Biomedical subjects

C V Rao

Publications and source records attributed to C V Rao.

At least 19 recordsLinked to original sources

PubMed1995

Enhancement of experimental colon carcinogenesis by dietary 6-phenylhexyl isothiocyanate.

Naturally occurring and related synthetic isothiocyanates are known to exert chemopreventive effects in several organs in rodent models. The present study was designed to investigate the efficacy of 6-phenylhexyl isothiocyanate (PHITC), a potent chemopreventive agent in the lung tumor model in strain A mice, on azoxymethane-induced colon tumorigenesis. Another aim was to study the modulating effect of PHITC on colonic mucosal and tumor phospholipase A2 (PLA2), phosphatidylinositol-specific phospholipase C (PI-PLC), lipoxygenase (LOX), and cyclooxygenase (COX) activities. At 5 weeks of age, groups of male F344 rats were fed control diet or diets containing 320 or 640 ppm of PHITC representing 40 and 80% maximum tolerated dose levels, respectively. At 7 weeks of age, all animals except those in the vehicle-treated groups were given two weekly s.c. injections of azoxymethane at a dose rate of 15 mg/kg body weight/week. All animals continued on their respective dietary regimen for 52 weeks after the carcinogen treatment; then the study was terminated. Colonic mucosa and tumors were analyzed for PLA2, PI-PLC, prostaglandin (PG) E2, COX, and LOX activities. Intestinal tumors were evaluated histopathologically and classified as invasive or noninvasive adenocarcinomas. Intestinal tumor incidence (percentage of animals with tumors) and tumor multiplicity (tumors/animal; tumors/tumor-bearing animal) were compared among the dietary groups. At the 640-ppm dose level, dietary PHITC significantly increased the incidence of intestinal (small intestine plus colon) adenocarcinomas (P < 0.05) as well as the multiplicities of invasive and noninvasive adenocarcinomas of the colon (P < 0.05 to 0.01). At the 320-ppm dose level, PHITC increased the multiplicity (tumors/animal) of noninvasive adenocarcinomas and total (invasive plus noninvasive) adenocarcinomas of the colon (P < 0.05). Dietary PHITC also increased the colon tumor volume (2- to 4.3-fold) in a dose-dependent manner. Moreover, PHITC significantly enhanced the activities of PLA2 (50-100%) and levels of PGE2 (2-fold) in the colonic mucosa and in tumors, but it had no significant effect (P > 0.05) on PI-PLC activity. The formation of COX metabolites, particularly PGE2, PGF2 alpha, PGD2, 6-keto PGF1 alpha, and thromboxane B2, as well as LOX metabolites such as 8(S)-, 12(S)- and 15 (S)-hydroxyeicosatetraenoic acids, were significantly increased in the colonic mucosa and tumors of animals that were fed 640 ppm of PHITC. Although the exact mechanism by which PHITC promotes colon tumorigenesis remains to be elucidated, it is likely that the tumor-promoting effects of PHITC may, at least in part, be related to increased eicosanoid metabolism in the colon.

Animals↗

PubMed1995

Chemoprevention of colon carcinogenesis by phenylethyl-3-methylcaffeate.

Previous studies from this laboratory have established that caffeic acid esters present in propolis, a natural resin produced by honey bees, are potent inhibitors of human colon adenocarcinoma cell growth, carcinogen-induced biochemical changes, and preneoplastic lesions in the rat colon. The present study was designed to investigate the chemopreventive action of dietary phenylethyl-3-methylcaffeate (PEMC) on azoxymethane-induced colon carcinogenesis and to examine the modulating effect of PEMC on phosphatidylinositol-specific phospholipase C (PI-PLC), phospholipase A2, lipoxygenase (LOX), and cyclooxygenase activities in the colonic mucosa and tumor tissues in male F344 rats. At 5 weeks of age, groups of rats were fed the control (modified AIN-76A) diet, or a diet containing 750 ppm of PEMC. At 7 weeks of age, all animals except those in the vehicle (normal saline)-treated groups were given 2 weekly s.c. injections of azoxymethane at a dose rate of 15 mg/kg body weight/week. All groups were maintained on their respective dietary regimen until the termination of the experiment 52 weeks after the carcinogen treatment. Colonic tumors were evaluated histopathologically. Both colonic mucosa and tumors were analyzed for PI-PLC, phospholipase A2, cyclooxygenase, and LOX activities. The results indicate that dietary administration of PEMC significantly inhibited the incidence and multiplicity of invasive, noninvasive, and total (invasive plus noninvasive) adenocarcinomas of the colon (P < 0.05-0.004). Dietary PEMC also suppressed the colon tumor volume by 43% compared to the control diet. Animals fed the PEMC diet showed significantly decreased activities of colonic mucosal and tumor PI-PLC (about 50%), but PEMC diet had no effect on phospholipase A2. The production of 5(S)-, 8(S)-, 12(S)-, and 15(S)-hydroxyeicosatetraenoic acids via the LOX pathway from arachidonic acid was reduced in colonic mucosa and tumors (30-60%) of animals fed the PEMC diet as compared to control diet. PEMC had no effect on the formation of colonic mucosal cyclooxygenase metabolites but inhibited the formation in colonic tumors by 15-30%. The precise mechanism by which PEMC inhibits colon tumorigenesis remains to be elucidated. It is likely that the chemopreventive action may be related, at least in part, to the modulation of PI-PLC-dependent signal transduction and LOX-mediated arachidonic acid metabolism.

Animals↗

PubMed1995

Novel expression of luteinizing hormone/chorionic gonadotropin receptor gene in rat prostates.

The reverse transcription-polymerase chain reaction (RT-PCR) amplified an expected 255 bp luteinizing hormone/chorionic gonadotropin (LH/CG) receptor sequence from rat prostates. Northern blotting demonstrated that prostates contain 4.3, 3.3, 2.6, 1.8, 0.8 and 0.2 kb LH/CG receptor transcripts. Western immunoblotting and ligand blotting demonstrated that prostates also contain an 80 kDa receptor protein which can bind 125I-labeled hCG and this binding was inhibited by excess unlabeled hCG. In situ hybridization and immunocytochemistry revealed that while the receptors are most abundant in epithelial cells, they are scarcely found in the stroma. The ventral lobe contained more receptors than the lateral lobe and the receptors in peripheral acini of the ventral lobe are higher which progressively decreased towards central acini. In summary, prostate glands express LH/CG receptor gene. The cellular, topographical and lobular distribution of receptors suggest that LH may directly regulate prostate functions.

Animals↗

PubMed1995

Lymphocytes from pregnant women express human chorionic gonadotropin/luteinizing hormone receptor gene.

Previous studies have indicated that human chorionic gonadotropin (hCG) has immunoregulatory properties and alters the functions of human lymphocytes. However, it has not been determined whether the gene encoding the receptors for hCG/luteinizing hormone (LH) is expressed in human lymphocytes. Total peripheral mononuclear lymphocytes isolated from blood samples of pregnant women by Ficoll-Paque gradient centrifugation contained mRNA transcripts encoding the hCG/LH receptors and a 50 kDa receptor protein which can bind 125I-hCG. T-Lymphocytes isolated from total mononuclear cell fractions also contained these receptor mRNA transcripts as well as the receptor protein. The levels of receptor transcripts and receptor protein are lower in lymphocytes than in ovarian tissue. These findings suggest that the immunoregulatory actions of hCG are probably mediated by specific receptors in T-lymphocytes from pregnant women.

Blotting, Western↗

PubMed1995

Signaling and transacting factors in the transcriptional inhibition of gonadotropin releasing hormone gene by human chorionic gonadotropin in immortalized hypothalamic GT1-7 neurons.

We recently demonstrated that immortalized GT1-7 neurons co-express luteinizing hormone (LH)/human chorionic gonadotropin (hCG) receptor and gonadotropin releasing hormone (GnRH) genes. Treatment of GT1-7 neurons with LH/hCG resulted in a transcriptional inhibition of GnRH gene. In the present study, we investigated the signaling and transacting factors involved in the action of hCG. Eight-bromo-cyclic AMP can mimic the down-regulating action of hCG on GnRH mRNA levels. H-89, a protein kinase (PK) A inhibitor, but not bisindolylmaleimide, a PKC inhibitor, blocked the down- regulating actions of hCG as well as of 8-bromocyclic AMP. Treatment with the PKA inhibitor alone modestly decreased GnRH mRNA levels suggesting that PKA signaling also controls the basal expression of the GnRH gene. The direct measurement of PK activities revealed that hCG treatment of GT1-7 neurons increased the PKA but not the PKC activity. New protein synthesis is required for the down-regulating action of hCG on GnRH mRNA levels. Since some of the new proteins could be nuclear transcription or transacting factors, we investigated the effects of hCG on cyclic AMP response element binding protein (CREB), c-Fos and c-Jun protein levels. Treatment of GT1-7 neurons with hCG resulted in an increase of 43 kDa phosphorylated CREB, 50 kDa c-Fos and 40 kDa c-Jun proteins compared to the corresponding controls. The kinetics of increases were different and in all cases the increases of the proteins preceded the decrease of GnRH mRNA levels. In summary, PKA signaling and transacting factors such as CREB, Fos and Jun are probably involved in transcriptional inhibition of GnRH gene by hCG in GT1-7 neurons.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1995

Chemoprevention of colon carcinogenesis by sulindac, a nonsteroidal anti-inflammatory agent.

Epidemiological and laboratory animal model studies have suggested that nonsteroidal anti-inflammatory drugs reduce the risk of development of colon cancer. The present study was designed to investigate the chemopreventive action of 160 and 320 ppm (equivalent to 40 and 80% maximum tolerated doses) sulindac, a nonsteroidal anti-inflammatory drug, fed during initiation and postinitiation stages and 320 ppm sulindac fed during promotion/progression stages of azoxymethane-induced colon carcinogenesis in male F344 rats. Also investigated was the modulating effect of this agent on the colonic mucosal and tumor phospholipase A2, phosphatidylinositol-specific phospholipase C, lipoxygenase, and cyclooxygenase activities. At 5 weeks of age, groups of male F344 rats were fed control diet or diets containing 160 and 320 ppm of sulindac. At 7 weeks of age, all animals except those in the vehicle-treated groups were given two weekly s.c. injections of azoxymethane at a dose rate of 15 mg/kg body weight/week. Animals intended for tumor promotion/progression study were administered 320 ppm of sulindac in diet starting at 14 weeks after a second azoxymethane treatment. All animals continued on their respective dietary regimen until the termination of the experiment at 52 weeks after the carcinogen treatment. Colonic tumors were evaluated histopathologically. Colonic mucosa and tumors were analyzed for phospholipase A2, phosphatidylinositol-specific phospholipase C, prostaglandin E2, cyclooxygenase, and lipoxygenase activities. The levels of sulindac and its metabolites in stomach, cecal, and fecal contents and in serum were analyzed. The results indicate that dietary sulindac at 160 and 320 ppm levels inhibited the incidence of invasive and noninvasive adenocarcinomas of the colon (P < 0.01-0.001) as well as their multiplicity (P < 0.01-0.0001) in a dose-dependent manner. Also, feeding sulindac during promotion/progression stages significantly suppressed the incidence (P < 0.0001) and multiplicity (P < 0.0001) of colonic adenocarcinomas. Dietary sulindac also suppressed the colon tumor volume by > 52-62% compared to the control diet. Dietary sulindac significantly decreased the activities of phosphatidylinositol-specific phospholipase C (32-51%) and levels of prostaglandin E2 (> 40%) in the colonic mucosa and tumors, but it had no significant (P > 0.05) effect on phospholipase A2 activity. The formation of cyclooxygenase metabolites, particularly prostaglandin E2, prostaglandin F2 alpha, prostaglandin D2, 6-ketoprostaglandin F1 alpha, and thromboxane B2, and lipoxygenase metabolites such as 8(S)- and 12(S)-hydroxyeicosatetraenoic acids were significantly reduced in colonic mucosa and tumors of animals fed 320 ppm sulindac.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenocarcinoma↗

PubMed1995

Chemoprevention of colon carcinogenesis by dietary curcumin, a naturally occurring plant phenolic compound.

Human epidemiological and laboratory animal model studies have suggested that nonsteroidal antiinflammatory drugs reduce the risk of development of colon cancer and that the inhibition of colon carcinogenesis is mediated through the alteration in cyclooxygenase metabolism of arachidonic acid. Curcumin, which is a naturally occurring compound, is present in turmeric, possesses both antiinflammatory and antioxidant properties, and has been tested for its chemopreventive properties in skin and forestomach carcinogenesis. The present study was designed to investigate the chemopreventive action of dietary curcumin on azoxymethane-induced colon carcinogenesis and also the modulating effect of this agent on the colonic mucosal and tumor phospholipase A2, phospholipase C gamma 1, lipoxygenase, and cyclooxygenase activities in male F344 rats. At 5 weeks of age, groups of animals were fed the control (modified AIN-76A) diet or a diet containing 2000 ppm of curcumin. At 7 weeks of age, all animals, except those in the vehicle (normal saline)-treated groups, were given two weekly s.c. injections of azoxymethane at a dose rate of 15 mg/kg body weight. All groups were continued on their respective dietary regimen until the termination of the experiment at 52 weeks after the carcinogen treatment. Colonic tumors were evaluated histopathologically. Colonic mucosa and tumors were analyzed for phospholipase A2, phospholipase C gamma 1, ex vivo prostaglandin (PG) E2, cyclooxygenase, and lipoxygenase activities. The results indicate that dietary administration of curcumin significantly inhibited incidence of colon adenocarcinomas (P < 0.004) and the multiplicity of invasive (P < 0.015), noninvasive (P < 0.01), and total (invasive plus noninvasive) adenocarcinomas (P < 0.001). Dietary curcumin also significantly suppressed the colon tumor volume by > 57% compared to the control diet. Animals fed the curcumin diet showed decreased activities of colonic mucosal and tumor phospholipase A2 (50%) and phospholipase C gamma 1 (40%) and levels of PGE2 (> 38%). The formation of prostaglandins such as PGE2, PGF2 alpha, PGD2, 6-keto PGF1 alpha, and thromboxane B2 through the cyclooxygenase system and production of 5(S)-, 8(S)-, 12(S)-, and 15(S)-hydroxyeicosatetraenoic acids via the lipoxygenase pathway from arachidonic acid were reduced in colonic mucosa and tumors of animals fed the curcumin diet as compared to control diet. Although the precise mechanism by which curcumin inhibits colon tumorigenesis remains to be elucidated, it is likely that the chemopreventive action, at least in part, may be related to the modulation of arachidonic acid metabolism.

Animals↗

PubMed1995

Peritoneal papillary serous carcinoma arising in an infertile woman during ovulation-induction therapy: immunohistochemical expression of LH/hCG receptors.

A case of peritoneal papillary serous carcinoma arising in an infertile patient during ovulation-induction therapy is presented. A 34-year-old woman with past history of ovulation-induction therapy for infertility again received fertility drugs. During the use of gonadotropic hormones, massive ascites developed and an exploratory laparotomy revealed serous papillary carcinoma on the surface of bilateral ovaries, pelvic peritoneum, and omentum. Immunohistochemical analysis showed the tumor cells to be positive for luteinizing hormone/human chorionic gonadotropin receptors and negative for sex steroid receptors. A possible relationship between the use of gonadotropic hormones and cancer development is discussed.

Adult↗

PubMed1995

Peripheral and intracerebroventricular administration of human chorionic gonadotropin alters several hippocampus-associated behaviors in cycling female rats.

Our recent demonstration of receptors for luteinizing hormone (LH)/human chorionic gonadotropin (hCG) in rat brain with the highest density within the hippocampus and dentate gyrus suggests novel functional roles for gonadotropic hormones within the brain. The present study investigated whether 125I-hCG can cross the blood-brain barrier and reach hippocampus and the possible role of hCG in the regulation of several behavioral activities associated with the hippocampal formation in the rat. About 1/100th of peripherally injected 125I-hCG crossed the blood-brain barrier in an intact form and was found in cerebrospinal fluid and in hippocampus. Intraperitoneal (IP) or intracerebroventricular (ICV) injections of highly purified hCG on the morning of proestrus of cycling female rats resulted in changes in several hippocampus-associated behaviors. hCG-treated animals were generally less active and showed less exploratory behavior as compared to saline-injected control animals. There was no difference, however, in latency to enter the open field between hCG-treated and control animals. Taste neophobia was dramatically decreased following IP as well as ICV injection of hCG. No differences were found in the memory component of T-maze performance; however, the hCG-treated rats exhibited decreased stereotypic behavior. In summary, hCG can cross the blood-brain barrier, and peripheral or central administration of hCG affects several hippocampus-associated behaviors suggesting that hippocampal LH/hCG receptors are most likely involved in mediating these effects. Some of the observed behavioral changes have parallels in pregnant women.

Animals↗

PubMed1995

Regional chemoprevention of carcinogen-induced tumors in rat colon.

BACKGROUND & AIMS: Rat colon neoplasms are distributed 60% in the distal colon (DC) and 40% in the proximal colon (PC), similar to distribution of colon cancers in the industrialized world. The effects of chemopreventive agents that affect colon tumor incidence on the distribution of colon tumors were studied. METHODS: Colon tumor distribution, numbers, and volumes were measured in the DC and PC of rats administered azoxymethane (15 mg/kg subcutaneously 2x) as an initiating agent and fed diets containing various chemopreventive agents. RESULTS: In control rats, azoxymethane-induced tumor incidence in the DC exceeded that in the PC, but tumor volume was greater in the PC than the DC. Ellagic acid showed no chemopreventive effect and maintained the PC-DC colon tumor gradient. Oltipraz, a modestly effective chemopreventive agent, principally reduced the incidence of DC tumors. DL-d-difluoromethylornithine also greatly altered tumor number in the DC compared with the PC. In contrast, piroxicam (400 ppm) reduced PC tumors by 82% but DC tumors only by 57%. With all regimens, tumor volume remained greater in the PC than the DC. CONCLUSIONS: Chemopreventive agents have a selective regional effect on colon tumorigenesis in the rat. Elucidation of the mechanism for these differences may help clarify the modes of action of chemopreventive agents in colon cancer.

Animals↗

PubMed1995

Myometrial estradiol and progesterone receptor changes in preterm and term pregnancies.

OBJECTIVE: To determine if labor is associated with changes in myometrial estradiol (E2) and progesterone receptors. METHODS: Lower myometrial segments were obtained from women undergoing cesarean deliveries at term in labor (n = 10), term not in labor (n = 10), preterm in labor (n = 9), and preterm not in labor (n = 11). Western immunoblotting was used to determine the presence and molecular size of E2 and progesterone receptor proteins. Immunocytochemistry was used to determine E2 and progesterone receptor changes in preterm and term pregnancies. RESULTS: Myometrium from pregnant women contained 74-kilodalton (kDa) E2 receptor and 94- and 110-kDa progesterone receptor proteins. These receptors are present in both myometrial smooth muscle and myometrial blood vessels. The nuclei of myometrial smooth muscle cells primarily contain both receptors. The immunostaining for progesterone receptors was less in patients in labor compared with those not in labor in preterm and term pregnancies. In comparing patients not in labor, the immunostaining for progesterone receptors was less at term compared with preterm pregnancy. Unlike the differences in progesterone receptors, there are no obvious differences in E2 receptor immunostaining in myometrial samples from all four groups of women. CONCLUSION: A myometrial decrease in progesterone receptors, rather than an increase in E2 receptors, may play a role in the onset of labor in women with term or preterm pregnancies.

Adult↗

PubMed1995

Administration of subtoxic doses of t-butyl alcohol and trichloroacetic acid to male Wistar rats to study the interactive toxicity.

Tertiary butyl alcohol and trichloroacetic acid (TCA) are known to be contaminants in drinking water. In order to evaluate the interactive toxicity of t-butyl alcohol (TBA) with TCA, young male Wistar rats were dosed through water at a dose level of TBA (0.5% v/v), 25 ppm TCA and a combined dose of TBA+TCA (0.5% v/v TBA, 25 ppm TCA) for a period of 10 weeks ad libitum and were maintained on normal diet. The control animals received plain water and normal diet. There was remarkable loss of body weight and significantly decreased liver triglycerides in the treatment groups in the order of TBA+TCA, TCA, TBA and increased liver weights were observed. Serum succinate dehydrogenase (SDH) levels were significantly increased in TCA- and TBA+TCA-treated groups. There was no significant change in serum alanine (GPT), aspartate (GOT) aminotransferase, serum alkaline (ALP) and acid (ACP) phosphatase levels as well as liver glutathione (GSH) and liver and serum cholesterol levels in the treated groups. But serum triglycerides, liver glycogen, serum glucose (only in TBA- and TCA-treated animals) were significantly high in the treated groups. Lipid peroxidation measured by diene conjugation was significant in TBA+TCA-treated group and kidney GSH levels were significantly low in the treated groups. These results show that interaction of TBA+TCA does bring about alteration in biochemical parameters which may play a pivotal role in toxic responses on long-term exposure.

Alanine Transaminase↗

PubMed1995

Decreased expression of functional human chorionic gonadotropin/luteinizing hormone receptor gene in human uterine leiomyomas.

Human myometrium contains functional hCG/LH receptors. The purpose of this study was to determine whether or not leiomyomas that arise from myometrium also contain these receptors. Northern blotting demonstrated that leiomyomas and normal adjacent myometria contained hCG/LH receptor mRNA transcripts. Western immunoblotting showed that leiomyomas and corresponding normal myometria also contained 60- and 50-kDa receptor proteins. Ligand blotting revealed that only the 50-kDa receptor protein in leiomyomas and corresponding normal myometria could bind 125I-hCG and that this binding inhibited by excess unlabeled hCG. In situ hybridization and immunocytochemistry revealed that smooth muscle cells in leiomyomas and corresponding normal myometria contained the hCG/LH receptor mRNA transcripts and receptor proteins. The receptor levels were lower in leiomyomas than in corresponding normal myometria. However, the receptors were functional, as treatment with hCG resulted in a decrease of connexin-43 protein levels in leiomyomas as in normal myometria. In summary, human uterine leiomyomas express a functional hCG/LH receptor gene at a reduced level compared with that for normal corresponding myometria. This finding could be relevant to an understanding of the growth control mechanisms in leiomyomas and the manner in which medical therapy for leiomyomas might work.

Adult↗

PubMed1995

Missing anti-proliferative effect of fish oil on rectal epithelium in healthy volunteers consuming a high-fat diet: potential role of the n-3:n-6 fatty acid ratio.

Several studies have indicated dietary fish oil (FO) as a protective agent in colon carcinogenesis. Rectal cell proliferation as an intermediate biomarker of cancer risk was shown to be reduced by dietary FO in patients with adenomatous polyps and healthy subjects consuming a low-fat diet. Because the synthesis of prostaglandins (PG) which seem to be involved in this process is dependent on the ratio of n-3:n-6 fatty acids in the diet, the present study was designed to investigate whether this FO effect is also detectable in volunteers eating a high-fat diet (50% of energy) with a low n-3:n-6 ratio of 0.25. Twelve healthy volunteers received in addition to a controlled basal diet either FO (4.4 g n-3 fatty acids/day) or corn oil supplements (double-blind, crossover) for two 4-week periods. No significant differences between the two study periods were found for rectal cell proliferation as assessed by bromodeoxyuridine immunohistochemistry and ornithine decarboxylase activity, as well as for mucosal PGE2 release and mucosal membrane fatty acid composition. The results emphasize the importance of dietary n-3:n-6 ratio in determining the effects of FO on rectal cell proliferation.

Administration, Oral↗

PubMed1995

Functional nuclear epidermal growth factor receptors in human choriocarcinoma JEG-3 cells and normal human placenta.

Immunocytochemistry with a monoclonal antiepidermal growth factor (anti-EGF) receptor antibody directed against the extracellular domain which can inhibit ligand binding to the receptors showed that nuclei of choriocarcinoma JEG-3 cells and normal placental trophoblasts were distinctly immunostained for EGF receptors. This finding led us to investigate the structure and function of nuclear EGF receptors. Western immunoblotting revealed that cell membranes, isolated intact pure nuclei, and nuclear membranes contain a 170-kilodalton EGF receptor protein. Covalent receptor cross-linking demonstrated that the 170-kilodalton receptor protein in nuclei and nuclear membranes can bind [125I]EGF just as in cell membranes, and that this binding is inhibited by excess unlabeled EGF. As in cell membranes, the addition of EGF resulted in an increased receptor autophosphorylation in the nuclei and nuclear membranes. In addition, the activated receptor kinase stimulated, and in some cases inhibited, tyrosine phosphorylation of a number of lower molecular size proteins, especially in nuclei and nuclear membranes. Although the identity of these proteins is not known, none of them could bind [125I]EGF. The addition of EGF to isolated nuclei resulted in a time-dependent specific transcriptional inhibition of hCG/LH receptor gene. In summary, our data demonstrating the presence of functional nuclear EGF receptors are novel, potentially important, and go against the traditional concepts of growth factors action. The nuclear receptors have the capacity to transduce signals from EGF and may mediate intracrine and paracrine actions of EGF in the regulation of trophoblast functions.

Blotting, Western↗

PubMed1995

Expression of epidermal growth factor, transforming growth factor-alpha, and their common receptor genes in human umbilical cords.

Epidermal growth factor (EGF) and transforming growth factor-alpha (TGF alpha), present in amniotic fluid and/or in fetal blood, could potentially regulate cord functions. The present study investigated the possible presence of functional receptors and EGF and TGF alpha themselves in umbilical cord. The reverse transcription-polymerase chain reaction followed by Southern blotting demonstrated that human umbilical cords contain EGF, TGF alpha, and EGF/TGF alpha messenger ribonucleic acid (mRNA) transcripts. In situ hybridization revealed that these mRNA transcripts are present in vascular endothelial cells and smooth muscle, amnion, and myofibroblasts in Wharton's jelly. Western immunoblotting showed that the cords contain a 170-kilodalton EGF/TGF alpha receptor protein. Immunocytochemistry demonstrated that all of the cells that contained mRNA transcripts also contained corresponding proteins. Umbilical amnion contains more EGF, TGF alpha, and their receptors than any other part of the cord. In the cord, the fetal and middle portions contain more than the placental portion or the vessels inside the placental tissue. The cord receptors can bind [125I]EGF, stimulate receptor autophosphorylation, and increase cyclooxygenase-1 and -2 and prostaglandin E2, suggesting that the receptors are functional. In summary, our study demonstrates that human umbilical cord expresses EGF, TGF alpha, and their common receptor genes. The cord EGF/TGF alpha receptors are functional in terms of binding of EGF, activation of receptor autophosphorylation, and increasing the formation of vasoconstrictive eicosanoid. Thus, EGF, TGF alpha, and their receptors could be one of the determinants of human fetal growth and development by autocrine, paracrine, and endocrine actions in umbilical cord.

Dinoprostone↗

PubMed1995

Consequences of antisense human chorionic gonadotrophin-alpha subunit cDNA expression in human choriocarcinoma JAR cells.

The biosynthesis of human chorionic gonadotrophin (hCG) is a hallmark endocrine function of human choriocarcinoma cells. The present study investigated the consequences of greatly diminishing this synthesis in JAR cells by stably transfecting them with pRSV-antisense hCG-alpha cDNA expression vector. The vector directs the synthesis of antisense hCG-alpha subunit mRNA which would then bind to sense hCG-alpha subunit mRNA, thus blocking its translation and consequently dimer hCG protein synthesis. The transfection with pRSV-antisense hCG-alpha cDNA resulted in a dramatic decrease in hCG secretion as compared with untransfected parental cells or those transfected with an empty vector used for the selection of clones. The decreased secretion was due to a decreased synthesis which in turn was due to a fall in steady-state hCG-alpha and -beta subunit mRNA levels. The decrease of hCG-beta subunit transcripts was unexpected and it was not due to contamination of antisense hCG-alpha cDNA construct with hCG-beta sequence. The transcription of hCG-alpha and -beta subunit genes was not altered in transfected cells suggesting that increased degradation was responsible for decreased steady-state hCG subunit mRNA levels. Despite the decreased hCG levels, the transfected cells maintained normal hCG receptor levels, responded to epidermal growth factor stimulation of hCG synthesis and secretion and grew at the same rate as the control parental cells and those transfected with an empty vector.

Biomarkers, Tumor↗

PubMed1995

Analysis of epidermal growth factor action in human myometrial smooth muscle cells.

The present study investigated the mechanisms involved in the mitogenic action of epidermal growth factor (EGF) in cultured human myometrial smooth muscle cells. The cells contained EGF/transforming growth factor-alpha (TGF-alpha) receptors as well as EGF and TGF-alpha mRNA transcripts and the corresponding proteins. Culturing with human EGF resulted in concentration- and time-dependent increases in cell density. The maximal increase was seen at 1 nM followed by a decrease to control levels at 100 nM EGF. The EGF increased cell density from 4 to 8 days followed by a plateau coinciding with the cells reaching confluence. EGF treatment concomitantly decreased the average size of cells. TGF-alpha mimicked EGF and there was no synergism between the two, suggesting a common mechanism of action. Although the presence of 10% fetal bovine serum enhanced overall cell growth, it was not required for EGF and TGF-alpha action. The receptor antibody, which is directed against the extracellular domain and can inhibit ligand binding to the receptors, dramatically inhibited the basal cell growth and exogenous EGF reversed the antibody effect. While TGF-alpha antibody was only marginally effective, EGF antibody had no effect on basal cell growth. Lavendustin (a tyrosine kinase inhibitor), calphostin (a protein kinase C inhibitor), but not H-89 (a protein kinase A inhibitor), inhibited EGF action. Indomethacin, a cyclo-oxygenase inhibitor, completely inhibited, whereas nordihydroguaiaretic acid, a lipoxygenase inhibitor, slightly inhibited EGF action. While estradiol-17 beta modestly inhibited basal as well as EGF-stimulated myometrial smooth muscle cell density, progesterone had no effect. In summary, mitogenic action of EGF in human myometrial smooth muscle cells does not require serum components and it involves tyrosine kinase and protein kinase C signaling and eicosanoids from the cyclooxygenase pathway of arachidonic acid metabolism.

Antibodies, Monoclonal↗