bluuebunny/crossref_metadata_2025 huggingface dataset:
Epithelial c-jun and c-fos are temporally and spatially regulated by estradiol during neonatal rat oviduct differentiation
Authors: Okada A , Ohta Y , Brody SL , Iguchi T
Published: December 2004

Expression of transcription factors binding to the activating protein-1 (AP-1) site is induced by estrogens in association with epithelial proliferation in the uterus, but, in the oviduct, the relationship between cell proliferation and differentiation and AP-1 transcription factors is not well understood. In the developing rat oviduct, we found that proliferation and differentiation of epithelial cells were region-dependently regulated by 17beta-estradiol (E2). To determine the role of AP-1 transcription factors in the development of rat oviduct, we performed immunohistochemistry for epithelial c-jun and c-fos proteins in E2-untreated and -treated newborn rats. E2 increased the expression of c-jun and c-fos during proliferation of undifferentiated epithelial cells, but diminished both proteins during accelerated differentiation of ciliated epithelial cells. A pure estrogen receptor (ER) antagonist, ICI 182,780, inhibited changes in their expression during both cell proliferation and differentiation. Importantly, no reduction of c-jun was noted in the epithelial cells of the foxj1-deficient oviduct, which lacks cilia development. This study shows that c-jun and c-fos are regulated during epithelial cell proliferation and differentiation in a region-specific manner. This provides critical information for understanding the molecular and cellular mechanisms of the development of the neonatal oviduct.
Effects of peroxisome proliferator activated receptors-alpha and -gamma agonists on estradiol-induced proliferation and hyperplasia formation in the mouse uterus
Authors: Gunin AG , Bitter AD , Demakov AB , Vasilieva EN , Suslonova NV
Published: December 2004

It is suggested that the action of peroxisome proliferator-activated receptors (PPARs) cross-talks with estrogen signaling in the uterus. However, it is not known how PPAR agonists affect estrogen-dependent processes in the uterus, especially proliferation and morphogenetic changes. The effects of agonists of PPAR-alpha and -gamma on proliferative and morphogenetic reactions in the uterus under short- and long-term estrogen treatments were therefore examined. Ovariectomized mice were treated with estradiol dipropionate (4 micro g/100 g, s.c., once a week) or vehicle and rosiglitazone (PPAR-gamma agonist) or fenofibrate (PPAR-alpha agonist) or with no additional treatment for 2 days or for 30 days. Treatment with estradiol and PPAR agonists for 2 days did not affect uterine mass. In mice treated with estradiol and rosiglitazone for 2 days, proliferation was enhanced and levels of estrogen receptors-alpha and beta-catenin were decreased in all uterine tissues. Treatment with estradiol and fenofibrate for 2 days had the opposite effects on the parameters tested. In animals treated with estradiol and rosiglitazone for 30 days, uterine mass was increased, abnormal uterine glands and atypical endometrial hyperplasia were found more often and levels of estrogen receptors-alpha and beta-catenin were decreased. In animals treated with estradiol and fenofibrate for 30 days, uterine mass was decreased, most of the uterine glands had a normal structure, no cases of atypical hyperplasia were diagnosed, proliferative activity was declined and the levels of estrogen receptors-alpha and beta-catenin were markedly higher. Treatment with rosiglitazone or fenofibrate did not affect the serum estradiol level in the mice which received estradiol together with PPAR agonists for 30 days. Thus, rosiglitazone exerted the proliferative and morphogenetic effects of estradiol, but fenofibrate had the opposite effect. The actions of rosiglitazone and fenofibrate are associated with changes in the expression of estrogen receptors-alpha and beta-catenin in the uterus.
Expression of thyroid hormone receptor alpha in 3T3-L1 adipocytes; triiodothyronine increases the expression of lipogenic enzyme and triglyceride accumulation
Authors: Jiang W , Miyamoto T , Kakizawa T , Sakuma T , Nishio S , Takeda T , Suzuki S , Hashizume K
Published: December 2004

Thyroid hormone receptors (TR) are members of the nuclear receptor superfamily. There are at least two TR isoforms, TRalpha and TRbeta, which act as mediators of thyroid hormone in tissues. However, the relative expression of each TR isoform in target tissues is still elusive. Herein, we have developed an RT-PCR and restriction enzyme digestion method to determine the expression of TRalpha1 and TRbeta1. We analyzed the expression of TR isoforms in 3T3-L1 preadipocytes induced to differentiate by an adipogenic cocktail in the presence or absence of 100 nM triiodothyronine (T(3)). The TRalpha1 isoform was predominantly expressed in 3T3-L1 adipocytes, and its expression was increased at the stage of development concomitant with the emergence of lipid droplets. Little, if any, TRbeta1 mRNA was detected in adipocytes. Administration of T(3) to the differentiating 3T3-L1 cells enhanced the accumulation of triglyceride. The expression profile of TRalpha1 in T(3)-treated adipocytes was similar to that in non-treated cells. The transcripts of adipogenic factors, CCAAT/enhancer binding protein beta (C/EBPbeta) and peroxisome proliferator activated receptor gamma (PPARgamma), were not altered by T(3). Lipid binding protein, aP2, that is downstream of these transcription factors was also unaffected by T(3). In contrast, the lipogenic enzyme, glyceraldehyde-3-phosphate dehydrogenase mRNA was significantly increased in the presence of T(3). Therefore, T(3) appears to be a hormone capable of modulating the expression of lipogenic enzyme and augments the accumulation of lipid droplets. We conclude that the TRalpha isoform might play an important role in the generation and maintenance of the mature adipocyte phenotype, regulating the expression of lipogenic enzymes.
Simultaneous changes in central and peripheral components of the hypothalamus-pituitary-thyroid axis in lipopolysaccharide-induced acute illness in mice
Authors: Boelen A , Kwakkel J , Thijssen-Timmer DC , Alkemade A , Fliers E , Wiersinga WM
Published: December 2004

During illness, major changes in thyroid hormone metabolism and regulation occur; these are collectively known as non-thyroidal illness and are characterized by decreased serum triiodothyronine (T(3)) and thyroxine (T(4)) without an increase in serum TSH. Whether alterations in the central part of the hypothalamus-pituitary-thyroid (HPT) axis precede changes in peripheral thyroid hormone metabolism instead of vice versa, or occur simultaneously, is presently unknown. We therefore studied the time-course of changes in thyroid hormone metabolism in the HPT axis of mice during acute illness induced by bacterial endotoxin (lipopolysaccharide; LPS).LPS rapidly induced interleukin-1beta mRNA expression in the hypothalamus, pituitary, thyroid and liver. This was followed by almost simultaneous changes in the pituitary (decreased expression of thyroid receptor (TR)-beta2, TSHbeta and 5'-deiodinase (D1) mRNAs), the thyroid (decreased TSH receptor mRNA) and the liver (decreased TRbeta1 and D1 mRNA). In the hypothalamus, type 2 deiodinase mRNA expression was strongly increased whereas preproTRH mRNA expression did not change after LPS. Serum T(3) and T(4) fell only after 24 h.Our results suggested almost simultaneous involvement of the whole HPT axis in the downregulation of thyroid hormone metabolism during acute illness.
Suppression of cell proliferation and regulation of estrogen receptor alpha signaling pathway by arsenic trioxide on human breast cancer MCF-7 cells
Authors: Chow SK , Chan JY , Fung KP
Published: December 2004

In recent years, breast cancers have aroused much concern. Together with a growing incidence all over the world, the development of drug resistance to tamoxifen, the most commonly prescribed chemotherapeutic drug for breast cancer patients, has highlighted the importance of developing a new chemotherapeutic drug in combating breast cancer. With the aim of treating breast cancers, the anti-tumor effects of arsenic trioxide in MCF-7 cells have been studied. MCF-7 cells are estrogen responsive cells which mimic breast cancers at the early stage. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and direct cell counting were used to measure cell proliferation. The mechanisms of action were elucidated through the measurement of estrogen receptor (ER) binding, mRNA and protein levels of ERalpha and its activity. We have demonstrated that arsenic trioxide was capable of reducing cell survival in MCF-7 cells via the suppression of the estrogen-induced growth stimulatory effects in MCF-7 cells. Arsenic trioxide was shown to suppress the action of estrogen through the regulation of the ERalpha signaling pathway. Arsenic trioxide could down-regulate ERalpha mRNA and protein levels without competing with estrogen for ERalpha binding. Arsenic trioxide also inhibited the transcription activity mediated by the ERalpha signaling pathway and ultimately it down-regulated c-myc protein expression and inhibited cell entry to S phase under estrogen's stimulation. In conclusion, arsenic trioxide could inhibit the growth of MCF-7 cells by reducing the growth stimulatory effect of estrogen. As estrogen is a primary risk factor in promoting the growth of breast tumor cells, the anti-estrogenicity exhibited by arsenic trioxide sheds light on the therapy of breast cancer.
Prohormone convertase 1 (PC1) processing and sorting: effect of PC1 propeptide and proSAAS
Authors: Lee SN , Prodhomme E , Lindberg I
Published: December 2004

Prohormone convertase 1 (PC1) is a serine proteinase responsible for the proteolytic processing of many precursor proteins within the regulated secretory pathway. The activity of PC1 is potentially regulated by two endogenous inhibitors, the PC1 propeptide and proSAAS. Here we have investigated the effect of proSAAS and propeptide-containing constructs on PC1 carboxy-terminal processing and activity. In AtT-20 cells, proSAAS expression inhibited both C-terminal PC1 processing and proopiomelanocortin (POMC) processing under pulse/chase conditions. SAAS CT peptide-propeptide chimeric constructs had no effect on the cleavage of PC1 and POMC under pulse/chase conditions. However, a construct containing the propeptide alone reduced C-terminal PC1 processing under pulse/chase conditions and also inhibited POMC processing. In contrast, experiments using HEK293 cells transiently expressing PC1 plus the respective constructs demonstrated significant inhibition of zymogen processing and decreased C-terminal processing of PC1 by the SAAS CT peptide portion of the chimera. Our results suggest that the PC1 propeptide expressed in trans is able to act as an endogenous inhibitor of PC1, but that SAAS CT peptide-containing/propeptide constructs cannot function as effective inhibitors of precursor maturation in the regulated pathway.
Leptin modulates fertility under the influence of elevated growth hormone as modeled in oMt1a-oGH transgenic mice
Authors: Thomas AD , Murray JD , Oberbauer AM
Published: December 2004

Elevated growth hormone (GH) concentrations suppress reproductive function in a variety of species, although it is unclear whether GH directly suppresses reproductive performance, or whether GH activates other pathways to achieve these effects. The ovine metallothionein 1a-ovine GH (oMt1a-oGH) transgenic mouse has been used to model the effects of GH on both body composition and reproductive function. A recent report has documented increased leptin levels in obese oMt1a-oGH mice. Given the importance of leptin in modulation of the reproductive endocrine axis, as well as the reports documenting reduced leptin signal transduction in animals with elevated leptin levels, we hypothesized that high leptin concentrations in response to elevated GH would reduce fertility. To determine the effects of high circulating leptin levels on the reproductive endocrine axis, we assessed hypothalamic neuropeptide Y (NPY) and GnRH expression. At weaning, oMt1a-oGH transgenic (TG) and wild-type (WT) female mice were allocated to one of four treatment groups: oMt1a-oGH females chronically expressing the transgene (TG ON); oMt1a-oGH females expressing the transgene from 3 to 8 weeks of age (TG ON/OFF); WT females receiving the transgene stimulus from 3 to 8 weeks of age (WT ON/OFF); and WT females never receiving the transgene stimulus (WT OFF). Eight-week-old females were housed with males for a 2-week period, after which females were isolated from males and allowed to carry pregnancies to term. Body and gonadal fat pad (GFP) weights, along with plasma leptin concentrations, estrous cyclicity, pregnancy rate and litter characteristics, were recorded for each female. Chronic expression of the oMt1a-oGH transgene resulted in larger leaner mice, and inactivation of the transgene produced obese females. Pregnancy rate was reduced in TG ON females when compared with all other groups, and infertility was associated with elevated leptin levels. In addition, high leptin levels were associated with increased NPY expression, suggesting reduced leptin-signaling capacity, which may contribute to suppression of the reproductive axis in oGH animals.
Increased in vivo phosphorylation of insulin receptor at serine 994 in the liver of obese insulin-resistant Zucker rats
Authors: Coba MP , Munoz MC , Dominici FP , Toblli JE , Pena C , Bartke A , Turyn D
Published: December 2004

Serine phosphorylation of the insulin receptor (IR) has been proposed to exert an inhibitory influence on its tyrosine kinase activity. Previous works using site-directed mutagenesis suggested that serine 994 of the IR (IR Ser 994) might be part of an inhibitory domain of the receptor. In this study we examined whether this residue is subjected to phosphorylation in vivo. We used a site-phosphospecific antibody to determine the extent of phosphorylation of IR Ser 994 in insulin target tissues from two animal models of insulin resistance with different IR kinase (IRK) activity: obese (fa/fa) Zucker rats and transgenic mice overexpressing bovine growth hormone (PEPCK-bGH mice).Phosphorylation at IR Ser 994 was markedly increased in liver of obese rats. This alteration appeared to be tissue-selective since no phosphorylation on Ser 994 was detected in IRs isolated from skeletal muscle of these animals. On the other hand, the phosphorylation level of IR Ser 994 was very low in liver of PEPCK-bGH mice and did not differ from that of the control group. We have also demonstrated that protein kinase (PK) C isoforms alpha, betaI and zeta are able to promote the in vitro phosphorylation of the IR on Ser 994. Differential findings in these two models of insulin resistance might thus reflect increased PKC activity resulting from increased lipid availability in obese Zucker rats. Our results suggest that Ser 994 is a novel in vivo IR phosphorylation site that might be involved in the regulation of the IRK in some states of insulin resistance.
Proteolytic degradation of IGF-binding protein (IGFBP)-2 in equine ovarian follicles: involvement of pregnancy-associated plasma protein-A (PAPP-A) and association with dominant but not subordinated follicles
Authors: Gerard N , Delpuech T , Oxvig C , Overgaard MT , Monget P
Published: December 2004

In the ovary of mammalian species, terminal follicular growth is accompanied by a decrease in intrafollicular levels of IGF-binding protein-2 (IGFBP-2) and IGFBP-4. The decrease in IGFBP-4 levels is essentially due to an increase in proteolytic cleavage by intrafollicular pregnancy-associated plasma protein-A (PAPP-A) in growing healthy follicles. The decrease in IGFBP-2 levels is partly due to a decrease in mRNA expression by follicular cells. In addition, we have recently shown that IGFBP-2 is also proteolytically cleaved by PAPP-A in bovine and porcine growing follicles. In the present work, we showed that follicular fluid from late dominant equine follicles (35 mm diameter) contains a proteolytic activity against IGFBP-2. First follicular fluid from dominant follicles contained lower levels of native IGFBP-2 than the corresponding serum, as assessed by Western ligand blotting. In contrast, immunoblotting experiments showed much higher levels of a 12 kDa proteolytic fragment in dominant follicular fluid than in the serum. Moreover, equine dominant follicular fluid was able to induce proteolysis of exogenous recombinant bovine (rb)IGFBP-2, this degradation being dose-dependently enhanced by IGFs. The proteolytic activity against IGFBP-2 in equine follicles was partially immunoneutralized by a polyclonal antibody raised against human PAPP-A. Moreover, cleavage of rbIGFBP-2 by equine follicular fluid was dose-dependently inhibited by a peptide derived from the heparin-binding domain of IGFBP-5, as well as by peptides derived from other heparin-binding domain-containing proteins such as connective tissue growth factor, vitronectin and heparin-interacting protein, previously shown to inhibit PAPP-A. Finally, the proteolytic activity was very low in subordinate follicles, was high in both early (25 mm diameter) and late (35 mm diameter) dominant follicles, and was slightly lower in preovulatory follicles recovered 35 h after human chorionic gonadotropin (hCG) treatment.Overall, these data show that in the equine ovary, the selection of dominant follicles is associated with an increase of the proteolytic degradation of IGFBP-2 by PAPP-A, as for IGFBP-4, and potentially other protease(s), probably contributing to the increase in IGF bioavailability. In atretic subordinate follicles, the decrease in the proteolytic degradation of IGFBP-2, probably due in part to a direct inhibition by peptides containing heparin-binding domains, contributes to the increase in IGFBP-2 levels and the decrease in IGF bioavailability. The expression of PAPP-A and IGFBP-2 mRNA during folliculogenesis remain to be investigated in the mare.
Thyroid papillary carcinoma: preliminary evidence for a germ-line single nucleotide polymorphism in the Fas gene
Authors: Basolo F , Giannini R , Faviana P , Fontanini G , Patricelli Malizia A , Ugolini C , Elisei R , Miccoli P , Toniolo A
Published: December 2004

The expression of Fas in thyroid tumours and Graves' disease was analysed by mRNA transcript expression. As compared with unaffected thyroid tissue, Fas expression was enhanced in Graves' disease, adenomas, and papillary and follicular carcinomas. This pattern was also reflected in immunohistochemical studies. The PCR single-strand conformational polymorphism (SSCP) method and DNA sequencing were used to analyse Fas exons 1-9. The study was carried out on five different histotypes of thyroid tumours (n=93) and tissue from Graves' disease patients. As compared with a group of healthy blood donors (n=64), a significant association (P=0.006) emerged between papillary thyroid carcinoma and a silent single nucleotide polymorphism (SNP, 988C-->T) in exon 7 of the Fas gene. Other forms of thyroid pathology were not associated with the above polymorphism. Patients with neoplasia showed the same SNP in tumour tissue, in the unaffected contralateral thyroid lobe, and in peripheral blood cells. Thus, the 988C-->T polymorphism appeared to be of germ-line origin.
Identification of the structural and functional determinants of the extracellular domain of the human follicle stimulating hormone receptor
Authors: Kene PS , Nalavadi VC , Dighe RR , Iyer KS , Mahale SD
Published: December 2004

The extracellular domain (ECD) of the human follicle-stimulating hormone receptor (hFSHR) is believed to be the major determinant of hormone selectivity. Different discrete, discontinuous regions on the ECD of the hFSHR have been suggested to be crucial for hormone binding. However, the role of the ECD in signal transduction is not well understood. This study provides some insight into these aspects of the structure-function relationship of the ECD of hFSHR. Ten peptides were selected from the ECD on the basis of their ability to be surface oriented, synthesized by the solid-phase method using fluorenylmethyloxycarbonyl chemistry, purified and characterized. They were further studied for their ability to modulate both human follicle-stimulating hormone (hFSH)-FSHR binding and cAMP generation. Competitive inhibition studies showed that, of all the peptides studied, peptides 285-300 and 297-310 hFSHR were able to inhibit hFSH binding to FSHR. Both peptides function as weak competitive inhibitors of hFSH-FSHR binding. Peptides 285-300 hFSHR, 216-235 hFSHR, 184-195 hFSHR, 79-89 hFSHR and 15-31 hFSHR were observed to inhibit FSH-induced cAMP production. In summary, this study suggests that discrete, functional domains of the ECD have a role in hormone binding and signal transduction. Region 285-300 has been identified as a novel region crucial for both FSH binding and cAMP generation.
Type II iodothyronine deiodinase protein in chicken choroid plexus: additional perspectives on T3 supply in the avian brain
Authors: Verhoelst C H J , Darras V M , Roelens S A , Artykbaeva G M , Van der Geyten S
Published: December 2004

It is widely accepted that type II iodothyronine deiodinase (D2) is mostly present in the brain, where it maintains the homeostasis of thyroid hormone (TH) levels. Although intensive studies have been performed on activity and mRNA levels of the deiodinases, very little is known about their expression at the protein level due to the lack of specific antisera. The current study reports the production of a specific D2 polyclonal antiserum and its use in the comparison of D2 protein distribution with that of type I (D1) and type III (D3) deiodinase protein in the choroid plexus at the blood–brain barrier level. Immunocytochemistry showed very high D2 protein expression in the choroid plexus, especially in the epithelial cells, whereas the D1 and D3 proteins were absent. Furthermore, dexamethasone treatment led to an up-regulation of the D2 protein in the choroid plexus. The expression of D2 protein in the choroid plexus led to a novel insight into the working mechanism of the uptake and transport of thyroid hormones along the blood–brain barrier in birds. It is hypothesized that D2 allows the prohormone thyroxine (T4) to be converted into the active 3,5,3′-triiodothyronine (T3). Within the choroidal epithelial cells. T3 is subsequently bound to its carrier protein, transthyretin (TTR), to allow transport through the cerebrospinal fluid. Neurons can thus not only be provided with a sufficient T3 level via the aid of the astrocytes, as was hypothesized previously based on in situ hybridization data, but also by means of T4 deiodination by D2, directly at the blood–brain barrier level.
Thyroid hormones modulate the endocrine and autocrine/paracrine actions of leptin on thyrotropin secretion
Authors: da Veiga M A L Costa , de Jesus Oliveira K , Curty F H , de Moura C C Pazos
Published: December 2004

We investigated the influence of hypo- and hyperthyroidism on the ability of leptin to modulate TSH secretion. Two hours after receiving leptin (8 μg leptin/100 g BW; s.c.), hyperthyroid rats (10 μg thyroxine (T4)/100 g body weight (BW) for 5 days) showed a 1.7-fold increase in serum TSH (P<0.05); in hypothyroid rats, leptin had no effect. Hemi-pituitaries of hyperthyroid rats incubated with 10−9 and 10−7M leptin showed reductions in TSH release of 40 and 50% respectively (P<0.05); incubation with 1:2000 and 1:500 dilutions of antiserum against leptin resulted in 3- and 4-fold higher TSH release (P<0.05 and P<0.001 respectively). However, in hypothyroid pituitaries leptin or the antiserum had no effect. The results suggest that the in vivo and in vitro responsiveness of TSH to leptin is abolished in hypothyroidism and is preserved in short-term hyperthyroidism, in comparison to previous reports in euthyroidism. In addition, the inhibitory action of pituitary leptin is enhanced in hyperthyroid glands, which may suggest a role for locally produced leptin in the suppression of TSH release associated with hyperthyroidism.
Lentiviral vectors efficiently transduce human gonadotroph and somatotroph adenomas in vitro. Targeted expression of transgene by pituitary hormone promoters
Authors: Roche Catherine , Zamora Alfredo J , Taïeb David , Lavaque Esteban , Rasolonjanahary Ramahefarizo , Dufour Henri , Bagnis Claude , Enjalbert Alain , Barlier Anne
Published: December 2004

Despite important advances in human therapeutics, no specific treatment for both non-functioning gonadotroph and resistant somatotroph adenomas is available. Gene transfer by viral vectors can be considered as a promising way to achieve a specific and efficient treatment. Here we show the possibility of efficient gene transfer in human pituitary adenoma cells in vitro using a human immunodeficiency virus (HIV)-type 1-derived vector. Using enhanced green fluorescent protein (eGFP) gene as a marker placed under the phosphoglycerate kinase (PGK) promoter, gonadotroph and somatotroph adenomas were transduced even with moderate viral loads. The expression started at day 2, reached a peak at day 5, and it was still present at day 90. For targeting somatotroph and gonadotroph adenomas, human growth hormone (GH) promoter (GH −481, +54 bp) and two fragments of the human glycoprotein hormone α-subunit promoter (α-subunit 1 −520, +33 bp, and α-subunit 2 −907, +33 bp) were tested. In gonadotroph adenomas, the percentage of identified fluorescent cells and the fluorescence intensity analyzed by fluorescence-activated cell sorting indicated that the strength of the α-subunit 1 and α-subunit 2 promoters were comparable to that of the PGK promoter. Primary cultures of rat pituitary cells showed that α-subunit 1 is more selective to thyreotroph and gonadotroph phenotypes than α-subunit 2. GH promoter activity appeared weak in somatotroph adenomas. The human GH enhancer did not increase the GH promoter activity at all but the human prolactin promoter (−250 bp) allowed 4-fold more fluorescent cells to be obtained than the GH promoter. Several cell lines appeared too permissive to test cell-specificity of pituitary promoters. However, on human non-pituitary cell cultures, the tested pituitary promoters seemed clearly selective to target endocrine pituitary phenotypes. This study gives a starting point for a gene-therapy program using lentiviral vectors to transfer therapeutic genes in human pituitary adenomas.
Androgen generation in adipose tissue in women with simple obesity – a site-specific role for 17β-hydroxysteroid dehydrogenase type 5
Authors: Quinkler Marcus , Sinha Binayak , Tomlinson Jeremy W , Bujalska Iwona J , Stewart Paul M , Arlt Wiebke
Published: December 2004

Women with polycystic ovary syndrome (PCOS) have high circulating androgens, thought to originate from ovaries and adrenals, and frequently suffer from the metabolic syndrome including obesity. However, serum androgens are positively associated with body mass index (BMI) not only in PCOS, but also in simple obesity, suggesting androgen synthesis within adipose tissue. Thus we investigated androgen generation in human adipose tissue, including expression of 17β-hydroxysteroid dehydrogenase (17β-HSD) isozymes, important regulators of sex steroid metabolism. Paired omental and subcutaneous fat biopsies were obtained from 27 healthy women undergoing elective abdominal surgery (age range 30–50 years; BMI 19.7–39.2 kg/m2). Enzymatic activity assays in preadipocyte proliferation cultures revealed effcient conversion of androstenedione to testosterone in both subcutaneous and omental fat. RT-PCR of whole fat and preadipocytes of subcutaneous and omental origin showed expression of 17β-HSD types 4 and 5, but no relevant expression of 17β-HSD types 1, 2, or 3. Microarray analysis confirmed this expression pattern (17β-HSD5>17β-HSD4) and suggested a higher expression of 17β-HSD5 in subcutaneous fat. Accordingly, quantitative real-time RT-PCR showed significantly higher expression of 17β-HSD5 in subcutaneous compared with omental fat (P<0.05). 17β-HSD5 expression in subcutaneous, but not omental, whole fat correlated significantly with BMI (r=0.51, P<0.05). In keeping with these findings, 17β-HSD5 expression in subcutaneous fat biopsies from six women taking part in a weight loss study decreased significantly with weight loss (P<0.05). A role for 17β-HSD5 in adipocyte differentiation was further supported by the observed increase in 17β-HSD5 expression upon differentiation of stromal preadipocytes to mature adipocytes (n=5; P<0.005), which again was higher in cells of subcutaneous origin. Functional activity of 17β-HSD5 also significantly increased with differentiation, revealing a net gain in androgen activation (androstenedione to testosterone) in subcutaneous cultures, contrasting with a net gain in androgen inactivation (testosterone to androstenedione) in omental cultures. Thus, human adipose tissue is capable of active androgen synthesis catalysed by 17β-HSD5, and increased expression in obesity may contribute to circulating androgen excess.
Reproductive failure in mice lacking inter-alpha-trypsin inhibitor (ITI) – ITI target genes in mouse ovary identified by microarray analysis
Authors: Suzuki Mika , Kobayashi Hiroshi , Tanaka Yoshiko , Kanayama Naohiro , Terao Toshihiko
Published: December 2004

Bikunin, a Kunitz-type protease inhibitor, is found in blood and urine. It has been established by two laboratories independently that the bikunin knockout female mice display a severe reduction in fertility: the cumulus oophorus has a defect in forming the extracellular hyaluronan-rich matrix during expansion. Proteins of the inter-alpha-trypsin inhibitor (ITI) family are eliminated in mice in which the bikunin gene has been inactivated, since bikunin is essential for their biosynthesis. Proteins of the ITI family may contribute to the microenvironment in which ovulation takes place. It is not clear, however, whether a single mechanism affects the reproductive function including ovulation. For identifying the full repertoire of the ITI deficiency-related genes, a cDNA microarray hybridization screening was conducted using mRNA from ovaries of wild-type or bik−/− female mice. A number of genes were identified and their regulation was confirmed by real-time RT-PCR analysis. Our screen identified that 29 (0.7%) and 5 genes (0.1%) of the genes assayed were, respectively, up- and down-regulated twofold or more. The identified genes can be classified into distinct subsets. These include stress-related, apoptosis-related, proteases, signaling molecules, aging-related, cytokines, hyaluronan metabolism and signaling, reactive oxygen species-related, and retinoid metabolism, which have previously been implicated in enhancing follicle development and/or ovulation. Real-time RT-PCR analysis confirmed that these genes were up- and down-regulated two- to tenfold by bikunin knockout. These studies demonstrate that proteins of the ITI family may exert potent regulatory effects on a major physiological reproductive process, ovulation.
Comparative effects of amino acids and glucose on insulin secretion from isolated rat or mouse islets
Authors: Zawalich W S , Yamazaki H , Zawalich K C , Cline G
Published: December 2004

Glucose and the combination of leucine and glutamine were used to stimulate insulin secretion from rat islets during a dynamic perifusion and the responses obtained were compared with those elicited from mouse islets under identical conditions. In rat islets, glucose (15 mM) or the amino acid combination of 10 mM glutamine plus 20 mM leucine were most efficacious and peak second-phase insulin release responses were 20- to 30-fold above prestimulatory rates. In contrast to rat islet responses, sustained second-phase insulin secretory responses to the same agonists were minimally increased 1- to 2-fold from mouse islets. Parallel studies demonstrated that phospholipase C (PLC) was markedly activated in rat, but not mouse, islets by both high glucose concentrations and the amino acid combination.

Additional studies documented that glucose and amino acid responses of both rat and mouse islets were amplified by carbachol or forskolin. However, wortmannin, a phosphatidylinositol 3-kinase inhibitor, amplified only the responses to glucose leaving the responses to the amino acid mixture unaltered. These observations support the concept that mitochondrial metabolism alone is minimally effective in stimulating insulin secretion from islets. The activation of the supplementary second messenger systems (PLC and/or cAMP) appears essential for the emergence of their full secretory potential. The mechanism regulating the potency and specificity of wortmannin’s impact on glucose-induced secretion remains to be identified; however a unique mechanism is supported by these findings.
Additional case of Tsukahara's syndrome or new syndrome: Further delineation of the association of microcephaly and radio‐ulnar synostosis
Authors: Selicorni Angelo , Ferrarini Alessandra , Cagnoli Giacomo , Fratoni Alessia , Bottigelli Michaela , Milani Donatella
Published: December 2004

In 1994, Giuffrè et al. reported two unrelated families in which some of the members had microcephaly and radio‐ulnar synostosis, suggesting a new condition. Since this first report, Tsukahara et al. and Udler et al. described two distinct patients with a different condition characterized by radio‐ulnar synostosis, short stature, microcephaly, scoliosis, and mental retardation. Here we report on a new case of microcephaly and radio‐ulnar synostosis and discuss the possible relationship between Tsukahara's syndrome and the phenotype described by Giuffrè et al. © 2004 Wiley‐Liss, Inc.
Hand involvement in Schmid metaphyseal chondrodysplasia
Authors: Elliott Alison M. , Field Fiona M. , Rimoin David L. , Lachman Ralph S.
Published: December 2004

Schmid metaphyseal chondrodysplasia (Schmid MCD, MIM 156500) is caused by mutations in the COL10A1 gene and is clinically characterized by short stature, bowed legs, and a waddling gait. Radiographic findings include anterior cupping, sclerosis and splaying of the ribs, diffuse metaphyseal flaring, and irregularity that is most pronounced at the knees, coxa vara, and femoral bowing. We reviewed the radiographs of Schmid MCD patients at the International Skeletal Dysplasia Registry in Los Angeles for evidence of hand involvement. We found hand involvement in 47% (7/15) of cases included in our analysis. These changes were subtle and consisted of shortening of the tubular bones and metaphyseal cupping of the proximal phalanges and metacarpals. Mild hand involvement is a common feature of Schmid MCD. © 2004 Wiley‐Liss, Inc.
A semi‐supervised active learning algorithm for information extraction from textual data
Authors: Wu Tianhao , Pottenger William M.
Published: December 2004

In this article we present a semi‐supervised active learning algorithm for pattern discovery in information extraction from textual data. The patterns are reduced regular expressions composed of various characteristics of features useful in information extraction. Our major contribution is a semi‐supervised learning algorithm that extracts information from a set of examples labeled as relevant or irrelevant to a given attribute. The approach is semi‐supervised because it does not require precise labeling of the exact location of features in the training data. This significantly reduces the effort needed to develop a training set. An active learning algorithm is used to assist the semi‐supervised learning algorithm to further reduce the training set development effort. The active learning algorithm is seeded with a single positive example of a given attribute. The context of the seed is used to automatically identify candidates for additional positive examples of the given attribute. Candidate examples are manually pruned during the active learning phase, and our semi‐supervised learning algorithm automatically discovers reduced regular expressions for each attribute. We have successfully applied this learning technique in the extraction of textual features from police incident reports, university crime reports, and patents. The performance of our algorithm compares favorably with competitive extraction systems being used in criminal justice information systems.
Technology, security, and individual privacy: New tools, new threats, and new public perceptions
Authors: Strickland Lee S. , Hunt Laura E.
Published: December 2004

Highly portable information collection and transmission technologies such as radio frequency identification (RFID) tags and smart cards are becoming ubiquitous in government and business—employed in functions including homeland security, information security, physical premises security, and even the control of goods in commerce. And, directly or indirectly, in many of these applications, it is individuals and their activities that are tracked. Yet, a significant unknown is (a) whether the public understands these technologies and the manner in which personally identifiable information may be collected, maintained, used, and disseminated; and (b) whether the public consents to these information practices. To answer these and related questions, we surveyed a select group of citizens on the uses of this technology for business as well as homeland security purposes. We found a significant lack of understanding, a significant level of distrust even in the context of homeland security applications, and a very significant consensus for governmental regulation. We conclude that a primary objective for any organization deploying these technologies is the promulgation of a comprehensive Technology Privacy Policy, and we provide detailed specifications for such an effort.
Developmental gene expression profiling of mammalian, fetal orofacial tissue
Authors: Mukhopadhyay Partha , Greene Robert M. , Zacharias Wolfgang , Weinrich Martin C. , Singh Saurabh , Young William W. , Pisano M. Michele
Published: December 2004

BACKGROUND

The embryonic orofacial region is an excellent developmental paradigm that has revealed the centrality of numerous genes encoding proteins with diverse and important biological functions in embryonic growth and morphogenesis. DNA microarray technology presents an efficient means of acquiring novel and valuable information regarding the expression, regulation, and function of a panoply of genes involved in mammalian orofacial development.

METHODS

To identify differentially expressed genes during mammalian orofacial ontogenesis, the transcript profiles of GD‐12, GD‐13, and GD‐14 murine orofacial tissue were compared utilizing GeneChip arrays from Affymetrix. Changes in gene expression were verified by TaqMan quantitative real‐time PCR. Cluster analysis of the microarray data was done with the GeneCluster 2.0 Data Mining Tool and the GeneSpring software.

RESULTS

Expression of >50% of the ∼12,000 genes and expressed sequence tags examined in this study was detected in GD‐12, GD‐13, and GD‐14 murine orofacial tissues and the expression of several hundred genes was up‐ and downregulated in the developing orofacial tissue from GD‐12 to GD‐13, as well as from GD‐13 to GD‐14. Such differential gene expression represents changes in the expression of genes encoding growth factors and signaling molecules; transcription factors; and proteins involved in epithelial‐mesenchymal interactions, extracellular matrix synthesis, cell adhesion, proliferation, differentiation, and apoptosis. Following cluster analysis of the microarray data, eight distinct patterns of gene expression during murine orofacial ontogenesis were selected for graphic presentation of gene expression patterns.

CONCLUSIONS

This gene expression profiling study identifies a number of potentially unique developmental participants and serves as a valuable aid in deciphering the complex molecular mechanisms crucial for mammalian orofacial development. Supplementary material for this article can be found at http://www.mrw.interscience.wiley.com/suppmat/1542‐0752/suppmat/2004/70/v70.mukhopadhyay.html Birth Defects Research (Part A), 2004. © 2004 Wiley‐Liss, Inc.
Genetic approaches to identify disease genes for birth defects with cleft lip/palate as a model
Authors: Lidral Andrew C. , Murray Jeffrey C.
Published: December 2004

BACKGROUND

Understanding the etiology of birth defects is an important step toward developing improved treatment and preventive strategies. Most birth defects have an underlying genetic basis, ranging from single genes playing dominant or recessive roles in Mendelian disorders to a mixture of contributions from multiple genes and environmental triggers in complex traits. The purpose of this article is to provide an overview of genetic approaches to identifying disease genes for genetically complex birth defects.

METHODS

A review of the literature describing successes and limitations for identifying disease genes for complex traits was conducted.

RESULTS

Cleft lip and cleft palate are common congenital anomalies with significant medical, psychological, social, and economic ramifications. The Online Mendelian Inheritance in Man catalog (OMIM; http://www3.ncbi.nlm.nih.gov/Omim) lists more than 400 single‐gene causes of clefts of the lip and/or palate. Genetic causes of clefting also include chromosomal rearrangements, genetic susceptibility to teratogenic exposures, and complex genetic contributions of multiple genes.

CONCLUSIONS

Genetic causes of birth defects can be identified using an increasingly powerful combination of careful sample collection, molecular analytic methods, and statistical evaluations. We will describe a range of approaches to search for genetic factors of birth defects and use our own work with cleft lip and palate as a model. Birth Defects Research (Part A), 2004. © 2004 Wiley‐Liss, Inc.
Eigenspannungen in zweischichtigen Betonplatten, insbesondere mit nachträglich aufgebrachter zweiter Schicht
Authors: Sieber Edgar
Published: December 2004

Eigenspannungen in zweischichtigen Betonplatten infolge unterschiedlicher Volumenänderungen der Schichten aus Temperatureinwirkungen oder Schwinden können Ursache für Schäden sein. Es wird ein Rechenverfahren zur Bestimmung derartiger Eigenspannungen abgeleitet und an einem Beispiel, bei dem die zweite Schicht nachträglich aufgebracht worden ist, erläutert.
Risikountersuchung am Beispiel historischer Brücken unter Schiffsanprall
Authors: Curbach Manfred , Proske Dirk
Published: December 2004

Zur Bewertung der Sicherheit vorhandener Bauwerke wird der Sicherheitsbegriff in den Risikobegriff überführt. Anschließend werden verschiedene Parameter zur Abbildung von Risiken vorgestellt. Diese Parameter werden bei der Beurteilung der Sicherheit von zwei historischen Brücken über schiffbare Flüsse unter der möglichen Einwirkung Schiffsanprall beispielhaft verwendet.
Antioxidative Effects of Flavonols and Their Glycosides against the Free‐Radical‐Induced Peroxidation of Linoleic Acid in Solution and in Micelles
Authors: Zhou Bo , Miao Qing , Yang Li , Liu Zhong‐Li
Published: December 2004

The antioxidative effect of flavonols and their glycosides against the peroxidation of linoleic acid has been studied in homogeneous solution (tBuOH/H2O, 3:2) and in sodium dodecyl sulfate and cetyl trimethylammonium bromide micelles. The peroxidation was initiated thermally by the water‐soluble initiator 2,2′‐azobis(2‐methylpropionamidine) dihydrochloride, and the reaction kinetics were studied by monitoring the formation of linoleic acid hydroperoxides. The synergistic antioxidant effect of the flavonols with α‐tocopherol (vitamin E) was also studied by following the decay kinetics of α‐tocopherol and the α‐tocopheroxyl radical. Kinetic analysis of the antioxidative process demonstrates that the flavonols are effective antioxidants in solution and in micelles, either alone or in combination with α‐tocopherol. The antioxidative action involves trapping the initiating radicals in solution or in the bulk‐water phase of the micelles, trapping the propagating lipid peroxyl radicals on the surface of the micelles, and regenerating α‐tocopherol by reducing the α‐tocopheroxyl radical. It was found that the antioxidant activity of the flavonols and their glycosides depends significantly on the position and number of the hydroxy groups, the oxidation potential of the molecule, and the reaction medium. The flavonols bearing ortho‐dihydroxy groups possess significantly higher antioxidative activity than those without such functionalities, and the glycosides are less active than their parent aglycones. The activity of the flavonols is higher in micelles than in solution, while the activity of α‐tocopherol is lower in micelles than in solution. This is because the predominant factor for controlling the activity is the hydrogen‐bonding interaction of the antioxidant with the micellar surface in the case of hydrophilic flavonols, while it is the inter‐ and intramicellar diffusion in the case of lipophilic α‐tocopherol.
Dissecting the Intimate Mechanism of Cyanation of {2Fe3S} Complexes Related to the Active Site of All‐Iron Hydrogenases by DFT Analysis of Energetics, Transition States, Intermediates and Products in the Carbonyl Substitution Pathway
Authors: Zampella Giuseppe , Bruschi Maurizio , Fantucci Piercarlo , Razavet Mathieu , Pickett Christopher J. , De Gioia Luca
Published: December 2004

A bridging carbonyl intermediate with key structural elements of the diiron sub‐site of all‐iron hydrogenase has been experimentally observed in the CN/CO substitution pathway of the {2Fe3S} carbonyl precursor, [Fe2(CO)5{MeSCH2C(Me)(CH2S)2}]. Herein we have used density functional theory (DFT) to dissect the overall substitution pathway in terms of the energetics and the structures of transition states, intermediates and products. We show that the formation of bridging CO transitions states is explicitly involved in the intimate mechanism of dicyanation. The enhanced rate of monocyanation of {2Fe3S} over the {2Fe2S} species [Fe2(CO)6{CH2(CH2S)2}] is found to rest with the ability of the thioether ligand to both stabilise a μ‐CO transition state and act as a good leaving group. In contrast, the second cyanation step of the {2Fe3S} species is kinetically slower than for the {2Fe2S} monocyanide because the Fe2 atom is deactivated by coordination of the electron‐donating thioether group. In addition, hindered rotation and the reaction coordinate of the approaching CN− group, are other factors which explain reactivity differences in {2Fe2S} and {2Fe3S} systems. The intermediate species formed in the second cyanation step of {2Fe3S} species is a μ‐CO species, confirming the structural assignment made on the basis of FT‐IR data (S. J. George, Z. Cui, M. Razavet, C. J. Pickett, Chem. Eur. J. 2002, 8, 4037–4046). In support of this we find that computed and experimental IR frequencies of structurally characterised {2Fe3S} species and those of the bridging carbonyl intermediate are in excellent agreement. In a wider context, the study may provide some insight into the reactivity of dinuclear systems in which neighbouring group on‐off coordination plays a role in substitution pathways.