skin fibroblast and monocyte/macrophage.

The effect of surgical glove powders (Biosorb, Keoflo, and CaCO3) and Hydrocote (powder-free lubricating agent, Biogel) was examined on human skin fibroblasts and monocyte/macrophage cell lines (U937 and HL-60). Glove powders (0.1-100 micrograms/ml) in the presence of 10% fetal bovine serum (FBS) had no significant effect on the rate of 3H-thymidine uptake and proliferation of these cells after 48 h and 7 days of exposure, respectively. However, they inhibited HL-60 growth after 10 days, and Biosorb and CaCO3 inhibited U937 after 10-21 days of exposure compared with control. In the presence of low serum (0.5%), Biosorb, but not Keoflo, CaCO3, and Hydrocote, inhibited HL-60 cells after the third day of exposure (p < 0.05), whereas they were without any effect on U937 cells. Further incubation resulted in a significant decrease in cell density in all treatments, as well as controls, because of cell death. In the presence of 2% serum, glove powder-treated HL-60 significantly increased in cell numbers during the first 3 days, and the cells became stationary thereafter, whereas Keoflo and CaCO3-stimulated U937 reached a maximal by 9 days of treatment. Coculturing of fibroblasts directly with macrophages (0.4-5 x 10(5) cells per dish) or incubation with macrophage culture-conditioned media (CCM) stimulated quiescent fibroblast growth equal to that induced by 10% and 0.5% serum, respectively (p < 0.05). However, incubation of fibroblasts with glove powder-treated HL-60 CCM (except CaCO3) inhibited (p < 0.05) and CCM from Biosorb-treated U937 stimulated (p < 0.05) fibroblast proliferation. The CCM from glove powder-treated HL-60 and U937 did not have any significant effect on the rate of 3H-thymidine incorporation into fibroblasts compared with controls. The present observations suggest that glove powder action on fibroblast and macrophage growth in vitro depends on both the serum concentration of the culture medium and the length of exposure. The results imply that glove powders may have an adverse effect in vivo by directly influencing the biologic activity of macrophages, as well as other cell types, leading to alterations in the early phases of wound healing.

Calcium Carbonate↗

PubMed1994

Immunohistochemical localization of insulin-like growth factor (IGF-I), IGF-I receptor, and IGF binding proteins 1-4 in human fallopian tube at various reproductive stages.

Despite the existence of numerous data regarding the insulin-like growth factor (IGF) system in reproductive tissues, especially uterine and ovarian, very little information is available concerning their presence in oviductal/fallopian tube tissue. To elucidate this, the present study was undertaken to determine the presence and cellular distribution of IGF-I, IGF-I receptor (IGF-IR), and IGF binding proteins (IGFBP) 1-4 in human fallopian tubes during various reproductive stages, by means of immunohistochemical analysis with specific antibodies to IGF-I, IGF-IR and IGFBPs 1-4. The primary site of immunoreactivity for these proteins in fallopian tube tissue was in the epithelial lining of the tubes, with substantially lower intensity in the smooth muscle layer, fibroblasts of the serosal tissue, and arteriolar endothelial and smooth muscle cells. The immunostaining was associated with both ciliated and nonciliated tubal epithelial cells without substantial differences in their intensity. There were also no differences in immunoreactivity of IGF-I, IGF-IR, and IGFBPs 1-4 present in ampullary vs. isthmus region of the tubes. Specimens obtained 5-12 years post tubal ligation stained similarly to sections from unligated tubes taken during the same phase of the cycle. The intensity of immunostaining for the IGFBPs was greatest for IGFBP-1, followed by IGFBP-4, then IGFBP-2 and IGFBP-3, with 2 and 3 having similar intensities. The immunostaining intensity of IGF-I, IGF-IR, and IGFBPs in fallopian epithelial cells was cycle-dependent and considerably higher in late proliferative and early-mid secretory compared to late secretory phases, with little immunostaining in the early proliferative phase of the menstrual cycle and postmenopausal period. In summary, the immunohistochemical study reported here demonstrates the presence of IGF-I, IGF-IR, and IGFBPs 1-4 in the human fallopian tube during various reproductive stages. These data suggest an autocrine/paracrine role for the IGF-I system in fallopian tube function, and their cyclic dependency further implies ovarian steroidal regulation.

Animals↗

PubMed1994

Expression of messenger ribonucleic acid and presence of immunoreactive proteins for epidermal growth factor (EGF), transforming growth factor alpha (TGF alpha) and EGF/TGF alpha receptors and 125I-EGF binding sites in human fallopian tube.

Reverse transcription polymerase chain reaction (RT-PCR) revealed that the Fallopian tubes express epidermal growth factor (EGF), transforming growth factor (TGF alpha), and EGF receptor (EGF-R) mRNA. The RT-PCR product was verified by restriction enzyme digestion analysis. Immunohistochemically, EGF, TGF alpha, and EGF-R were localized in Fallopian tubes by use of specific antibodies to human EGF, mature fragments of human TGF alpha, and monoclonal antibodies to the extracellular binding domain of EGF-R. The tubal epithelial cells were the primary site of immunoreactive EGF, TGF alpha, and EGF-R, which were present to a lesser extent in the stromal cells, smooth muscle cell layers, fibroblasts of serosal tissue, and arterial endothelial and smooth muscle cells. Using antibodies generated against the amino and carboxy termini of TGF alpha precursor produced a similar cellular distribution to that observed for mature TGF alpha. The intensity of immunoreactive TGF alpha with these antibodies was similar to that seen with EGF. The ciliated and nonciliated epithelial cells in the ampullary and isthmus regions immunostained with similar intensity for EGF, TGF alpha, and EGF-R. The immunostaining for EGF, TGF alpha, and EGF-R was cycle-dependent, was considerably higher during late proliferative and early-to-mid-secretory phases than during early proliferative and late secretory phases of the menstrual cycle, and was reduced during the postmenopausal period. Specimens obtained 5-12 yr after tubal ligation immunostained for EGF, TGF alpha, and EGF-R similarly to sections from unligated tubes taken during the same phase of the cycle. Quantitative autoradiography of 125I-EGF binding generated a pattern similar to that of immunostaining for EGF-R binding. Net grain density/100 microns 2 calculated for different cell types indicated that the epithelial cells had a significantly higher grain density than did other tubal cell types (p < 0.05) without the cycle dependency seen in the immunohistochemical study. In summary, the results demonstrate that the human Fallopian tube expresses mRNA and contains immunoreactive proteins for EGF, TGF alpha, and EGF-R as well as binding sites for 125I-EGF. The cycle dependency and lower immunostaining in postmenopausal tubes suggest a potential regulation of their expression by ovarian steroids. The results imply the importance of EGF/TGF alpha in a variety of tubal biochemical and physiological functions and possibly early embryonic development.

Autoradiography↗

PubMed1994

Insulin-like growth factor I (IGF-I), IGF-I receptors, and IGF binding proteins 1-4 in human uterine tissue: tissue localization and IGF-I action in endometrial stromal and myometrial smooth muscle cells in vitro.

The objective of the present study was to elucidate the presence and cellular distribution of insulin-like growth factor I (IGF-I), IGF-I receptor (IGF-IR), and IGF binding proteins (IGFBPs) in human uterine tissue at various reproductive stages, and to determine the effect of IGF-I and its interaction with epidermal growth factor (EGF) and platelet-derived growth factor (PDGF) in endometrial stromal and myometrial smooth muscle cells in primary culture. Using specific antibodies, immunohistochemical observations indicated that luminal and glandular epithelial cells were the major sites of immunoreactive IGF-I, IGF-IR, and IGFBPs 1-4, followed by myometrial smooth muscle and endometrial stromal cells. The immunostaining intensity of IGF-I, IGF-IR, and IGFBPs in endometrial but not myometrial tissue was cycle-dependent and higher in the late proliferative and early/mid-secretory periods than in the late secretory and postmenopausal periods, with little immunostaining at the early proliferative phase of the menstrual cycle. Stromal and smooth cells in primary cell culture also contained immunoreactive IGF-I, IGF-IR, and IGFBPs. IGF-I at 10-100 ng/ml stimulated 3H-thymidine incorporation in quiescent stromal and smooth muscle cells with maximal effect at 100 ng/ml (p < 0.05). However, in the presence of 2% serum, which induces half-maximal stimulation, IGF-I (100 ng/ml) further increased the rate of 3H-thymidine incorporation in stromal but not smooth muscle cells (p < 0.05). The effect of IGF-I was significantly lower than that induced by EGF (10 ng/ml), PDGF-BB (10 ng/ml) and their combination (p < 0.005), and higher in stromal cells from proliferative, than secretory phase of the cycle in the presence of 2% fetal bovine serum, but not serum-free condition (p < 0.005). The effect of IGF-I on myometrial smooth muscle cells was significantly higher than that induced by EGF, but lower than that induced by PDGF-BB or by EGF+PDGF-BB, without the cycle specificity seen with stromal cells. EGF, PDGF-BB, and their combination with IGF-I, but not IGF-I alone, stimulated stromal and smooth muscle cell growth as determined by a cell proliferation assay. The results indicate that human uterine tissue at various reproductive stages contains immunoreactive IGF-I, IGF-IR, and IGFBPs 1-4. Although IGF-I alone was found to be a weak mitogenic factor for stromal and smooth muscle cells, by interacting with EGF and PDGF-BB in a cycle-dependent manner it may regulate the growth and differentiation of these and other uterine cell types.

Adult↗

PubMed1994

Characterization of the synthesis and secretion of transforming growth factor-alpha from salivary glands and saliva.

Whole saliva collected from rat, mouse, and human sources was found to contain high concentrations of transforming growth factor-alpha (TGF alpha) when analyzed by RIA. The concentrations of TGF alpha in unstimulated human saliva (age, 30-45 yr; n = 10; 1.5 +/- 3.1 nM) was reduced with age (age, 55-70 yr; n = 10; 0.4 +/- 0.1 nM), but increased in oral pathologies manifested in xerostomia (age, 57-70; n = 6; 0.8 +/- 0.2 nM) and Paget's disease (age, 58-76; n = 8; 2.0 +/- 0.6 nM). Immunohistochemical localization of TGF alpha in the salivary glands of rats and mice revealed specific immunostaining of the granular ductal cells of the parotid and submandibular glands. Reverse transcription followed by polymerase chain reaction amplification of total RNA from the parotid and submandibular glands of rats and mice demonstrated the presence of TGF alpha mRNA, suggesting endogenous synthesis by the salivary glands. Thus, salivary glands appear to be an exocrine source for a second member of the epidermal growth factor-like growth factor family in the oral cavity.

Adult↗

PubMed1994

Human uterine tissue throughout the menstrual cycle expresses transforming growth factor-beta 1 (TGF beta 1), TGF beta 2, TGF beta 3, and TGF beta type II receptor messenger ribonucleic acid and protein and contains [125I]TGF beta 1-binding sites.

The use of isoform-specific transforming growth factor-beta (TGF beta) primers, 35S-labeled 40-mer oligonucleotide probes and polyclonal antibodies, reverse transcription-polymerase chain reaction, in situ hybridization, and immunohistochemical observations has revealed that human uterine tissue at various reproductive stages expresses TGF beta s and TGF beta type II receptor messenger RNAs (mRNAs) and proteins. The reverse transcription-polymerase chain reaction revealed the predicted 443-, 310-, 524-, and 431-basepair fragments for TGF beta 1, TGF beta 2, TGF beta 3, and TGF beta type II receptor, respectively, in both endometrial and myometrial tissues, which were further verified by restriction enzyme analysis. In situ hybridization and immunohistochemical observations indicated that all uterine cell types express TGF beta s mRNAs and proteins. In the functionalis region, endometrial luminal and glandular epithelial cells are the primary cell types expressing TGF beta s mRNAs and proteins, with lesser expression in stromal cells, whereas in the basalis region, they are equally expressed in both cell types. In myometrium, TGF beta mRNA and protein expression in smooth muscle cells occurs at a substantially lower level than in endometrial tissue. In endometrial tissue, the highest level of TGF beta mRNA and protein expression appeared in the late proliferative and early to midsecretory phases of the menstrual cycle, with a considerable reduction during the late secretory and postmenopausal periods. The pattern and cellular distribution of TGF beta type II receptor protein were similar to those seen with TGF beta isoforms in both endometrial and myometrial tissues. Quantitative autoradiography (net grain density per 100 microns 2) of specific binding of [125I]TGF beta 1 for different uterine cell types indicated that the stromal cells contain a higher grain density than other uterine cell types (P < 0.05), without a significantly different density in the proliferative, compared with the secretory, phase of the menstrual cycle. These data suggest that TGF beta s acting through their specific receptors may play an important role in a variety of uterine functions in an autocrine/paracrine manner, and ovarian steroids may also regulate their expression in endometrial tissue.

Adult↗