Tritium levels in milk in the vicinity of chronic tritium releases.

Tritium is the radioactive isotope of hydrogen. It can be integrated into most biological molecules. Even though its radiotoxicity is weak, the effects of tritium can be increased following concentration in critical compartments of living organisms. For a better understanding of tritium circulation in the environment and to highlight transfer constants between compartments, we studied the tritiation of different agricultural matrices chronically exposed to tritium. Milk is one of the most frequently monitored foodstuffs in the vicinity of points known for chronic release of radionuclides firstly because dairy products find their way into most homes but also because it integrates deposition over large areas at a local scale. It is a food which contains all the main nutrients, especially proteins, carbohydrates and lipids. We thus studied the tritium levels of milk in chronic exposure conditions by comparing the tritiation of the main hydrogenated components of milk, first, component by component, then, sample by sample. Significant correlations were found between the specific activities of drinking water and free water of milk as well as between the tritium levels of cattle feed dry matter and of the main organic components of milk. Our findings stress the importance of the metabolism on the distribution of tritium in the different compartments. Overall, dilution of hydrogen in the environmental compartments was found to play an important role dimming possible isotopic effects even in a food chain chronically exposed to tritium.

Persistent arthralgia and its association with HIV infection in Rwanda.

A prospective study of persistent arthralgia was carried out on 331 consecutive female patients admitted to the Department of Internal Medicine of the Centre Hospitalier de Kigali in Kigali, Rwanda. The aim of this study was to determine its association with HIV-1 infection and to describe its clinical characteristics. Ten additional HIV-1-seropositive patients with this condition attending the outpatient clinic were also included in the clinical study. Persistent arthralgia was significantly more common in HIV-1-seropositive hospitalized patients (14 of 209, 6.7%) than in HIV-1-seronegative hospitalized patients (one of 122, 0.8%; p = 0.02) and had a specificity and a positive predictive value for HIV-1 infection of 99.1% and 93.3%, respectively. HIV-1-related persistent arthralgia, studied in 24 patients in early as well as late stages of HIV-1 infection, commonly affected several and mainly large joints, was mostly distributed symmetrically, and was usually relieved with nonsteroidal antiinflammatory drugs. Recurrencies were noted in eight patients. In areas highly endemic for HIV-1, persistent arthralgia should be considered a probable manifestation of HIV-1 infection, and although it is uncommon, it can be regarded as a predictor of HIV-1 infection.

Effect of somatostatin on prolactin in rainbow trout (Oncorhynchus mykiss) pituitary cells in primary culture.

To study the control of prolactin secretion in fish, an in-vitro technique using a monolayer cell culture system of rainbow trout pituitary glands was developed. Such secretion was characterized by measurement of both prolactin release and prolactin mRNA content using a trout prolactin cDNA as a probe. This cell culture technique, already used to study the regulation of gonadotrophin secretion in rainbow trout, was further validated by measuring total DNA and protein content. Both parameters appeared to be stable after 2 days of culture. Studying the effect of somatostatin (SRIF) on prolactin cells indicated that a maximal inhibitory effect (62%) was observed after 24 h of treatment. Significant inhibition of prolactin release was obtained for SRIF doses ranging from 50 nM to 1 microM. However, in the same experiment, SRIF was much more potent as an inhibitor of growth hormone release. Short-term (< 12 h) incubation with SRIF did not induce a significant change in prolactin release, whereas growth hormone release was reduced at as early as 1 h after SRIF exposure. SRIF did not have a significant effect on total prolactin content or prolactin mRNA levels, suggesting the absence of an effect on prolactin synthesis. No increase in the magnitude of the inhibitory effect of SRIF was observed when using pituitary cells from immature, mature male or mature female trout. When comparing effects on primary cultures containing cells from the whole pituitary with a prolactin cell-enriched population, SRIF appeared to have the same inhibitory effect on prolactin release, supporting a direct action of SRIF on prolactin cells. These results provide further support for SRIF being a prolactin-inhibiting factor in rainbow trout and acting as a modulator of a dominant stimulatory control of prolactin release.

Two complementary bioassays for screening the estrogenic potency of xenobiotics: recombinant yeast for trout estrogen receptor and trout hepatocyte cultures.

A relation between the chemical structure of a xenobiotic and its steroidal action has not yet been clearly established. Thus, it is not possible to define the estrogenic potency of different xenobiotics. An assessment may be accomplished by the use of different bioassays. We have previously developed a yeast system highly and stably expressing rainbow trout estrogen receptor (rtER) in order to analyze the biological activity of the receptor. The recombinant yeast system appears to be a reliable, rapid and sensitive bioassay for the screening and determination of the direct interaction between ER and estrogenic compounds. This system was used in parallel with a more elaborate biological system, trout hepatocyte aggregate cultures, to examine the estrogenic potency of a wide spectrum of chemicals commonly found in the environment. In hepatocyte cultures, the vitellogenin gene whose expression is principally dependent upon estradiol was used as a biomarker. Moreover, competitive binding assays were performed to determine direct interaction between rtER and xenobiotics. In our study, 50% of the 49 chemical compounds tested exhibited estrogenic activity in the two bioassays: the herbicide diclofop-methyl; the fungicides biphenyl, dodemorph, and triadimefon; the insecticides lindane, methyl parathion, chlordecone, dieldrin, and endosulfan; polychlorinated biphenyl mixtures; the plasticizers or detergents alkylphenols and phthalates; and phytoestrogens. To investigate further biphenyl estrogenic activity, its principal metabolites were also tested in both bioassays. Among these estrogenic compounds, 70% were able to activate rtER in yeast and hepatocytes with variable induction levels according to the system. Nevertheless, 30% of these estrogenic compounds exhibited estrogenic activity in only one of the bioassays, suggesting the implication of metabolites or different pathways in the activation of gene transcription. This paper shows that it is important to combine in vivo bioassays with in vitro approaches to elucidate the mechanism of xenoestrogen actions.

Full-length sequence and in vitro expression of rainbow trout estrogen receptor cDNA.

We previously reported the isolation of a partial cDNA clone encoding the rainbow trout estrogen receptor (rtER). A 0.4 kb 5'-end insert of this cDNA was used to screen the trout liver lambda gt10 cDNA library, and a full-length cDNA was isolated and sequenced. The principal structural characteristics of the complete coding sequence of the rtER are: first a remarkable homology of the DNA binding (C) and hormone binding (E) domains with those of other species, and second the lack of an A region, the function of which is not yet known but which is well conserved in other species. In vitro expression of the full-length rtER cDNA was carried out after transcription by T7 RNA polymerase and translation in rabbit reticulocyte lysate. Translation product analysis shows three major proteins, the largest one of which probably corresponds to the translation of the complete open reading frame of mRNA. The rtER in vitro translation products specifically bind estrogens (estradiol and diethylstilbestrol), without competition from testosterone or cortisol. The equilibrium dissociation constant (Kd), deduced from the Scatchard plot, is in the same order of magnitude as those determined heretofore in salmon livers during classical experiments. The tissue distribution of rtER mRNA shows that the same mRNA size (3.5 kb) is also present in the pituitary and hypothalamus. However, in the pituitary, a smaller sized mRNA (1.4 kb) is also detected.

Absence of direct regulation of prolactin cells by estradiol-17 beta in rainbow trout (Oncorhynchus mykiss).

The effects of estradiol-17 beta (E2) implants on plasma prolactin (PRL) concentrations, pituitary PRL content and pituitary PRL mRNA levels were examined in rainbow trout (Oncorhynchus mykiss). Intact immature fish treated with 1 mg estradiol-17 beta did not show significant changes in both PRL mRNA levels and pituitary PRL content after 3 days of treatment. In a similar experiment, no changes were observed in plasma PRL levels followed during 7 days. Similarly, lack of estradiol-17 beta effect on plasma PRL levels and on final PRL pituitary content was observed in ovariectomized female rainbow trout treated during 48 days with 25 mg estradiol-17 beta and in mature male fish over a 3-week treatment period. Localization of estradiol receptor (ER) mRNAs in the pituitary was carried out by Northern blot analysis using a full-length rainbow trout estrogen receptor (rtER) cDNA as a probe. The rostral pars distalis of the pituitary which contained mostly PRL cells showed the lower amount of rtER mRNA when compared to other parts of the pituitary. Moreover, two mRNAs of different size (3.5 and 1.4 kb) were detected in different parts of the pituitary. Further hybridization experiments using probes containing part of the rtER cDNA (E domain or C and D domains) indicated that the small-sized mRNA (1.4 kb) probably encodes a truncated ER protein lacking hormone binding domain or an ER-related protein. Thus, only the 3.56 kb mRNA appeared to be involved in the regulation of pituitary function by estradiol. In situ hybridization analysis allowed a more precise localization of this rtER mRNA in the pituitary.(ABSTRACT TRUNCATED AT 250 WORDS)

Localization of the estradiol receptor mRNA in the forebrain of the rainbow trout.

In situ hybridization was used to localize the cells that express the estradiol receptor gene (ER) in the forebrain (hypothalamus, preoptic area, telencephalon) of the rainbow trout (Oncorhynchus mykiss). Both sense and anti-sense [35S]UTP-labeled single-stranded RNA probes were generated from the estradiol binding domain of the ER cDNA. The sense probe was used to evaluate the background of the hybridization reaction. In the forebrain, specific signal appeared in three areas: the posterior hypothalamus, the preoptic area, and the ventral telencephalon. Our localization correlates with [3H]estradiol binding studies in other teleost species. In the pituitary, we observed a weak signal when compared to the signal observed in the forebrain (about ten grains/cell in the pituitary against 35 grains/cell in the posterior hypothalamus). A significant difference was also observed between the intensity of labeling per cell when different forebrain nuclei were compared. We provide here evidence for a tissue-specific regulation of the ER mRNA levels in the trout hypothalamo-pituitary axis.

The nosology-taxonomy of recent-onset arthritis: the experience of early-arthritis clinics.

OBJECTIVE: To compare the conclusions of studies addressing the outcome of early-arthritis cohorts. METHODS: The methodologies of previous reports on early-arthritis cohorts were examined, and their results and conclusions were compared. RESULTS: Thirty-four reports on 23 cohorts of early arthritis were found. The methodology was poor in most studies, with numerous inclusion and exclusion biases, frequently short follow-up periods, and a lack of precision about the rationale for diagnosis. However, similar conclusions were reached on several points: a large number of cases of early arthritis remained undifferentiated and/or resolved spontaneously, about 80% of cases initially classified as undifferentiated or rheumatoid arthritis retained this diagnosis during follow-up, and the incidence of psoriatic arthritis in most studies was similar (2% to 4%). Conversely, there were striking discrepancies among studies concerning the frequency of crystal arthropathies (0% to 18%), spondyloarthropathy (1% to 33%) and rheumatoid arthritis (15% to 47%). CONCLUSIONS: There appears to be a lack of agreement among researchers about the nosology and/or taxonomy of many cases of mild arthritis, despite the existence of classification criteria. RELEVANCE: Recognition of cultural bias in the diagnosis of early arthritis could be a prerequisite for the optimization of new sets of criteria for the diagnosis of early rheumatoid arthritis and spondyloarthropathy.

Expression of a human fetal anti-DNA antibody idiotype BEG-2 beta in the families of patients with rheumatoid arthritis.

BEG-2 is a monoclonal antibody produced by the human-human hybridoma technique from a 12 weeks old human fetus. A polyclonal antiserum was raised in an (NZW x Half-lop hybrid) rabbit against BEG-2 and the anti-BEG-2 anti-idiotype was purified and characterised. Using this rabbit reagent the expression of the BEG-2 beta idiotype was analysed in 12 patients with active rheumatoid arthritis and their close family members (n = 54). Twenty five sera from healthy controls were analysed to establish a normal range. Ten of 12 patients (83%) with rheumatoid arthritis expressed the BEG-2 idiotype as well as 11 of 54 healthy unaffected relatives (20%).

Anti-Epstein-Barr virus-nuclear antigen-1, -2A and -2B antibodies in rheumatoid arthritis patients and their relatives.

We have examined serum antibodies to Epstein-Barr virus Nuclear Antigen (EBNA)-1, -2A and -2B, in addition to antibodies to viral capsid antigen and early antigen in 100 rheumatoid arthritis patients and 50 of their relatives. Using indirect immunofluorescence on transfected cells and Western-blot technique, we have found increased frequency and titres of antibodies to EBNA-2B in patients and, to a lesser degree, in their family members, whereas other anti-Epstein-Barr virus antibodies appeared to be similar to controls. Cross-inhibition experiments were carried out and show that antibodies to EBNA-2A are distinct from those to -2B, and vice versa.

The relationship of anti-endothelial cell antibodies to anti-phospholipid antibodies in patients with giant cell arteritis and/or polymyalgia rheumatica.

Sera from patients with giant cell arteritis and/or polymyalgia rheumatica were tested for the presence of IgG, IgM and IgA antibody to endothelial cells (AEC), cardiolipin (ACL) and phosphatidylethanolamine (APE) using enzyme-linked immunosorbent assays. There were strong correlations between ACL and APE, but also between AEC and ACL IgM (p < 0.02) and between AEC and APE IgA (p < 0.003). Inhibition of AEC binding was achieved by absorption onto EC, but ACL and APE binding was also significantly reduced. In contrast, the binding of AEC antibody could not be inhibited by incubation with CL. Our data suggest that AEC constitute a heterogeneous population of autoantibodies.

Abnormal distribution of CD45 isoforms expressed by CD4+ and CD8+ T cells in rheumatoid arthritis.

CD45RO+ T cells are referred to as memory or helper-inducer while CD45RA+ T cells are regarded as naive or suppressor-inducer T cells. The former population predominates in the peripheral blood and even more in the synovial fluid of patients with rheumatoid arthritis, to the expense of the latter population. Within the CD45RB+ compartment, there appears to be more of the fully-differentiated than of the early-differentiated CD4+ T cells. In spite of the fact that these lymphocytes are close to undergoing apoptosis, this programmed cell death is inhibited in the rheumatoid synovium.

An assessment of the role of domain F and PEST sequences in estrogen receptor half-life and bioactivity.

The estrogen receptor (ER) is a rapidly turning over protein, with a half-life of ca. 3-4 h in estrogen target cells. Sequence analysis of the human ER reveals a putative PEST sequence, sequences rich in proline (P), glutamic acid (E), serine (S) and threonine (T), in the carboxy-terminal F domain of the protein. Since PEST sequences have been implicated in the rapid turnover of some proteins, we have used site-directed mutagenesis to investigate the role of the F region containing PEST residues in the stability and bioactivity of the receptor. A truncated form of ER lacking the last 41 amino acids of the protein and encompassing the PEST sequences (amino acids 555 to 567) was made by mutagenesis of the ER cDNA. Pulse-chase experiments, involving immunoprecipitation of [35S]methionine/[35S]cysteine labeled receptors or of receptors covalently labeled with tamoxifen aziridine followed by gel electrophoresis, were used to determine the half-life of the wild-type and truncated ERs. These experiments showed that the turnover rate of the receptors expressed in Chinese hamster ovary and monkey kidney (COS-1) cells was 3 to 5 h and that elimination of the PEST residues did not have a significant effect on the degradation rate of the protein. Moreover, deletion of the last 41 amino acids (F domain) of the ER did not affect transactivation ability, ligand binding affinity, or the phosphorylation pattern of the receptor. Therefore, the role of domain F in ER function remains unclear, but it is not a determinant of the relatively rapid rate of ER turnover in cells.

Antiestrogens: mechanisms and actions in target cells.

Antiestrogens, acting via the estrogen receptor (ER) evoke conformational changes in the ER and inhibit the effects of estrogens as well as exerting anti-growth factor activities. Although the binding of estrogens and antiestrogens is mutually competitive, studies with ER mutants indicate that some of the contact sites of estrogens and antiestrogens are likely different. Some mutations in the hormone-binding domain of the ER and deletions of C-terminal regions result in ligand discrimination mutants, i.e. receptors that are differentially altered in their ability to bind and/or mediate the actions of estrogens vs antiestrogens. Studies in a variety of cell lines and with different promoters indicate marked cell context- and promoter-dependence in the actions of antiestrogens and variant ERs. In several cell systems, estrogens and protein kinase activators such as cAMP synergize to enhance the transcriptional activity of the ER in a promoter-specific manner. In addition, cAMP changes the agonist/antagonist balance of tamoxifen-like antiestrogens, increasing their agonistic activity and reducing their efficacy in reversing estrogen actions. Estrogens, and antiestrogens to a lesser extent, as well as protein kinase activators and growth factors increase phosphorylation of the ER and/or proteins involved in the ER-specific response pathway. These changes in phosphorylation alter the biological effectiveness of the ER. Multiple interactions among different cellular signal transduction systems are involved in the regulation of cell proliferation and gene expression by estrogens and antiestrogens.

Prevalences of rheumatoid arthritis in Roman Catholic nuns and the general female population in Brittany, France: a pilot study.

OBJECTIVE: To evaluate the influence of lifestyle factors on the prevalence of rheumatoid arthritis (RA) by comparing Roman Catholic nuns and the general female population. METHOD: RA prevalence in the general population was evaluated using a standardized telephone survey in 1857 homes taken at random. Individuals who reported an inflammatory joint disease were contacted by a rheumatologist of our unit, missing data were collected from the general practitioner or rheumatologist with the patient's permission, and if necessary a physical examination was done by a rheumatologist. The 9 largest Roman Catholic nun communities in Brittany were screened using the same standardized questionnaire administered face-to-face; nuns who reported an inflammatory joint disease were interviewed and examined by rheumatologists. In both populations, RA was diagnosed when (1) the rheumatologist of our unit who interviewed the patient considered the RA classification criteria positive and (2) the rheumatologist who examined the patient gave a diagnosis of RA independently from RA classification criteria. RESULTS: Data were available for 1706 adult females in the general population and 721 nuns. Of the 20 nuns who reported RA or polyarthritis, 11 received a diagnosis of RA (prevalence 1.52%). The prevalences adjustedfor the French population after 40 years were 1.66% (95% confidence interval, 0.84-2.44) and 1.33 (0.27-2.40) among the nuns and the general female population, respectively. CONCLUSION: Although our nun population was too small for definite conclusions, we found no evidence of a difference in RA prevalence among nuns and the general female population in Brittany.

The antiperinuclear factor. II. Variability of the perinuclear antigen.

Since the antiperinuclear factor (APF) test on human buccal cells is rather unpredictable, we have investigated the possible factors determining the expression of appropriate antigens by the cells. We failed to find any relationship of the expression of perinuclear antigens to the donor's smoking habits, the degree of contamination with saprophytic bacteria, the presence or absence of blood group substances in saliva, or the titers of serum antibodies to Epstein-Barr virus. Family studies were also performed to further elucidate a genetic predisposition to the expression of the APF antigen.

The antiperinuclear factor. I. Clinical and serologic associations.

The antiperinuclear factor (APF) test raises two main problems: the unpredictability of the cells used as substrate and the difficulty in expressing the results. We propose that 10% of the cells have to be stained by a given serum in order for it to be considered positive. APF were found to be present in 76% of rheumatoid arthritis (RA) patients, 3% of healthy controls and occasionally in disease controls. The production of APF was significantly (p less than 0.01) related to the presence of rheumatoid factor in RA, and IgG antibody was predominant in the APF test.

Tiopronine-induced reduction of rheumatoid factor functional affinity and asialylated IgG in patients with rheumatoid arthritis.

Serum and synovial fluid (SF) from 16 rheumatoid arthritis patients were evaluated before and after a 2-month treatment with tiopronine (TP). The levels of rheumatoid factor (RF) declined, as did the functional affinity of the remaining RF (p less than 0.01 in serum and p less than 0.05 in SF). Concomitant restoration of the sialylation of IgG was observed (p less than 0.05 in serum and SF). Following an initial increase, a significant reduction was observed in the level of soluble interleukin-2 receptors in serum (p less than 0.05) and SF (p less than 0.05) after treatment with TP.

Antiperinuclear factor in Sjögren's syndrome in the presence or absence of rheumatoid arthritis.

Serum from 91 patients with Sjögren's syndrome (SS) was examined for antiperinuclear factor (APF). Nine out of 29 patients with SS alone (31.0%) were positive as opposed to 27 out of 40 with SS + rheumatoid arthritis (67.5%) and to 3 out of 22 with SS + other autoimmune disorders (13.6%). A clear relationship was established between APF and agglutinating or non-agglutinating rheumatoid factor.

Confidence in the diagnosis of early spondylarthropathy: a prospective follow-up of 270 early arthritis patients.

OBJECTIVE: To study the confidence of office-based rheumatologists (OBR) and a college of 5 experts in their diagnosis of spondylarthropathy (SpA) for early arthritis after more than 2 years of follow-up; to determine whether at that time the degree of confidence was improved by the fulfilment of the ESSG criteria. METHODS: 270 patients with early-onset (< 1 year) arthritis were prospectively followed-up for 29+/-11 months. At the final examination, OBR and the college of 5 experts rated their confidence in the diagnosis of SpA on a 0-10 analogue scale and on a 1-4 Likert scale, respectively. RESULTS: After 29+/-11 months OBR had classified 56 patients (21%) as SpA, while a collegial diagnosis of probable (N = 32) or certain SpA (N = 14) was made for 46 patients (17%). At the final examination OBR confidence in their diagnosis (gold standard) was only 6.7+/-2.4 for all 56 cases of SpA. The cumulative fulfilment of ESSG criteria for SpA after 29+/-11 months correlated with the confidence of OBR and the experts in SpA, but improved only slightly the final confidence of OBR (7.1+/-2.3 versus 6.7+/-2.4 for all 56 SpA). Similarly, OBR confidence for the 18/56 SpA patients positive for HLA-B27 was only 7.1+/-2.0. Only 21 of these 56 patients were considered as SpA at baseline, although 37/56 (66%) had fulfilled ESSG criteria since thefirst examination. CONCLUSION: This study indicates a probable lack of consensus on the nosology of early SpA and the limited help provided by the ESSG criteria to differentiate early SpA from otherforms of arthritis at baseline.