On the relationship between farmland biodiversity and land-use intensity in Europe.

Worldwide agriculture is one of the main drivers of biodiversity decline. Effective conservation strategies depend on the type of relationship between biodiversity and land-use intensity, but to date the shape of this relationship is unknown. We linked plant species richness with nitrogen (N) input as an indicator of land-use intensity on 130 grasslands and 141 arable fields in six European countries. Using Poisson regression, we found that plant species richness was significantly negatively related to N input on both field types after the effects of confounding environmental factors had been accounted for. Subsequent analyses showed that exponentially declining relationships provided a better fit than linear or unimodal relationships and that this was largely the result of the response of rare species (relative cover less than 1%). Our results indicate that conservation benefits are disproportionally more costly on high-intensity than on low-intensity farmland. For example, reducing N inputs from 75 to 0 and 400 to 60kgha-1yr-1 resulted in about the same estimated species gain for arable plants. Conservation initiatives are most (cost-)effective if they are preferentially implemented in extensively farmed areas that still support high levels of biodiversity.

[Assessment of postural control and balance in persons with temporomandibular disorders: A systematic review]. / Valoración del control postural y del equibrio en personas con trastornos temporomandibulares: revisión sistemática.

The stomatognathic system is the anatomo-functional unit of the cranio-cervico-facial region. Some dysfunctions affect its motor control. The aim of this study was to analyse the clinical usefulness of the various scales and instruments used in the assessment of postural control in people with temporomandibular disorders. A systematic review was carried out by 2independent reviewers in the PubMed, Medline, Ebsco, Science Direct and PEDro databases, selecting observational studies published between January 2006 and March 2017. The risk of bias and methodological quality was analysed following Cochrane indications and the Downs and Black quality scale. Ten studies were included, of which 9used computerised platforms, one added photogrammetry and one used electromyography. Seven studies were classified as moderate quality and 3as low quality. Posturography was the most widely used assessment instrument. Methodological differences did not allow determination of their clinical implications or the relationship between balance and the presence of temporomandibular disorders.

Evidence for selective accumulation of intrathyroidal T lymphocytes in human autoimmune thyroid disease based on T cell receptor V gene usage.

We have investigated the T cell receptor V alpha and V beta gene family usage by T lymphocytes infiltrating affected thyroids in patients with autoimmune thyroid disease. We show that the intrathyroidal T lymphocytes from patients (n = 6) with autoimmune thyroid disease display a widespread usage of V beta gene families with an average of 14.4/19 V beta gene families similar to the peripheral T lymphocytes of the same patients. Because we recently reported that the utilization of V alpha gene families is markedly reduced within these mitogen-stimulated intrathyroidal T cell populations, as well as within intact tissue from similar patients (n = 4) (overall mean of 4.0/18 families detected), these results indicate that in thyroids of patients with autoimmune thyroid disease the lymphocytes are selectively accumulating based on their V alpha rather than V beta elements. This preferential hTcR V alpha and widespread V beta gene usage was not mimicked in most 7-d autologous mixed lymphocyte reactions using non-T cell stimulators (n = 6) or EB-virus immortalized autologous B cell lines (n = 3). Hence, the selective V gene utilization by intrathyroidal T cells is likely to be secondary to multiepitopic thyroidal autoantigens activating thyroid infiltrating T cells or to the presence of a superantigenlike thyroidal self-antigen, capable of determining a selective infiltration or activation of a variety of T lymphocytes on the basis of their V alpha gene usage.

Endogenous antigen presentation by autoantigen-transfected Epstein-Barr virus-lymphoblastoid cells. I. Generation of human thyroid peroxidase-reactive T cells and their T cell receptor repertoire.

To develop a model for endogenous thyroid autoantigen presentation, we transfected EBV-transformed B lymphoblastoid cell lines (EBV-LCL), established from patients with autoimmune thyroid disease and normal controls, with cDNA for the human thyroid autoantigen thyroid peroxidase (hTPO). hTPO-antigen presentation to patient peripheral blood T cells was demonstrated after stimulation in vitro for 7 d with irradiated hTPO-transfected or untransfected autologous EBV-LCL. Anti-hTPO-reactive T cells were subsequently cloned in the presence of irradiated, autologous hTPO-transfected EBV-LCL and IL-2.10 T cell-cloned lines exhibited specific hTPO-induced proliferation (stimulation indices of 2.1-7.9) towards autologous hTPO-transfected EBV-LCL, and were subjected to human T cell receptor (hTCR) V gene analysis, using the PCR for the detection of V alpha and V beta hTcR gene families. The results indicated a preferential use of hTCR V alpha 1 and/or V alpha 3 in 9 of the 10 lines. In contrast, hTCR V beta gene family use was more variable. These data demonstrate a model for the endogenous presentation of human thyroid peroxidase in the absence of other thyroid specific antigens. The high frequency of antigen-specific T cells obtained from PBMC using this technique will facilitate further studies at both the functional and hTCR V gene level.

Recombinant human thyroid-stimulating hormone: initial bioactivity assessment using human fetal thyroid cells.

We have assessed the bioactivity of newly available recombinant human TSH (rec-hTSH) using human fetal thyroid cells, with the longer term aim of assessing its use for clinical applications. Rec-hTSH caused a consistent and dose-related increase in thyroid monolayer cell cAMP release and human thyroglobulin (hTg) secretion, confirming its bioactivity. Repetitive studies (n = 5) allowed us to derive an estimated biopotency for the rec-hTSH preparation examined of 5.6 IU/mg compared to 10 IU/mg for commercially available bovine TSH for human use. The rec-hTSH had a bioimmune ratio of 0.55, similar to that of purified pituitary hTSH standards, Furthermore, rec-hTSH induced thyroid epithelial cell growth, as evidenced by a decrease in thyroid cell doubling time from 54 +/- 2.1 to 31 +/- 1.7 h (P less than 0.005). Hence, rec-hTSH is a potent glycoprotein hormone preparation when measured in a homologous human thyroid cell culture system. Rec-hTSH could serve as a future definitive International Standard and has the potential for a useful diagnostic and therapeutic reagent.

Linkage analysis of candidate genes in autoimmune thyroid disease: 1. Selected immunoregulatory genes. International Consortium for the Genetics of Autoimmune Thyroid Disease.

Graves' and Hashimoto's diseases are autoimmune thyroid diseases in which the genetic contribution is complex. For this reason, identification of necessary susceptibility genes has been difficult. However, a number of immunoregulatory genes have been implicated by association studies, including: CTLA-4, a recently described protein involved in antigen presentation, located on chromosome 2q33; the T-cell receptor V alpha and V beta gene complexes, located on 14q11 and 7q35, respectively; and the Ig gene complex (IgH), located on 15q11. We used polymorphic microsatellite markers located within these genes, or gene complexes, to test for linkage (rather than association), to each of these candidates. Using markers within the loci allowed us to assume a fixed recombination fraction of 0.01 in the tested model. Three hundred eight subjects from 48 multiplex families were studied, with 142 affected subjects. Using this set of families, we have previously shown evidence of linkage with a major susceptibility locus for Graves' disease (GD-1) on 14q24.3-31, with a maximum lod (logarithm + odds) score of 2.1, at a penetrance of 80% and with a dominant mode of inheritance. In the present study, we obtained consistently negative lod scores for each of the candidate genes, assuming either dominant or recessive modes of inheritance. These data, therefore, showed evidence against linkage with all the candidate genes. Unlike association studies, linkage analyses detect major genetic influences on disease susceptibility exerted by the linked loci. The lack of linkage for the immunoregulatory genes that were studied indicated, therefore, that they were not major contributors to disease etiology.

Positive regulation of human thyrotropin receptor mRNA by thyrotropin.

We have assessed the influence of natural and recombinant thyrotropin (TSH) on mRNA specific for the human TSH receptor (TSHR) in normal and abnormal adult human thyroid monolayer cells. Using physiological concentrations of TSH (less than 100 mU/L), a marked increase in the level of TSHR mRNA was observed within 12 hours and reached a greater than 1000% increase after 24 hours exposure. At high TSH concentrations (greater than 1000 mU/L), this increase in TSHR-specific mRNA was markedly reduced. However, at no time were basal TSHR mRNA levels suppressed even with 100 U/L of TSH for 48 hours. These observations demonstrate ligand-induced up-regulation of the human TSHR mRNA by physiologically relevant concentrations of TSH.

Linkage analysis of candidate genes in autoimmune thyroid disease. III. Detailed analysis of chromosome 14 localizes Graves' disease-1 (GD-1) close to multinodular goiter-1 (MNG-1). International Consortium for the Genetics of Autoimmune Thyroid Disease.

The autoimmune thyroid diseases [Graves' and Hashimoto's diseases (GD and HT)] develop in genetically susceptible individuals, but the genes responsible for this susceptibility remain unknown. To identify such genes, we have been testing candidate genes and chromosomal regions using highly polymorphic microsatellite markers. We recently reported evidence for the first locus linked to GD (GD-1) on chromosome 14q31 in a small group of families. We have now extended these studies and analyzed 53 multiplex families with GD and/or HT (323 individuals). Chromosome 14 was screened using 16 microsatellite markers spanning the entire chromosome. Three additional markers located inside candidate genes on chromosome 14 were also studied. Microsatellite markers were amplified using fluorescent-labeled primers and separated on an ABI-310 genetic analyzer. The data were analyzed using LIPED software for two-point logarithm of odds (LOD) score analysis and GeneHunter software for multipoint linkage analysis. No linkage of any marker was found to HT or autoimmune thyroid diseases (GD+HT). The previously identified GD-1 locus on 14q31 continued to show evidence of linkage to GD in this much larger set of families. The maximum LOD score was 2.1 obtained for marker D14S81 (theta=0.01), assuming a recessive mode of inheritance and a penetrance of 0.3. Multipoint analysis yielded a maximum LOD score of 2.5 between markers D14S81 and D14S1054. There was no evidence for heterogeneity in our sample. These data again suggest the presence of a major Graves' disease susceptibilitygene (GD-1) on chromosome 14q31. This locus is close to the recently identified multinodular goiter-1 locus.

Linkage analysis of candidate genes in autoimmune thyroid disease. II. Selected gender-related genes and the X-chromosome. International Consortium for the Genetics of Autoimmune Thyroid Disease.

Hashimoto's thyroiditis (HT) and Graves' disease (GD) are autoimmune thyroid diseases (AITD) in which multiple genetic factors are suspected to play an important role. Until now, only a few minor risk factors for these diseases have been identified. Susceptibility seems to be stronger in women, pointing toward a possible role for genes related to sex steroid action or mechanisms related to genes on the X-chromosome. We have studied a total of 45 multiplex families, each containing at least 2 members affected with either GD (55 patients) or HT (72 patients), and used linkage analysis to target as candidate susceptibility loci genes involved in estrogen activity, such as the estrogen receptor alpha and beta and the aromatase genes. We then screened the entire X-chromosome using a set of polymorphic microsatellite markers spanning the whole chromosome. We found a region of the X-chromosome (Xq21.33-22) giving positive logarithm of odds (LOD) scores and then reanalyzed this area with dense markers in a multipoint analysis. Our results excluded linkage to the estrogen receptor alpha and aromatase genes when either the patients with GD only, those with HT only, or those with any AITD were considered as affected. Linkage to the estrogen receptor beta could not be totally ruled out, partly due to incomplete mapping information for the gene itself at this time. The X-chromosome data revealed consistently positive LOD scores (maximum of 1.88 for marker DXS8020 and GD patients) when either definition of affectedness was considered. Analysis of the family data using a multipoint analysis with eight closely linked markers generated LOD scores suggestive of linkage to GD in a chromosomal area (Xq21.33-22) extending for about 6 cM and encompassing four markers. The maximum LOD score (2.5) occurred at DXS8020. In conclusion, we ruled out a major role for estrogen receptor alpha and the aromatase genes in the genetic predisposition to AITD. Estrogen receptor beta remains a candidate locus. We found a locus on Xq21.33-22 linked to GD that may help to explain the female predisposition to GD. Confirmation of these data in HT may require study of an extended number of families because of possible heterogeneity.

T-cell receptor V gene use in autoimmune thyroid disease: direct assessment by thyroid aspiration.

We have examined the hTcR V gene family use of T-cells present in the aspiration thyroid biopsy specimens of patients with hyperthyroid Graves' disease (n = 8) and Hashimoto's autoimmune thyroiditis (n = 5). Nine of the 13 specimens had cytologically identified thyroid follicular cells, and 12 of the 13 contained human thyroglobulin-specific mRNA, confirming successful sampling. Of 18 hTcR V alpha and 19 hTCR V beta gene families tested for in the individual aspirates, a mean +/- SEM of 6.8 +/- 0.9 V alpha and 9.6 +/- 1.4 V beta gene families were present in the Graves' aspirates, while 12.2 +/- 1.7 and 16.8 +/- 0.4 V alpha and V beta gene families were present in the aspirates of patients with Hashimoto's thyroiditis. These samples, which offer a window onto the natural history of autoimmune thyroid disease, demonstrate significant hTcR V alpha and beta gene restriction in hyperthyroid Graves' disease, but much less restriction of both V alpha and V beta gene families in Hashimoto's disease. Such data extend our earlier information based only on examination of highly selected surgical specimens of patients with autoimmune thyroid disease to the much more typical patient. We conclude that hTcR V gene restriction of varying degrees is present in the majority of patients with autoimmune thyroid disease, but appears to be more easily detected in Graves', rather that Hashimoto's, disease.

Preservation of functioning human thyroid organoids in the scid mouse: II. Biased use of intrathyroidal T cell receptor V genes.

The severe combined immunodeficiency (scid) mouse, which lacks functional B cells and T cells, has proven a useful model for exploring the survival of transplanted human lymphocytes and thyrocytes. In order to further characterize T cell infiltrates in reconstituted sc human thyroid organoids, we examined for the presence of 18 human T cell receptor (hTcR) V alpha and 21 hTcR V beta gene families using polymerase chain reaction (PCR) analysis. Human TcR V gene activity was confirmed by Southern blot analysis of the PCR fragments from all but one of the thyroid organoids, confirming the continued survival of human T cells within the thyroid organoids. However, only 3.5 out of 18 V alpha and 5.9 out of 21 V beta gene families were detected in these human thyroid organoids indicating a marked bias in T cell survival. Sequencing of the V-D-J regions of the amplified TcR fragments showed that approximately 60% of the sequences were representative of clonally expanded T cells. Hence, these passenger T cells exhibited highly biased use of particular TcR V gene families similar to that observed previously in thyroid tissue and intrathyroidal T cell cultures. Furthermore, variations in the V-D-J regions of sequences from similar V gene families indicated that the V gene region was important in T cell selection rather than the CD3 region.

CTLA-4 and not CD28 is a susceptibility gene for thyroid autoantibody production.

One of the hallmarks of the human autoimmune thyroid diseases (AITDs) is the production of high titers of autoantibodies against thyroglobulin and thyroid peroxidase that often precedes the development of clinical disease. A high percentage of family members of patients with AITDs have significant titers of thyroid antibodies (TAbs), suggesting a genetic predisposition for their development, and segregation analyses have favored a dominant mode of inheritance. The aim of the present study was to identify the susceptibility genes for TAb production. We completed a genome-wide scan in 56 multiplex families (323 individuals) in which all family members with AITDs and/or detectable TAbs were considered affected. The highest 2-point logarithm of odds (LOD) score of 3.6 was obtained for marker D2S325 on chromosome 2q33 at 210.9 centimorgans. This locus showed no evidence for linkage to Graves' disease or Hashimoto's thyroiditis (2-point LOD scores, 0.42 for Graves' disease and -0.60 for Hashimoto's thyroiditis), demonstrating that the gene in this region conferred susceptibility to TAbs, but that clinical disease development required additional genetic and/or environmental factors. We then fine-mapped the region linked with TAbs using 11 densely spaced microsatellite markers. Multipoint linkage analysis using these markers showed a maximum LOD score of 4.2 obtained for marker D2S155 at 209.8 centimorgans (with heterogeneity, alpha = 0.41). As the linked region contained the CTLA-4 and CD28 genes, we then tested whether they were the susceptibility genes for TAbs on chromosome 2q33. The CD28 gene was sequenced in 15 individuals, and a new C/T single nucleotide polymorphism (SNP) was identified in intron 3. Analysis of this SNP revealed no association with TAbs in the probands of the linked families (families that were linked with D2S155) compared with controls. The CTLA-4 gene was analyzed using the known A/G(49) SNP, and the results showed a significantly increased frequency of the G allele in the probands of the linked families compared with the probands of the unlinked families or with controls (P = 0.02). We concluded that 1) a major gene for thyroid autoantibody production was located on chromosome 2q33; 2) the TAb gene on chromosome 2q33 was most likely the CTLA-4 gene and not the CD28 gene; and 3) CTLA-4 contributed to the genetic susceptibility to TAb production, but there was no evidence that it contributed specifically to Graves' or Hashimoto's diseases.

Mapping the major susceptibility loci for familial Graves' and Hashimoto's diseases: evidence for genetic heterogeneity and gene interactions.

The autoimmune thyroid diseases (AITDs), comprising Graves' disease (GD) and Hashimoto's thyroiditis (HT), appear to develop as a result of a complex interaction between predisposing genes and environmental triggers. The goals of the present study were to identify the susceptibility loci for GD and HT and to study the relationships between them. We performed a whole genome linkage study on a dataset of 56 multiplex, multigenerational AITD families (354 individuals), using 387 microsatellite markers. We identified 6 loci that showed evidence for linkage to AITD. Only one locus, on chromosome 6 [AITD-1; 80 centimorgans (cM)], was linked with both GD and HT [maximum LOD score (MLS), 2.9]. This locus was close to, but distinct from, the human leukocyte antigen region. One locus on chromosome 13 (HT-1; 96 cM) was linked to HT (MLS, 2.1), and another locus on chromosome 12 (HT-2; 97 cM) was linked to HT in a subgroup of the families (MLS, 3.8). Three loci showed evidence for linkage with GD: GD-1 on chromosome 14 (99 cM; MLS, 2.5), GD-2 on chromosome 20 (56 cM; MLS, 3.5), and GD-3 on chromosome X (114 cM; MLS, 2.5). Since GD-2 showed the strongest evidence for linkage to GD we fine-mapped this locus to a 1-cM interval between markers at 55 and 56 cM on chromosome 20. These results demonstrated that 1) Graves' and Hashimoto's diseases are genetically heterogeneous, with only one locus in common to both diseases on chromosome 6; 2) only one HT locus was identified in all families, probably due to heterogeneity of the HT phenotype; and 3) three loci were shown to induce genetic susceptibility to GD by interacting with each other. One of them (GD-2) was fine-mapped to a 1-cM interval.

Evidence for association of polycystic ovary syndrome in caucasian women with a marker at the insulin receptor gene locus.

The polycystic ovary syndrome (PCOS) is one of the commonest female endocrinopathies affecting 5-10% of women of reproductive age. The disorder, characterized by chronic anovulation and signs of hyperandrogenism, results from a complex interaction between genetic predisposing factors and environmental triggers. We have studied 85 Caucasian PCOS patients and 87 age-matched Caucasian control women for associations with four candidate genes: follistatin, CYP19 (aromatase), CYP17a, and the insulin receptor (INSR). These genes were analyzed using microsatellite markers located near or inside the genes. We found that only the insulin receptor gene marker D19S884 was significantly associated with PCOS (p=0.006 and even after a conservative correction p=0.042). The INSR gene region was then fine mapped with an additional panel of 9 markers but only marker D19S884, located 1 cM telomeric to the INSR gene, was again associated with PCOS. In conclusion, our results suggested that a susceptibility gene for PCOS was located on chromosome 19p13.3 in the insulin receptor gene region. It remains to be determined if this susceptibility gene is the insulin receptor gene itself or a closely located gene. Since insulin stimulates androgen secretion by the ovarian stroma it is likely that INSR function in the ovary is involved in the genetic susceptibility ot PCOS.

Endogenous antigen presentation by autoantigen-transfected Epstein-Barr virus-lymphoblastoid cells: T cell receptor N-region hydrophobicity relates to thyroid antigen recognition.

Seven human T cell lines from a patient with Graves' disease were raised against endogenously generated human thyroid peroxidase (hTPO) with stimulation indices ranging from 2.1 to 7.6. Clonal expansion within these T cell lines was demonstrated by sequencing multiple bacterial colonies containing RT-PCR-generated fragments derived from the expressed hTcRs. Some lines had more than one human T cell receptor (hTcR) alpha and beta chain mRNAs as judged by RT-PCR. Stopcodons present in several hTcR sequences indicated that only one V alpha and one V beta gene were translated. Both the V alpha/beta gene families and the J alpha/beta gene segments differed amongst the lines and no characteristic recognition sequences were discernable in the CDR3 regions. Using Kyte-Doolittle analysis we found hydrophobic peaks in most N alpha-regions (but not N beta regions) suggesting that hydrophobic interactions may be important in the recognition of hTPO. However, increasing affinity values, as measured by SI, were strongly correlated with decreasing hydrophobicity in the N alpha region (1st order regression, r = -0.93138, p < 0.01). Thus, lower affinity, self-reactive, T cells may be more hydrophobic ('sticky') in their N alpha regions while higher affinity cells may be characterized by TcRs with lower hydrophobicity. These findings demonstrate a substantial role for hydrophobic interactions in hTPO-reactive T cell receptors and further support a role for the TcR alpha chain in the recognition of thyroid autoantigen.

Graves' disease thyroid tissue transplants in scid mice: persistent selectivity in hTcR Va gene family use.

We have analyzed the human T-cell receptor (hTcR) V alpha gene repertoire in thyroid tissue transplants of a patient with hyperthyroid Graves' disease. Blocks of thyroid tissue were transplanted subcutaneously into 10 mice with severe immunodeficiency (scid) and 4 weeks later 5 of the mice were injected intraperitoneally with autologous peripheral blood mononuclear cells (PBMC) (10(7) cells per mouse). After a further 3 weeks, mice were sacrificed and total cellular RNA and cDNA prepared from each of the explants. We used specific olingonucleotides in polymerase chain reactions (PCR) to amplify 18 different human hTcR V alpha gene families and the identity of the PCR fragments was confirmed by Southern blot analysis. Different samples of the donor thyroid tissue consistently expressed 9-10 of the 18 hTcR V alpha gene families screened (V alpha 1-7, 11, 12 & 15). A more marked bias in hTcR V gene family use was seen in each of the explants with a mean of only 2.8 V alpha gene families detected. After 7 weeks of transplantation, the thyroid explants largely reflected some of the same genes seen in the hTcR V gene repertoire of the donor tissue with particularly pronounced expression of V alpha 2 and V alpha 3 gene families. The transplantation of PBMC into the scid mice showed evidence for their accumulation within the transplanted thyroid tissues as judged by the appearance of additional hTcR V gene families expressed in these samples although the specificity of such accumulation remains unclear.(ABSTRACT TRUNCATED AT 250 WORDS)

Lack of disease associated HLA-DQ restriction fragment length polymorphisms in families with autoimmune thyroid disease.

Autoimmune thyroid disease (AITD) is often familial and serological HLA disease associations have been described in many different populations. However, such HLA disease associations are weak and the precise molecular contribution of HLA antigens to thyroid disease susceptibility remains unknown. Much of the data available are cross-sectional and few studies have explored familial inheritance of AITD at the molecular level. We have, therefore, examined the inheritance of AITD in multiplex and multi-generational families using restriction fragment length polymorphism (RFLP) analysis of DNA digested with the restriction enzyme BamH1 and probed with a full length human HLA-DQ beta cDNA probe. Thirty seven subjects in 7 informative families were available for study. Eleven subjects had Graves' disease and 4 were diagnosed as having Hashimoto's thyroiditis. Segregation of polymorphic fragments enabled genotyping of each individual to produce fully informative families. LOD scores were computed, using the LIPED program, for dominant and recessive models of inheritance, for recombination fractions of 0.01 to 0.5 for each sex, and for penetrances of 0.1 to 1.0. The results showed that maximum LOD scores were negative for all of the inheritance models tested. If the primary locus for AITD were in the HLA region, LOD scores would be highly positive. These data, therefore, provide strong evidence against a disease locus for AITD in linkage disequilibrium with the HLA-DQ beta locus.

Development of a scale to predict decline in patients with mild Alzheimer's disease.

OBJECTIVES: To develop a scale that can assist in predicting likelihood of decline from mild dementia over 1 year in patients with Alzheimer's disease (AD). DESIGN: Retrospective cohort study. SETTING: University Memory and Aging Center. PARTICIPANTS: Patients with probable or possible AD and Clinical Dementia Rating (CDR) of 1 at baseline, divided into development and validation cohorts (n = 118 each). MEASUREMENTS: The CDR and neurological and neuropsychological assessments were given at baseline and 1 year later. RESULTS: In the development cohort, high education, low Mini-Mental State Examination score, poor insight, psychotic symptoms, and greater activity of daily living impairment predicted decline in CDR from 1 to 2 or 3. Receiver operating characteristics (ROC) curve analysis identified cutoff scores that maximized sensitivity and specificity for each significant predictor of decline. Based on the cutoff, raw scores were recoded to reflect risk for decline, weighted, and summed to create a final scale score. ROC curve analysis established a cutoff to indicate risk for decline on the final scale score. Sensitivity, specificity, and area under the ROC were 0.76, 0.74, and 0.83 in the development cohort and 0.77, 0.69, and 0.80 in the validation cohort, respectively. Positive and negative predictive values were 0.71 and 0.78 in the development cohort and 0.68 and 0.78 in the validation cohort, respectively. CONCLUSIONS: Decline from mild to moderate or severe impairment represents significant clinical change, with implications for patient and caregiver quality of life and treatment options. The clinical scale developed uses data to enhance prediction about change from mild to moderate or severe stages of AD.

Detection of the gene rearrangement in chronic myelogenous leukemia with biotinylated gene probes.

The breakpoint cluster region gene rearrangement associated with chronic myelogenous leukemia is becoming important in the diagnosis and management of the disease. At this time, the ability to demonstrate the gene rearrangement is limited to a few research laboratories. The problem results partially from unfamiliarity of medical laboratory personnel with DNA technology, but more because of the restricted use of radiolabeled phosphorus in hospital laboratories. With the introduction of biotinylated deoxynucleotides, nucleic acid hybridization procedures can now be performed without the use of radioisotopically labeled gene probes. This article describes the use of biotin-labeled gene probes to detect the gene rearrangement of the breakpoint cluster region of chromosome 22 in chronic myelogenous leukemia. The techniques are reproducible, sensitive, and safe. With the procedures described in this article, the assay can become more available to medical laboratories interested in offering this diagnostic and decision-making tool.

Limited genetic susceptibility to severe Graves' ophthalmopathy: no role for CTLA-4 but evidence for an environmental etiology.

Graves' disease (GD) is an autoimmune thyroid disease (AITD) characterized by hyperthyroidism and by the occurrence of a distinctive ophthalmopathy (orbitopathy), which presents with varying degrees of severity. Graves' disease clusters in families but the importance of heredity in the pathogenesis of the associated ophthalmopathy is unclear. We have studied the family history of 114 consecutive, ethnically mixed patients with severe Graves' ophthalmopathy (GO). Patients were selected by unambiguous single ascertainment. Seventy-seven percent of patients were female and 59% smoked. The mean age at onset of their GD was 43 years (range 17-78 years). Forty-one patients (36%) had a family history of AITD, defined as a first- and/or a second-degree relative affected with either Graves' disease (GD) or Hashimoto's thyroiditis (HT). The segregation ratio for AITD in nuclear families in our ascertained Graves' ophthalmopathy families was 0.07 (0.12 in Caucasians only). Hence, the higher prevalence of AITD among relatives of Graves' ophthalmopathy patients agreed with the known genetic predisposition to AITD and this predisposition was stronger in women than in men. However, only 3 of the 114 patients had a family history of severe Graves' ophthalmopathy (all second-degree relatives) and the segregation ratio for GO was 0. These data did not support a major role for familial factors in the development of severe Graves' ophthalmopathy distinct from Graves' disease itself. In addition, we tested 4 candidate genes, human leukocyte antigen (HLA), tumor necrosis factor-beta (TNF-beta), CTLA-4 and the thyrotropin receptor (TSHR), for association with Graves' ophthalmopathy. These were negative except for the HLA and CTLA-4 genes, which were found to be weakly associated with GO giving similar relative risk (RR) as in GD patients without ophthalmopathy. These data suggested that environmental factors, rather than major genes, were likely to predispose certain individuals with AITD to severe Graves' ophthalmopathy. Smoking remains one example of such potential external insults.