[Hormonal contraception and vascular risk: CNGOF Contraception Guidelines]. / Contraception hormonale et risque vasculaire. RPC Contraception CNGOF.

Venous thromboembolism and arterial ischemic events are the main deleterious diseases associated with the use of combined hormonal contraceptives (CHC). Even though their composition has been substantially improved, the vascular risk persists with the most recent CHCs use. If the vascular risk associated with CHCs containing 50µg EE is significantly higher than with those containing less than 50µg, there is no evidence that the CHCs containing either 30 or 20µg of EE induce different venous risks. CHC containing gestodene, desogestrel, drospirenone or cyproterone acetate are associated with a higher risk of venous thrombosis compared with levonorgestrel-containing CHCs. CHC containing norgestimate are associated with similar venous thrombosis risk than CHC containing levonorgestrel. Venous thrombosis risk of non-oral routes of administration of CHC appears to be equivalent to the risk of CHC containing gestodene or desogestrel, but this result is based on a small number of epidemiological studies. Before prescribing a CHC, it is important to determine all vascular risk factors. Family history of ischemic arterial event or venous thromboembolism disease should be routinely sought before any CHC prescription. All CHCs are contraindicated in women with biological thrombophilia, in women with combined vascular risk factors, in women with first-degree family history of arterial or venous event (under age 50) as well as in women suffering of migraine with aura. Progestin-only contraceptives are not associated with vascular risk (arterial or venous) outside of medroxyprogesterone acetate. In women with higher vascular risk, progestin-only contraceptives (administered by oral, sous-cutaneous or intra-uterine routes) can be prescribed.

A family-based strategy to identify genes for diabetic nephropathy.

Diabetic nephropathy (DN) clusters in families and specific ethnic groups, suggesting a genetic basis of disease transmission. Identification of DN susceptibility loci should reveal new therapeutic targets but requires accurate phenotyping. A powerful family-based strategy, which is novel to the pursuit of nephropathy genes in type 2 diabetes, is being used to collect a sample for candidate gene and genome scan analyses. Sib pairs that include DN index cases plus (1) sibs concordant for type 2 diabetes and DN (affected sib pairs [ASPs]) and (2) sibs concordant for type 2 diabetes but discordant for DN (discordant sib pairs [DSPs]) are targeted specifically for recruitment. Type 2 diabetes and DN phenotype criteria for index cases include diabetes onset after 38 years of age, duration 10 years or longer, no initial insulin treatment, diabetic retinopathy, end-stage renal disease (ESRD), and history of nephrotic proteinuria. ESRD patients were screened by questionnaire and medical record review (n = 2114). Of 666 patients with ESRD secondary to DN, 227 had a family history of ESRD, 150 had a living diabetic sib, and 124 families were enrolled. Sixty-five families, with 86 diabetic relative pairs (69 sibs, 17 children), have been completely phenotyped. If nephropathy in diabetic sibs is defined as albuminuria greater than 0.3 g/24 h, 31 ASPs and 26 DSPs (diabetic sib with albuminuria <0.3 g/24 h) were identified. Applying more stringent criteria, only 12 ASPs (sib with diabetes >10 years, diabetic retinopathy, and nephrotic proteinuria) and 9 DSPs (sib with diabetes >10 years and normal urine albumin excretion) were identified. Extrapolating from the number of subjects recruited using stringent phenotyping criteria, nearly 10,000 ESRD patients are required for screening to achieve adequate statistical power for linkage analysis (80% power to detect locus-specific relative risk of 2.2 at a lod score of 3.0). Careful phenotyping requires a large recruitment effort but is necessary to reduce population heterogeneity, a strategy that increases the likelihood of identifying DN loci.

Sp1 binds to the rat luteinizing hormone beta (LHbeta) gene promoter and mediates gonadotropin-releasing hormone-stimulated expression of the LHbeta subunit gene.

The hypothalamic hormone gonadotropin-releasing hormone (GnRH) plays a critical role in reproductive function by regulating the biosynthesis and secretion of the pituitary gonadotropins. Although it is known that GnRH induces luteinizing hormone beta (LHbeta) gene transcription, the mechanisms by which this occurs remain to be elucidated. We have shown previously that GH3 cells transfected with the rat GnRH receptor cDNA (GGH3-1' cells) support the expression of a cotransfected fusion gene composed of 797 base pairs of rat LHbeta gene 5'-flanking sequence and the first 5 base pairs of the 5'-untranslated region fused to a luciferase reporter (-797/+5LHbetaLUC) and respond to a GnRH agonist with a 10-fold stimulation of activity. Furthermore, we have shown that DNA sequences at -490/-352 confer GnRH responsiveness to the rat LHbeta gene. We have now identified two putative binding sites for Sp1, a three-zinc-finger transcription factor, within this region. Using electrophoretic mobility shift assay, DNase I footprinting, and methylation interference assays, we demonstrate that Sp1 can bind to these sites and that Sp1 is responsible for DNA-protein complexes formed using GGH3-1' and alphaT3-1 nuclear extracts. Mutations of the Sp1 binding sites, which block binding of Sp1, blunt the stimulation of the LHbeta gene promoter by GnRH. These data define GnRH-responsive elements in the LHbeta 5'-flanking sequence and suggest that Sp1 plays an important role in conferring GnRH responsiveness to the LHbeta subunit gene.

Identification of cis-acting deoxyribonucleic acid elements that mediate gonadotropin-releasing hormone stimulation of the rat luteinizing hormone beta-subunit gene.

GnRH plays a critical role in reproductive development and function by regulating the biosynthesis and secretion of the pituitary gonadotropins, LH and FSH. Although it is known that GnRH induces gonadotropin subunit gene transcription, the mechanism by which this occurs has not been elucidated. Studies have been hindered by the lack of available cell lines that express the LH and FSH subunit genes and respond to GnRH. We have transfected the rat pituitary GH3 cell line with the rat GnRH receptor complementary DNA. These cells, when cotransfected with regulatory regions of the LH or FSH subunit genes fused to a luciferase reporter gene, respond to GnRH with an increase in promoter activity comparable to that seen in primary rat pituitary cells. In this study, we have used this cell model to identify cis-acting elements of the LHbeta gene that mediate stimulation by GnRH. Analysis of a series of 5'-deletion and internal deletion constructs has revealed two regions of the rat LHbeta gene promoter involved in mediating the response to GnRH, region A (-490/-352) and region B (-207/-82). Fusion of region A upstream of a heterologous minimal promoter linked to the luciferase gene conferred GnRH responsiveness to the promoter, whereas region B did not. However, the presence of both regions A and B conferred a greater GnRH response than region A alone. Electrophoretic mobility shift assay revealed the presence of a protein(s) binding to region A using GH3 as well as alphaT3-1 nuclear extracts. Thus, region A (-490/-352) confers GnRH responsiveness to the LHbeta subunit gene and binds to a protein(s) present in pituitary cell lines. DNA sequences in region B (-207/-82) also contribute to GnRH responsiveness. The identification of putative GnRH response elements in the rat LHbeta gene promoter will aid in elucidation of the mechanisms of regulation of gene expression by GnRH.

Differential use of signal transduction pathways in the gonadotropin-releasing hormone-mediated regulation of gonadotropin subunit gene expression.

The regulation of LH and FSH subunit gene expression is under the control of GnRH. Physiological changes in the frequency of pulsatile GnRH release from the hypothalamus result in differential stimulation of alpha-, LHbeta-, and FSHbeta-gene expression. Previous studies indicate that the GnRH receptor couples to G proteins of the G(q/11) family, with phosphoinositide turnover and its resultant increase in intracellular calcium concentration and protein kinase C (PKC) activation, to stimulate secretion of LH and FSH. However, the molecular mechanisms by which GnRH mediates its transcriptional effects remain largely unknown. We used GH3 cells, constitutively expressing the rat GnRH receptor (GGH(3)-1' cells) and transiently transfected with a luciferase reporter gene controlled by either the alpha, LHbeta, or FSHbeta gene regulatory region (alphaLUC, LHbetaLUC, and FSHbetaLUC, respectively), to examine the roles of several signal transduction pathways in the GnRH-mediated stimulation of gonadotropin subunit gene expression. Activation of PKC by phorbol, 12-myristate, 13-acetate resulted in an increase in the luciferase activity of all three gonadotropin subunit gene reporter constructs. Phorbol, 12-myristate, 13-acetate had a greater stimulatory effect, relative to the maximal stimulation with GnRH, for the beta-subunit genes than for the alpha-subunit gene. Depletion of PKC, or inhibition of PKC by GF109203X, demonstrated that PKC-dependent pathways play a larger role in the GnRH-mediated transcriptional control of the LHbeta- and FSHbeta-genes than the alpha-subunit gene. In contrast, an L-type calcium channel agonist, Bay K 8644, was able to stimulate alphaLUC but not LHbetaLUC or FSHbetaLUC. Nimodipine, an L-type calcium channel antagonist, had a larger inhibitory effect on the GnRH response of alphaLUC, relative to LHbetaLUC or FSHbetaLUC. We conclude from these results that the differential regulation of gonadotropin subunit gene expression by GnRH is caused, in part, by differential use of signal transduction pathways, activated upon GnRH binding.

[Treatment of idiopathic pulmonary fibrosis with colchicine]. / Tratamiento de la fibrosis pulmonar idiopática con colchicina.

BACKGROUND: Experimental and clinical evidences suggest that colchicine can be effective in the treatment of patients with idiopathic pulmonary fibrosis. AIM: To assess the effect of colchicine in the treatment of idiopathic pulmonary fibrosis. PATIENTS AND METHODS: Patients with clinically diagnosed idiopathic pulmonary fibrosis were treated with colchicine in doses of 0.5 to 1 mg/day, according to tolerance and followed for periods ranging from 7 to 40 months. The clinical and radiological score reported by Watters et al was used for the longitudinal assessment of patients. Maintenance or improvement in forced vital capacity and maintenance or decrease in alveolar arterial O2 gradient during follow up, were considered as positive therapeutic responses. RESULTS: Seventeen patients (10 male, aged 61 to 81 years old) were studied. Their basal score for dyspnea was 5.8 (over 20), for the chest X ray examination was 2.4 (over 3) and for CT scan was 2.8 (over 3). Basal FVC was 77% of predicted value (range 51-108%), basal FEV, was 82% (range 59-117%) and FEV1/FVC was 0.82 (range 0.68-0.95). PaO2 at rest was 78 mm Hg (ranges 63-97). Alveolar-arterial PO2 gradient was 16 mm Hg (range 5-31.6) at rest and 31 mm Hg (range 5.7-51.4) after exercise. Six patients (35%) had a positive response to therapy. CONCLUSIONS: The response rates of these patients to colchicine are at least similar to those obtained with steroids, but with less side effects.

A mechanism for the differential regulation of gonadotropin subunit gene expression by gonadotropin-releasing hormone.

The hypothalamic hormone gonadotropin-releasing hormone (GnRH) is released in a pulsatile fashion, with its frequency varying throughout the reproductive cycle. Varying pulse frequencies and amplitudes differentially regulate the biosynthesis and secretion of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) by pituitary gonadotropes. The mechanism by which this occurs remains a major question in reproductive physiology. Previous studies have been limited by lack of available cell lines that express the LH and FSH subunit genes and respond to GnRH. We have overcome this limitation by transfecting the rat pituitary GH3 cell line with rat GnRH receptor (GnRHR) cDNA driven by a heterologous promoter. These cells, when cotransfected with regulatory regions of the common alpha, LH beta, or FSH beta subunit gene fused to a luciferase reporter gene, respond to GnRH with an increase in luciferase activity. Using this model, we demonstrate that different cell surface densities of the GnRHR result in the differential regulation of LH and FSH subunit gene expression by GnRH. This suggests that the differential regulation of gonadotropin subunit gene expression by GnRH observed in vivo in rats may, in turn, be mediated by varying gonadotrope cell surface GnRHR concentrations. This provides a physiologic mechanism by which a single ligand can act through a single receptor to regulate differentially the production of two hormones in the same cell.

A look at women's health issues.

Women's health issues have become the focus of much debate. A critical question in that debate centers on gender bias in medical research, which may result in bias in treatment. In this article, several disease processes are examined as they relate to women today. The areas of uncertainty in treatment, as well as areas of needed research, are highlighted.

A comparison of dual-energy digital radiography and screen-film imaging in the detection of subtle interstitial pulmonary disease.

An ROC study is described which compares the performance of three types of images--conventional screen-film, single-energy digital and dual energy bone cancelled (soft tissue) digital--in detecting subtle interstitial pulmonary disease. Marginally detectable nodular and reticulonodular patterns (12 different patterns of each) were superimposed over the lungs of a frozen human chest phantom to simulate the clinical situation. The digital images were formatted on film at full size (ie, 35 cm X 43 cm). A total of 156 images (52 of each type, of which 28 were normal and 24 had simulated pathology) were used in the study and read by five experienced chest radiologists. Using a paired t-test, the areas under the individual ROC curves were compared for three combinations of images--single-energy digital and conventional, soft tissue digital and conventional, and soft tissue and single-energy digital. No statistically significant difference was observed between the conventional and single-energy digital images. The readers performed better with both conventional and single-energy digital images than with the soft tissue digital images at statistically significant levels (P = 0.05 for conventional vs. soft tissue digital and P = 0.02 for single-energy digital vs. soft tissue digital). The results suggest that there is no advantage in employing dual-energy soft tissue images to assist in diagnosing interstitial disease in the clinical setting. They also suggest that spatial resolution requirements are less demanding in digital chest systems that obtain scatter-free images than in digital systems utilizing conventional scatter control techniques.

Relative incidence of intraoral pemphigus vulgaris, mucus membrane pemphigoid and lichen planus.

A retrospective analysis of patients was conducted in order to determine the relative incidence of oral pemphigus vulgaris, mucus membrane pemphigoid and lichen planus. The analysis yielded a relative incidence of 1:2:15 of pemphigus vulgaris, mucus membrane pemphigoid and lichen planus patients. The dangers of comparing and relating incidence statistics from one study to other studies were demonstrated.

Mineral content of bone: measurement by energy subtraction digital chest radiography.

Several methods are presently available for measuring the mineral content of bone. Those in widespread use include dual-photon absorptiometry and quantitative CT. The feasibility of using dual-energy digital chest radiography for determination of the mineral content of posterior ribs on digital chest images was studied by using a prototype unit. The results showed a significant difference in the mineral density of the posterior ribs of control subjects and those of patients who had osteoporosis (251.1 +/- 36 mg Ca2+/cm2 of rib vs 158.8 +/- 48 mg Ca2+/cm2, p less than or equal to .01) and a close correlation with values obtained by dual-photon absorptiometry of the lumbar spine (r = .77). The results suggest that this technique can provide an accurate assessment of the presence or absence of osteoporosis.

Dual-energy digital radiographic quantification of calcium in simulated pulmonary nodules.

The presence or absence of calcium in solitary pulmonary nodules may indicate whether a nodule is benign or malignant. Because current techniques for measuring the amount of calcium in these nodules are unsatisfactory, a study was carried out to assess the capability of dual-energy digital chest radiography to identify and quantify the calcium content of simulated pulmonary nodules of known calcium content. Measurements were carried out on 280 nodules of various sizes and calcium content that were placed within the lungs of a frozen human-chest phantom. A new calcium quantification technique that uses a parallelogram was developed to eliminate the problem of nodule superimposition over ribs. Nodules containing more than 35 mg of calcium per square centimeter (i.e., 7, 30, 60, and 110 mg of calcium for spherical nodules 0.5, 1.0, 1.5, and 2.0 cm in diameter) were measured with a high degree of accuracy and reasonable precision. Dual-energy digital radiography is a simple and accurate method of measuring the calcium content of solitary pulmonary nodules in humans.

Simulated pulmonary nodules: detection with dual-energy digital versus conventional radiography.

Performance of a prototype dual-energy digital chest radiography unit in detecting calcified and noncalcified simulated pulmonary nodules was compared with that of a highly optimized, conventional system. Nodules ranging in size (0.5, 1.0, and 1.6 cm), in number (five to 11), and in calcium content (0-25 mg) were superimposed over the lungs of a frozen, unembalmed, human chest phantom. For each technique, six observers examined 50 posteroanterior projections with different randomized nodule locations. Detection consisted of locating and assigning a level of confidence to each perceived nodular opacity. The resulting plots of the true-positive fraction versus the mean number of false-positive calls per projection indicate that for both calcified and noncalcified nodules, the digital unit performed significantly better (P less than .01).

Calcification in pulmonary nodules: detection with dual-energy digital radiography.

The authors undertook a clinical study to determine the accuracy of dual-energy digital radiography in revealing nodule calcification because calcification in a pulmonary nodule almost excludes the possibility of malignancy. Over a 6-month period, 61 patients with pulmonary nodules (less than or equal to 3 cm) or masses (greater than 3 cm) were examined on a prototype scanned projection unit using a dual-energy detector. In 49 of 61 patients, nodules were noncalcified, and in 12, they were calcified. In 57 patients, the benignancy or malignancy of nodules was established beyond reasonable doubt by pathologic confirmation in 38 and by strong inference in 19 (four patients with noncalcified solitary pulmonary nodules either refused further investigation or surgery or their follow-up was too short to permit exclusion of malignancy). Dual-energy radiography was found to be highly accurate in assessing the presence or absence of calcification in pulmonary nodules and thus in determining their benignancy or possible malignancy.

Potential value of digital radiography. Preliminary observations on the use of dual-energy subtraction in the evaluation of pulmonary nodules.

The radiographic demonstration of calcification in a solitary pulmonary nodule renders the possibility of malignancy extremely unlikely, although rare exceptions have been reported. Conventional roentgenograms and tomograms sometimes provide inconclusive evidence although CT can be highly accurate in both identifying and quantifying calcium content. An alternative method is dual-energy subtraction utilizing scanned projection digital radiography. Forty-one patients with solitary (occasionally multiple) pulmonary nodules were examined with the technique, employing second-generation fan-beam equipment: 28 nodules or masses were noncalcified and 13 calcified. Of the former, 20 were pathologically proved, 16 being malignant and 4 benign (2 granulomas, 2 bronchiectasis); in 3 of the remaining 8, a presumptive diagnosis was reasonably certain (1 granuloma, 2 metastases), while in 5 the diagnosis was not made. In 8 of the 13 calcified lesions, the diagnosis can reasonably be regarded as confirmed as granulomas; 5 are being followed up with that presumptive diagnosis.

3-Mercaptopropionic acid, a potent inhibitor of fatty acid oxidation in rat heart mitochondria.

The effects of several short-chain mercapto acids on the rate of respiration supported by either palmitoylcarnitine, octanoate, or pyruvate was studied with coupled rat heart mitochondria. 3-Mercaptopropionic acid was found to be a potent inhibitor of respiration sustained by palmitoylcarnitine or octanoate, whereas under identical conditions respiration with pyruvate as a substrate was unaffected. 2-Mercaptoacetic acid also inhibits palmitoylcarnitine-supported respiration, but only at much higher concentrations of the inhibitor. 2-Mercaptopropionic acid has virtually no effect. Incubation of mitochondria with 3-mercaptopropionic acid did not cause the irreversible inactivation of any beta-oxidation enzyme. Since 3-mercaptopropionic acid did not inhibit beta-oxidation in uncoupled mitochondria, it appears that this compound must first be metabolized in an energy-dependent reaction before it becomes inhibitory. 3-Mercaptopropionyl-CoA and three of its S-acyl derivatives, all of which are likely mitochondrial metabolites of 3-mercaptopropionic acid, were tested for their capacity to inhibit the individual enzymes of beta-oxidation. 3-Mercaptopropionyl-CoA inhibits only acyl-CoA dehydrogenase, whereas S-myristoyl-3-mercaptopropionyl-CoA inhibits reversibly several beta-oxidation enzymes. All observations together lead us to suggest that the inhibition of beta-oxidation by 3-mercaptopropionic acid in coupled rat heart mitochondria is most likely a consequence of the reversible inhibition of acyl-CoA dehydrogenase by long-chain S-acyl-3-mercaptopropionyl-CoA thioesters and possibly by 3-mercaptopropionyl-CoA.