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1.
BJOG ; 118(1): 17-23, 2011 Jan.
Article in English | MEDLINE | ID: mdl-21091926

ABSTRACT

OBJECTIVE: To test the feasibility and efficacy of an approach that foregoes the routine use of ultrasound for the determination of eligibility for medical termination of pregnancy. DESIGN: Prospective trial. SETTING: Ten termination of pregnancy clinics in the USA. POPULATION: A total of 4484 women seeking termination of pregnancy with mifepristone-misoprostol. METHODS: Women provided estimates of the date of their last menstrual period and underwent pelvic bimanual and ultrasound examinations. We compared estimates of gestational age using these three methods. MAIN OUTCOME MEASURE: Proportion of women of ≤9 weeks' gestation by woman or provider estimate, but >9 weeks' gestation by ultrasound. RESULTS: The reliance on women's report of their last menstrual period together with physical examination to determine their eligibility for termination of pregnancy with mifepristone-misoprostol would result in few women (63/4008 or 1.6%) accepted for treatment outside the current limits of standard mifepristone-misoprostol regimens used for early termination of pregnancy (i.e. ≤63 days' gestation on ultrasound). CONCLUSIONS: Last menstrual period and physical examination alone, without the routine use of ultrasound, are highly effective for the determination of women's eligibility for early termination of pregnancy with mifepristone-misoprostol.


Subject(s)
Abortifacient Agents, Nonsteroidal/administration & dosage , Abortion, Induced/methods , Mifepristone/administration & dosage , Misoprostol/administration & dosage , Drug Therapy, Combination , Feasibility Studies , Female , Gestational Age , Humans , Physical Examination , Pregnancy , Prospective Studies , Self Administration , Ultrasonography, Prenatal
2.
J Endocrinol ; 171(1): 109-18, 2001 Oct.
Article in English | MEDLINE | ID: mdl-11572795

ABSTRACT

Prostate cancer is the most diagnosed invasive malignancy in males. Androgens and oestrogens have been implicated in the pathogenesis of prostate cancer. We report herein that the pure anti-oestrogen ICI 182,780 (ICI) reduces Ki-67 labelling index and IGF-I receptor levels in rat prostate. Increase of IGF-I mRNA and IGF-binding protein 3 (IGFBP-3) accumulation occur without any effect on prostate weight. Finasteride significantly decreases prostate weight and inhibits IGF-I gene expression. IGFBP-3 mRNA, Akt and phospho-Akt are not affected by finasteride. Co-administration of ICI plus finasteride reduces prostate weight by approximately 50% and causes acinar dilation with decreased luminal epithelial cell thickness. The acinar epithelial cells became atrophic and inactive with minimal cytoplasm. We also demonstrate a synergistic effect of ICI and finasteride on induction of IGFBP-3 accumulation and inhibition of Akt phosphorylation. Because the IGF and IGFBP-3 system plays an important role in prostate epithelial cell proliferation, apoptosis and tumour progression, the inhibitory effects of finasteride and ICI on IGF system may contribute to their anti-proliferative activity. These observations support a potential use of ICI in conjunction with finasteride in the prevention and/or treatment of prostate cancer.


Subject(s)
5-alpha Reductase Inhibitors , Estradiol/analogs & derivatives , Estradiol/pharmacology , Estrogen Antagonists/pharmacology , Finasteride/pharmacology , Insulin-Like Growth Factor I/metabolism , Prostate/metabolism , Protein Serine-Threonine Kinases , Animals , Blotting, Northern/methods , Cell Division/drug effects , Drug Synergism , Enzyme Inhibitors/pharmacology , Epithelial Cells/drug effects , Fulvestrant , Insulin-Like Growth Factor Binding Protein 3/metabolism , Male , Organ Size/drug effects , Phosphorylation , Prostate/anatomy & histology , Prostate/drug effects , Proto-Oncogene Proteins/metabolism , Proto-Oncogene Proteins c-akt , Rats , Receptor, IGF Type 1/metabolism , Statistics, Nonparametric
3.
Can J Physiol Pharmacol ; 79(6): 533-44, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11430591

ABSTRACT

Doxorubicin (DOX) and VP16 are DNA topoisomerase II inhibitors yet only DOX induces an irreversible cardiotoxicity, likely through DOX-induced oxidative stress. Egr-1 is overexpressed after many stimuli that increase oxidative stress in vitro and after DOX-injection into adult mice in vivo. To investigate Egr-1 function in the heart, we compared the molecular and histological responses of wild type (+/+) and Egr-1 deficient (-/-) female mice to saline, DOX, VP16, the cardioprotectant dexrazoxane (DZR), or DOX+DZR injection. DOX, and to a lesser extent VP16, induced characteristic increases in cardiac muscle and non-muscle genes typical of cardiac damage in +/+ mice, whereas only beta-MHC and Sp1 were increased in -/- mice. DZR-alone treated +/+ mice showed increased cardiomyocyte transnuclear width without a change to the heart to body weight (HW/BW) ratio. However, DZR-alone treated -/- mice had an increased HW/BW, increased cardiomyocyte transnuclear width, and gene expression changes similar to DOX-injected +/+ mice. DZR pre-injection alleviated DOX-induced gene changes in +/+ mice; in DZR+DOX injected -/- mice the increases in cardiac and non-muscle gene expression were equal to, or exceeded that, detected after DOX-alone or DZR-alone injections. We conclude that Egr-1 is required for DOX-induced molecular changes and for DZR-mediated cardioprotection.


Subject(s)
Antibiotics, Antineoplastic/pharmacology , Cardiovascular Agents/pharmacology , DNA-Binding Proteins/deficiency , DNA-Binding Proteins/genetics , Doxorubicin/pharmacology , Immediate-Early Proteins , Razoxane/pharmacology , Transcription Factors/deficiency , Transcription Factors/genetics , Animals , DNA/biosynthesis , DNA/genetics , Early Growth Response Protein 1 , Etoposide/pharmacology , Female , Gene Expression Regulation/drug effects , Mice , Phenotype , RNA, Messenger/biosynthesis , Reverse Transcriptase Polymerase Chain Reaction
4.
Endocrinology ; 142(6): 2409-18, 2001 Jun.
Article in English | MEDLINE | ID: mdl-11356689

ABSTRACT

Estrogens play a role in mammary gland function and are implicated in mammary carcinogenesis. We report the cloning of a novel gene [steroid-sensitive gene 1 (SSG1)] that is regulated by E(2) in the rat uterus and mammary gland. The full-length SSG1 complementary DNA has an open reading frame of 1158 nucleotides encoding a putative protein of 385 amino acids. A SSG1-specific antibody recognizes a 40-kDa protein localized to myoepithelial cells of normal mammary tissue and to endothelial cells of 7,12-dimethylbenz(a)antracene-induced mammary tumors. Treatment of rats with E(2) at 1.2 or 2.4 microg/kg.day for 21 days increases SSG1 protein levels in mammary tissue by 16-fold compared with controls. Removal of E(2) after a 14-day treatment decreases SSG1 protein levels 6-fold and 3-fold at 120 and 144 h, respectively. Treatment of rats with the estrogen antagonists tamoxifen or ICI 182,780 did not affect SSG1 protein levels compared with controls. SSG1 protein levels in 7,12-dimethylbenz(a)antracene-induced rat mammary tumors were 23-fold greater than SSG1 levels in resting mammary tissue, and 8-fold higher than protein levels expressed in lactating mammary glands. We propose that SSG1 plays a role in estrogen functions, and its overexpression is correlated with mammary carcinogenesis.


Subject(s)
Cloning, Molecular , Estradiol/pharmacology , Gene Expression Regulation/drug effects , Mammary Neoplasms, Experimental/genetics , Neoplasm Proteins/genetics , 9,10-Dimethyl-1,2-benzanthracene , Amino Acid Sequence , Animals , Base Sequence , Carcinogens , Endothelium, Vascular/chemistry , Female , Fluorescent Antibody Technique , Mammary Glands, Animal/blood supply , Mammary Glands, Animal/chemistry , Mammary Glands, Animal/drug effects , Mammary Neoplasms, Experimental/blood supply , Mammary Neoplasms, Experimental/chemically induced , Molecular Sequence Data , Neoplasm Proteins/analysis , Neoplasm Proteins/chemistry , Ovariectomy , RNA, Messenger/analysis , Rats , Rats, Sprague-Dawley , Tumor Suppressor Proteins
5.
J Biol Chem ; 276(17): 14124-32, 2001 Apr 27.
Article in English | MEDLINE | ID: mdl-11278765

ABSTRACT

Human hepatitis B virus is a risk factor for the development of hepatocellular carcinoma. The hepatitis B virus x protein (HBx) has been shown to inactivate the p53 tumor suppressor protein and impair DNA repair, cell cycle, and apoptosis mechanisms. Herein we report that HBx represses two components of the transcription-repair factor TFIIH, XPB (p89), and XPD (p80), both in p53-proficient and p53-deficient liver cells. This inhibition is observed while HBx maintains its transactivation function. Expression of HBx in liver cells results in down-regulation of endogenous XPB and XPD mRNAs and proteins; this inhibition is not observed with other TFIIH subunits, XPA or PCNA. In liver tissue from HBx transgenics, XPB and XPD proteins are down-regulated in comparison to matched normal liver tissue. HBx has been shown to interact with Sp1 transcription factor and affects its DNA binding activity. Sp1 is essential for the basal promoter activity of XPB in liver cells and Drosophila SL2 cells. In the Sp1-deficient SL2 cells, HBx-induced XPB and XPD inhibition is Sp1-dependent. In summary, our results provide evidence that HBx represses the expression of key TFIIH proteins at least in part through Sp1 elements; this repression may impair TFIIH function in DNA repair mechanisms.


Subject(s)
DNA Helicases , DNA-Binding Proteins/metabolism , Drosophila Proteins , Gene Expression Regulation, Viral , Liver/metabolism , Proteins/metabolism , TATA-Binding Protein Associated Factors , Trans-Activators/genetics , Trans-Activators/metabolism , Transcription Factor TFIID , Transcription Factors, TFII , Transcription Factors/genetics , Transcription, Genetic , Animals , Animals, Genetically Modified , Apoptosis , Blotting, Western , Cell Line , Chloramphenicol O-Acetyltransferase/metabolism , DNA Repair/genetics , Down-Regulation , Drosophila , Female , Flow Cytometry , Humans , Immunohistochemistry , Male , Mice , Mice, Transgenic , Models, Genetic , Plasmids/metabolism , Promoter Regions, Genetic , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Sp1 Transcription Factor/metabolism , Transcription Factor TFIIH , Transcriptional Activation , Transfection , Tumor Cells, Cultured , Viral Regulatory and Accessory Proteins , Xeroderma Pigmentosum Group D Protein
6.
J Mol Endocrinol ; 26(1): 1-10, 2001 Feb.
Article in English | MEDLINE | ID: mdl-11174849

ABSTRACT

Androgens play an important role in prostate gland development and function, and have been implicated in prostate carcinogenesis. We report the regulation of the gap junctional intercellular communication gene connexin 43 (Cx43) by androgens in the prostate gland. In rat ventral prostate tissue, only trace levels of Cx43 mRNA were detected. Castration, however, resulted in a high increase in Cx43 mRNA and protein. Cx32 was unchanged. Castration-induced Cx43 mRNA and protein were abolished by administration of dihydrotestosterone (DHT). Following castration, prostate weights were approximately 16% of sham-treated controls. However, DHT replacement resulted in prostate weights which were not different from sham-treated controls. Under similar castration conditions, Cx43 induction coincided with pronounced apoptosis in the prostate gland cells, and DHT prevented the induction of apoptosis. Given the physiological role of gap junctions and androgens in the regulation of prostate tissue homeostasis, our observations are relevant to the understanding of androgen-dependent prostate carcinogenesis.


Subject(s)
Connexin 43/genetics , Gene Expression Regulation , Prostate/metabolism , Animals , In Situ Hybridization , In Situ Nick-End Labeling , Male , RNA, Messenger/genetics , Rats , Rats, Sprague-Dawley
7.
Am J Physiol Heart Circ Physiol ; 278(3): H796-805, 2000 Mar.
Article in English | MEDLINE | ID: mdl-10710348

ABSTRACT

Unmanipulated early growth response-1 (Egr-1)-deficient -/- mice have similar heart-to-body weight ratios but express lower amounts of atrial natriuretic factor (ANF), beta-myosin heavy chain (beta-MHC), skeletal actin, NGF1-A binding protein (NAB)-2, Sp1, c-fos, c-jun, GATA-4, and Nkx2.5 than +/+ or +/- mice. alpha-MHC, tubulin, and NAB-1 expression was similar. Isoproterenol (Iso) and phenylephrine (PE) infusion into +/+ and -/- mice increased heart weight, ANF, beta-MHC, skeletal actin, Sp1, NAB-2, c-fos, and c-jun expression, but induction in -/- mice was lower. Only Iso + PE-treated +/+ mice showed induction of NAB-1, GATA-4, and Nkx2.5. Foci of fibrosis were found in Iso + PE-treated -/- and +/+ mice. Surprisingly, vehicle-treated -/- mice displayed fibrosis and increased Sp1, skeletal actin, Nkx2.5, and GATA-4 expression without hypertrophy. Minipump removal caused the agonist-treated hearts and gene expression to regress to control or near-control levels. Thus Egr-1 deficiency caused a blunted catecholamine-induced hypertrophy response and increased sensitivity to stress.


Subject(s)
Cardiomegaly/metabolism , DNA-Binding Proteins/deficiency , Immediate-Early Proteins , Neoplasm Proteins , Receptors, Adrenergic/physiology , Transcription Factors/deficiency , Actins/genetics , Adrenergic alpha-Agonists/pharmacology , Adrenergic beta-Agonists/pharmacology , Animals , Atrial Natriuretic Factor/genetics , Cardiomegaly/etiology , Early Growth Response Protein 1 , Gene Expression/drug effects , Genes, fos/genetics , Genes, jun/genetics , Isoproterenol/pharmacology , Male , Mice , Mice, Mutant Strains , Myosin Heavy Chains/genetics , Phenylephrine/pharmacology , Repressor Proteins/genetics , Sp1 Transcription Factor/genetics
8.
Biochem Biophys Res Commun ; 267(2): 482-7, 2000 Jan 19.
Article in English | MEDLINE | ID: mdl-10631087

ABSTRACT

Expression of the human parathyroid hormone (PTH)/PTH-related peptide receptor (PTHR) gene is controlled by three promoters, P1-P3. P1 functions specifically in kidney, whereas P2 is ubiquitously active. P3 is also widely active, although more so in kidney than other tissues. However, only P2 functions at midgestation. We examined the role of methylation in controlling PTHR promoter activity. Function of all promoters was inhibited by CpG methylation in vitro. Significantly, P1 is selectively hypomethylated in adult kidney in vivo, strongly suggesting that demethylation is required for renal P1 function. Moreover, this pattern is established by 11. 75 weeks of fetal age, several weeks prior to the onset P1 activity. P3 is unmethylated at midgestation, although it is inactive at this stage of development, and thus exhibits characteristics of both tissue-specific and ubiquitously active promoters. These results show that adult methylation patterns of P1 and P3 are established several weeks prior to their induction, indicating that their function requires factors expressed late in development.


Subject(s)
DNA Methylation , DNA/chemistry , DNA/genetics , Promoter Regions, Genetic , Receptors, Parathyroid Hormone/genetics , Adult , Animals , COS Cells , CpG Islands , Embryonic and Fetal Development/genetics , Female , Gene Expression Regulation, Developmental , Gene Transfer Techniques , Genes, Reporter , Gestational Age , Humans , In Vitro Techniques , Kidney/metabolism , Luciferases/genetics , Mice , Pregnancy , Receptor, Parathyroid Hormone, Type 1 , Restriction Mapping , Tissue Distribution
10.
Cancer Res ; 59(16): 4104-10, 1999 Aug 15.
Article in English | MEDLINE | ID: mdl-10463615

ABSTRACT

Gap junctions are intercellular channels that are formed from members of a family of proteins, the connexins (Cxs). Gap junctions play an important role in vital functions, including the regulation of cell growth and cell differentiation. Here, we examined the expression of Cx43, a major Cx in breast tissue, in 32 surgical specimens obtained from breast cancer patients who underwent a primary surgical resection prior to chemotherapy or radiotherapy treatments. The expression of Cx43 gap junctions was compared to the levels of estrogen, progesterone, and erbB2 tyrosine kinase receptors. In addition, a panel of breast cancer cell lines and a series of normal rat mammary tissues and rat mammary tumors induced in vivo by dimethylbenz(a)anthracene were studied. We demonstrated that the lack of Cx43 gap junctions is a common feature of human mammary cancer tissues compared to nonneoplastic breast tissues surrounding primary tumors. Cx43 gap junctions were not observed in ductal carcinomas in situ, infiltrating ductal carcinomas, and infiltrating lobular carcinomas, and they seem to be independent of estrogen, progesterone, and erbB2 receptor status. In breast cancer cell lines and rodent mammary carcinoma tissues, down-regulation of Cx43 occurs at the mRNA level, suggesting a transcriptional mechanism for the decrease of Cx43 protein in breast cancer. In summary, this study provides evidence of decreased expression of Cx43 gap junctions in breast cancer at various stages of progression as well as breast cancer cell lines and raises the possibility that Cx43 may be a useful marker for detecting early oncogenesis in the breast. Because Cx43 gap junctions are lacking in breast cancer and restoration of Cx43 has been shown to reverse the malignant phenotype in vitro, pharmacological up-regulation of Cx43 may prove beneficial in cancer therapeutics.


Subject(s)
Biomarkers, Tumor , Breast Neoplasms/metabolism , Connexin 43/deficiency , Mammary Neoplasms, Animal/metabolism , Adult , Aged , Aged, 80 and over , Animals , Breast Neoplasms/pathology , Down-Regulation , Female , Humans , Immunohistochemistry , Mammary Neoplasms, Animal/pathology , Middle Aged , Rats , Tumor Cells, Cultured
11.
Br J Pharmacol ; 127(5): 1165-76, 1999 Jul.
Article in English | MEDLINE | ID: mdl-10455263

ABSTRACT

Adrenoreceptor agonists induce a hypertrophic phenotype in vitro and in vivo. To investigate the molecular remodeling in chronic cardiac hypertrophy we infused adult male mice with vehicle. isoproterenol, phenylephrine or both agonists for 3, 7 or 14 days. All drugs increased cardiac mass. After minipump removal cardiac mass regressed to control levels within 7 days after PE and ISO treatment whereas ISO + PE treated hearts were incompletely regressed. ANF and beta-MHC, but not alpha-MHC, expression were increased by agonists at all time points. GATA-4, Nkx-2.5, Egr-1, c-jun and c-fos expression were increased after 3, 7 and 14 days of treatment. Expression was greatest after ISO+PE> >ISO>PE>vehicle infusion suggesting a synergistic effect of adrenoreceptor stimulation and indicating a greater effect of beta- than alpha-adrenergic action in vivo. After PE or ISO drug withdrawal the HW/BW was normal and Egr-1, c-jun, c-fos and GATA-4, but not Nkx2.5, expression dropped to control levels. HW/BW regression was incomplete after ISO+PE and elevated levels of Egr-1, c-jun and Nkx2.5 expression remained. A hydralazine-mediated reduction in blood pressure had no effect on the agonist-induced cardiac hypertrophy or gene expression. In conclusion, we found that continued agonist stimulation, and not blood pressure. is responsible for the maintained increase in gene expression. Further, we found the decrease in gene expression in the regression after drug withdrawal was gene specific.


Subject(s)
Cardiomegaly/metabolism , DNA-Binding Proteins/genetics , Gene Expression Regulation , Genes, Immediate-Early , Homeodomain Proteins/genetics , Receptors, Adrenergic/physiology , Transcription Factors/genetics , Xenopus Proteins , Animals , Antihypertensive Agents/pharmacology , Atrial Natriuretic Factor/biosynthesis , Cardiomegaly/chemically induced , GATA4 Transcription Factor , Homeobox Protein Nkx-2.5 , Isoproterenol/pharmacology , Male , Mice , Mice, Inbred C57BL , Myosin Heavy Chains/biosynthesis , Phenylephrine/pharmacology , Transcription Factors/biosynthesis
12.
Age Ageing ; 28(2): 193-7, 1999 Mar.
Article in English | MEDLINE | ID: mdl-10350418

ABSTRACT

INTRODUCTION: the age reported by or on behalf of centenarians may be suspect unless proven correct. We report the validity of age reports in a population-based sample of centenarians living in New England and the prevalence of centenarians in an area within the North Eastern USA. METHODS: cohort study. All centenarians in a population-based sample detected by local censuses. Ages were confirmed by birth certificate. Type of residence and whether the subject was living independently were also recorded. RESULTS: from a population of about 450,000 people, 289 potential centenarians were reported by the censuses of the eight towns participating in the study. Of these, 186 (64%) had died at the time centenarian prevalence was determined. Of the 80 still alive, 13 (16%) had incorrect birth years recorded by the censuses. The specificity of the censuses for stating the number of centenarians alive and living in the sample was 28-31%. Using additional sources, only four more centenarians were located, indicating that the sensitivity of the censuses approached 100%. We had an 83% success rate in obtaining proof of age in those families we interviewed. In all instances, age and birth order of children were an important source of corroborative evidence and in no case did we detect inconsistencies with the families' reported ages of the centenarian subjects. Therefore, there were at least 46 centenarians or approximately 1 centenarian per 10,000 people. CONCLUSIONS: age validation can be performed for most centenarians in the North Eastern USA. Self or family reports of those between the ages of 100 and 107 years were dependable.


Subject(s)
Aged , Censuses , Age Distribution , Aged, 80 and over , Cohort Studies , Humans , New England , Reproducibility of Results
13.
Am J Physiol ; 276(3): H803-14, 1999 03.
Article in English | MEDLINE | ID: mdl-10070062

ABSTRACT

Differential display identified that gene fragment HA220 homologous to the transcriptional activator factor II 250 (TAFII250) gene, or CCG1, was increased in hypertrophied rodent heart. To determine whether TAFII250 gene expression is modified after cardiac damage, we measured TAFII250 expression in vivo in mouse hearts after injection of the cardiotoxic agent doxorubicin (DXR) and in vitro in DXR-treated isolated rat neonatal cardiomyocytes. In vivo atrial natriuretic factor (ANF), beta-myosin heavy chain (beta-MHC), Egr-1, and TAFII250 expression increased with dose and time after a single DXR injection, but only ANF and beta-MHC expression were increased after multiple injections. After DXR treatment of neonatal cardiomyocytes we found decreased ANF, alpha-MHC, Egr-1, and TAFII250 expression. Expression of the TAFII250-regulated genes, the D-type cyclins, was increased after a single injection in adult mice and was decreased in DXR-treated cardiomyocytes. Thus expression of Erg-1, TAFII250, and the D-type cyclins is modulated after cardiotoxic damage in adult and neonatal heart.


Subject(s)
Cyclins/genetics , DNA-Binding Proteins/genetics , Doxorubicin/pharmacology , Gene Expression/drug effects , Heart/physiopathology , Immediate-Early Proteins , Nuclear Proteins/genetics , TATA-Binding Protein Associated Factors , Transcription Factor TFIID , Transcription Factors/genetics , Amino Acid Sequence/genetics , Animals , Animals, Newborn/genetics , Cardiomegaly/physiopathology , Cyclin D , DNA-Binding Proteins/physiology , Early Growth Response Protein 1 , Gene Expression/physiology , Gene Expression Regulation/physiology , Heart/drug effects , Histone Acetyltransferases , Mice , Mice, Inbred Strains , Molecular Sequence Data , Myocardium/metabolism , Myocardium/pathology , Nuclear Proteins/physiology , RNA/metabolism , Rats , Rats, Sprague-Dawley , Time Factors
14.
J Clin Invest ; 102(5): 958-67, 1998 Sep 01.
Article in English | MEDLINE | ID: mdl-9727064

ABSTRACT

The parathyroid hormone (PTH)/PTH-related peptide (PTHrP) receptor (PTHR) functions in skeletal development and mediates an array of other physiological responses modulated by PTH and PTHrP. PTHR gene transcription in mouse is controlled by two promoters: P1, which is highly and selectively active in kidney; and P2, which functions in a variety of tissues. P1 and P2 are conserved in human tissue; however, P1 activity in kidney is weak. We have now identified a third human promoter, P3, which is widely expressed and accounts for approximately 80% of renal PTHR transcripts in the adult. No P3 activity was detected in mouse kidney, indicating that renal PTHR gene expression is controlled by different signals in human and mouse. During development, only P2 is active at midgestation in many human tissues, including calvaria and long bone. This strongly suggests that factors regulating well conserved P2 control PTHR gene expression during skeletal development. Our results indicate that human PTHR gene transcription is upregulated late in development with the induction of both P1 and P3 promoter activities. In addition, P2-specific transcripts are differentially spliced in a number of human cell lines and adult tissues, but not in fetal tissues, giving rise to a shorter and less structured 5' UTR. Thus, our studies show that both human PTHR gene transcription and mRNA splicing are developmentally regulated. Moreover, our data indicate that renal and nonrenal PTHR gene expression are tightly coordinated in humans.


Subject(s)
Gene Expression Regulation, Developmental/genetics , Promoter Regions, Genetic/genetics , Receptors, Parathyroid Hormone/physiology , Up-Regulation/physiology , Bone and Bones/physiology , Cells, Cultured , Cloning, Molecular , Fetus/physiology , Genes, Reporter/genetics , Humans , Kidney/physiology , Parathyroid Hormone/physiology , RNA Splicing/genetics , RNA, Messenger/metabolism , Receptor, Parathyroid Hormone, Type 1 , Ribonucleases/metabolism , Sequence Analysis, DNA , Transfection/genetics
15.
Clin Cancer Res ; 4(1): 131-8, 1998 Jan.
Article in English | MEDLINE | ID: mdl-9516961

ABSTRACT

5-Oxo-L-prolinase (5-OPase) is an enzyme of the gamma-glutamyl cycle involved in the synthesis and metabolism of glutathione (GSH), which is known to protect cells from the cytotoxic effects of chemotherapy and radiation. Previous studies on rats have shown that administration of the cysteine prodrug L-2-oxothiazolidine-4-carboxylate, a 5-oxo-L-proline analogue that is metabolized by 5-OPase, preferentially increases the GSH content of normal tissues while paradoxically decreasing it in the tumor and results in an enhanced in vivo tumor response to the anticancer drug melphalan. These observations initiated the present study of 5-OPase in experimental models and clinical specimens to investigate the potential role of this enzyme in the selective modulation of GSH in normal and tumor tissues. First, 5-OPase activity was measured in tissues of tumor-bearing rats, in the peripheral mononuclear cells of normal human subjects, and in surgically resected tumor and the adjacent normal tissues from patients. We found that the activity of 5-OPase in human kidney, liver, and lung is significantly lower than that found in rats. Second, we have raised a polyclonal IgG anti-5-OPase antibody by immunizing rabbits with purified 5-OPase from rat kidney. This antibody has very high affinity (shown by immunoprecipitation) and specificity (shown by Western blot) and cross-reacts with human 5-OPase (shown by Western blot and immunohistochemistry). It was then used to examine the distribution of 5-OPase in paired normal and neoplastic human specimens using Western blot and immunohistochemistry. Examination of paired normal and neoplastic tissues of stomach and lung revealed a significantly lower level of 5-OPase in tumor tissues than in the paired normal tissues. In colon tissues, there is no significant difference in 5-OPase level between the normal and tumor tissues. These findings could have implications for both carcinogenesis and therapy.


Subject(s)
Glutathione/metabolism , Neoplasms/enzymology , Pyroglutamate Hydrolase/metabolism , Animals , Female , Humans , Immunohistochemistry , Pyroglutamate Hydrolase/immunology , Pyrrolidonecarboxylic Acid , Rats , Rats, Inbred F344 , Thiazoles/pharmacology , Thiazolidines
16.
17.
Lab Invest ; 76(5): 703-16, 1997 May.
Article in English | MEDLINE | ID: mdl-9166289

ABSTRACT

Human carcinoembryonic antigen (CEA) is overexpressed in a wide variety of epithelial malignancies including colon cancer. CEA can function in vitro as a homotypic intercellular adhesion molecule and can block the terminal differentiation of rodent myoblasts, thus raising the possibility that deregulated expression of CEA might directly contribute to malignant progression. To address this question, the expression pattern and cell-surface levels of CEA were studied during malignant transformation of the colonic epithelium in sporadic and familial adenomatous polyposis-related neoplasms. The level of immunohistochemically detected CEA was higher in 30% to 62% of microadenomas and small adenomas from familial adenomatous polyposis patients compared with adjacent normal mucosa, and this proportion was positively correlated with lesion size and degree of dysplasia. Cytofluorometric analysis of highly purified single epithelial cell suspensions from freshly excised carcinomas versus adjacent normal tissue demonstrated up to a 20-fold increase of mean cell-surface CEA in a group of colon carcinomas representative of the overall majority of such tumors--from Dukes' stages A to D and ranging mainly from well to moderately differentiated, the degree of overproduction was inversely correlated with tumor differentiation and directly correlated with stage. A marked tendency toward nonpolarized versus apical cell-surface expression with progression was also noted. Nonspecific cross-reacting antigen (NCA), a CEA family member, is also a homotypic adhesion molecule and blocks terminal myogenic differentiation, whereas biliary glycoprotein is a CEA family adhesion molecule that does not. Cell-surface NCA showed even greater overexpression (up to 70-told) in dedifferentiated tumors, whereas total-cell biliary glycoprotein showed approximately 2-fold lower levels than was normal in more differentiated tumors and approximately 2-fold higher levels than in further progressed tumors. These results therefore support the suggestion that CEA and NCA can directly contribute to colon carcinogenesis by inhibiting colonocyte differentiation.


Subject(s)
Biomarkers, Tumor/analysis , Carcinoembryonic Antigen/biosynthesis , Carcinoma/chemistry , Cell Adhesion Molecules , Colonic Neoplasms/chemistry , Membrane Glycoproteins/analysis , Adenoma/chemistry , Adenoma/pathology , Antigens, CD , Antigens, Neoplasm/biosynthesis , Carcinoembryonic Antigen/analysis , Carcinoma/pathology , Cell Differentiation , Colonic Diseases/pathology , Colonic Neoplasms/pathology , Epithelial Cells , Epithelium/metabolism , Epithelium/pathology , Fluorescent Antibody Technique, Indirect , Glycoproteins/analysis , Humans , Immunohistochemistry , Membrane Glycoproteins/biosynthesis , Neoplasm Staging , Precancerous Conditions/chemistry , Precancerous Conditions/pathology
18.
Clin Cancer Res ; 3(5): 661-7, 1997 May.
Article in English | MEDLINE | ID: mdl-9815734

ABSTRACT

Glutathione S-transferase (GST) represents a multifunctional enzyme family consisting of four known cytosolic isoforms (alpha, mu, pi, and Phi) that detoxify a variety of xenobiotic chemicals and may confer resistance to both chemotherapeutic drugs and carcinogens in various experimental models. GST-pi has already been extensively studied in clinical specimens, including breast cancer. We studied the immuno-histochemical distribution and relative immunopositivity of GST-alpha and GST-mu, based on a grading system for immunointensity, in samples of 51 neoplastic and 46 normal breast samples and 12 lymph node metastases from patients treated with intensive chemotherapy and bone marrow transplant. In normal breast tissue, GST-alpha localized predominantly to the cytoplasm of scattered cells lining the luminal aspects of the ducts. Occasional cells showed both cytoplasmic and nuclear GST-alpha immunoreactivity. GST-mu was stained in myoepithelial cells preferentially as well as in occasional ductal cells (including apocrine epithelium), vascular smooth muscle, and plasma cells. GST-alpha and GST-mu were detected in 22 of 51 (43%) and 24 of 48 (50%) invasive cancers, respectively. In paired samples of normal and malignant tissue from the same patient, GST-alpha immunostaining in cancers was significantly less intense compared to that of normal breast tissue in 13 of 41 (32%) cases. No such trend was found for GST-mu in paired samples. Neither GST-alpha nor GST-mu immunopositivity in tumor or nonneoplastic breast was found to correlate with relapse-free or overall survival in this clinical context; however, the apparent decreased expression of GST-alpha in malignant versus normal breast epithelial cells could have important implications in breast carcinogenesis.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Bone Marrow Transplantation , Breast Neoplasms/enzymology , Breast Neoplasms/therapy , Breast/enzymology , Glutathione Transferase/analysis , Hematopoietic Stem Cell Transplantation , Isoenzymes/analysis , Adult , Breast Diseases/enzymology , Breast Diseases/pathology , Breast Neoplasms/mortality , Breast Neoplasms/pathology , Female , Follow-Up Studies , Humans , Immunohistochemistry/methods , Lymphatic Metastasis , Menopause , Middle Aged , Receptors, Estrogen/analysis , Receptors, Progesterone/analysis , Survival Analysis , Time Factors
19.
Cancer Res ; 56(21): 4865-70, 1996 Nov 01.
Article in English | MEDLINE | ID: mdl-8895735

ABSTRACT

Recently, we reported that breast cancer cell lines fail to express the gene encoding the fatty acid binding protein mammary derived growth inhibitor (MDGI) and that transfection with an MDGI expression vector results in suppression of the malignant phenotype, suggesting that MDGI is a tumor suppressor gene. We also demonstrated that homozygous deletion and point mutation are not common mechanisms for silencing of the MDGI gene in human breast neoplasms. We now report that hypermethylation of HpaII and HhaI sites upstream of the first exon of the MDGI gene, and a SacII site in the first intron, occurs frequently in human breast cancer cell lines. This distinct methylation pattern is associated with loss of transcription and is reversible by treatment with 5-aza-deoxycytidine. Primary breast tumors also exhibited methylation of the SacII site (19 of 35, 54.3%) and the HpaII and HhaI sites (21 of 35, 66%). Hypermethylation of these sites was correlated with the absence of MDGI mRNA in these tumors. Our results suggest that epimutation of the MDGI gene leads to silencing, which, in turn, may initiate or contribute to progression of breast cancer.


Subject(s)
Breast Neoplasms/genetics , Carrier Proteins/genetics , Growth Inhibitors/genetics , Azacitidine/pharmacology , Breast Neoplasms/metabolism , DNA Methylation , Fatty Acid Binding Protein 3 , Fatty Acid-Binding Proteins , Female , Humans , Polymerase Chain Reaction , Tumor Cells, Cultured
20.
J Clin Endocrinol Metab ; 81(8): 2841-7, 1996 Aug.
Article in English | MEDLINE | ID: mdl-8768840

ABSTRACT

PTH-related peptide (PTHrP) is the major factor responsible for humoral hypercalcemia of malignancy. This paraneoplastic syndrome has been described in association with a number of malignancies, but rarely with carcinoma of the colon. Moreover, little is known about the molecular mechanisms that underlie PTHrP overexpression in tumors. Here we report a patient who presented with hypercalcemia 6 months after resection of a neuroendocrine colonic carcinoma (tumor I). At the time of admission, intact PTH was decreased, circulating PTHrP levels were elevated, and there was tumor recurrence (tumor II). Immunohistochemical staining of paraffin-embedded sections from tumor I did not stain for PTHrP, whereas cells from tumor II stained intensely positive. Southern blot analysis and differential PCR of genomic DNAs from tumor specimens and the patient's leukocytes demonstrated amplification of the PTHrP gene in tumor II. Moreover, staining for p53 protein was evident in tumor II, but not in tumor I, consistent with the presence of a mutant form of p53 and associated loss of tumor suppressor function in the recurrent tumor. PTHrP gene amplification was also detected in one of five other tumors associated with humoral hypercalcemia of malignancy. These findings suggest that a potential mechanism contributing to PTHrP overexpression in malignancies is gene amplification, which could arise from increased genomic instability associated with the progressive stages of neoplasia.


Subject(s)
Carcinoma/genetics , Colonic Neoplasms/genetics , Gene Amplification , Proteins/genetics , Aged , Base Sequence , Blotting, Southern , Carcinoma/metabolism , Carcinoma/pathology , Colonic Neoplasms/metabolism , Colonic Neoplasms/pathology , Disease Progression , Humans , Immunohistochemistry , Male , Molecular Probes/genetics , Molecular Sequence Data , Parathyroid Hormone/genetics , Parathyroid Hormone-Related Protein , Polymerase Chain Reaction , Tumor Suppressor Protein p53/physiology
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