Functional Domains of Autoimmune Regulator (AIRE) Modulate INS-VNTR Transcription in Human Thymic Epithelial Cells.

INS-VNTR (insulin-variable number of tandem repeats) and AIRE (autoimmune regulator) have been associated with the modulation of insulin gene expression in thymus, which is essential to induce either insulin tolerance or the development of insulin autoimmunity and type 1 diabetes. We sought to analyze whether each functional domain of AIRE is critical for the activation of INS-VNTR in human thymic epithelial cells. Twelve missense or nonsense mutations in AIRE and two chimeric AIRE constructs were generated. A luciferase reporter assay and a pulldown assay using biotinylated INS-class I VNTR probe were performed to examine the transactivation and binding activities of WT, mutant, and chimeric AIREs on the INS-VNTR promoter. Confocal microscopy analysis was performed for WT or mutant AIRE cellular localization. We found that all of the AIRE mutations resulted in loss of transcriptional activation of INS-VNTR except mutant P252L. Using WT/mutant AIRE heterozygous forms to modulate the INS-VNTR target revealed five mutations (R257X, G228W, C311fsX376, L397fsX478, and R433fsX502) that functioned in a dominant negative fashion. The LXXLL-3 motif is identified for the first time to be essential for DNA binding to INS-VNTR, whereas the intact PHD1, PHD2, LXXLL-3, and LXXLL-4 motifs were important for successful transcriptional activation. AIRE nuclear localization in the human thymic epithelial cell line was disrupted by mutations in the homogenously staining region domain and the R257X mutation in the PHD1 domain. This study supports the notion that AIRE mutation could specifically affect human insulin gene expression in thymic epithelial cells through INS-VNTR and subsequently induce either insulin tolerance or autoimmunity.

Bullous skin lesions in an adult male: a diagnostic dilemma.

BACKGROUND: Henoch-Schönlein Purpura (HSP) is an IgA small-vessel vasculitis that is primarily a disease of childhood. Its presentation in adulthood is rare and has a more severe disease course. We present a case with an atypical presentation of this disease that was a diagnostic challenge for multiple providers. CASE REPORT: A 42-year-old man noticed bullous lesions over his ankles that spread to his entire legs over a few weeks. They later became necrotic and ulcerated areas. His primary care physician and 2 dermatologists could not reach a definitive diagnosis. He then presented to our hospital with new abdominal pain, rectal bleeding, and a new elevation in liver enzymes. A biopsy of his skin lesions led to the diagnosis of HSP. CONCLUSIONS: We discuss this highly unusual initial presentation with bullous skin lesions and liver enzyme abnormalities and explore the medical literature to understand its pathogenesis. Clinicians need to be aware of this rare presentation to avoid a delay in diagnosis and management.

Fibroblast activation protein-α: a key modulator of the microenvironment in multiple pathologies.

Fibroblast activation protein-α (FAP) is a serine protease that can provide target specificity to therapeutic agents because in adults its expression is restricted to pathologic sites, including cancer, fibrosis, arthritis, wounding, or inflammation. It is not expressed in most normal tissues. The majority of FAP is expressed by activated fibroblasts responding to the pathologic situations. FAP is typically found as a type II transmembrane protein physically attached to cells and with the bulk of the protein, including the catalytic domain, exposed to the extracellular space and accessible to small molecules. In this chapter, we review the structure, substrate specificities, signaling functions, and current design of FAP inhibitors. Evidence indicating the presence of FAP in multiple cancers, arthritis, fibrosis, keloids, and other pathologies is described and indicates possible roles for FAP in facilitating cell invasion and growth. Separate sections are devoted to the role of FAP in coordinating the stromal response to cancer, including a role in angiogenesis and a potential role in modulation of the antitumor immune response. Finally studies attempting to demonstrate the clinical potential of FAP are discussed, as well as some novel applications employing FAP in therapy or diagnosis. Throughout this review, effort is made to highlight areas where information is lacking and to highlight important questions that require further investigation.

Fibroblast activation protein-α promotes tumor growth and invasion of breast cancer cells through non-enzymatic functions.

Fibroblast activation protein-α (FAP) is a cell surface, serine protease of the post-prolyl peptidase family that is expressed in human breast cancer but not in normal tissues. Previously, we showed that FAP expression increased tumor growth rates in a mouse model of human breast cancer. Here the role of the proteolytic activities of FAP in promoting tumor growth, matrix degradation and invasion was investigated. Mammary fat pads of female SCID mice were inoculated with breast cancer cells that express FAP and the mice treated with normal saline or Val-boroPro (talabostat); Glu-boroPro (PT-630); or 1-[[(3-hydroxy-1-adamantyl)amino]acetyl]-2-cyano-(S)-pyrrolidine (LAF-237) that inhibit prolyl peptidases. Other mice were injected with breast cancer cells expressing a catalytically inactive mutant of FAP and did not receive inhibitor treatment. PT-630 and LAF-237 did not slow growth of tumors produced by any of the three cell lines expressing FAP. Talabostat slightly decreased the growth rates of the FAP-expressing tumors but because PT-630 and LAF-237 did not, the growth retardation was likely not related to the inhibition of FAP or the related post-prolyl peptidase dipeptidyl peptidase IV. Breast cancer cells expressing a catalytically inactive mutant of FAP (FAP(S624A)) also produced tumors that grew rapidly. In vitro studies revealed that cells expressing wild type FAP or FAP(S624A) degrade extracellular matrix (ECM) more extensively, accumulate higher levels of matrix metalloproteinase-9 (MMP-9) in conditioned medium, are more invasive in type I collagen gels, and have altered signaling compared to control transfectants that do not express FAP and form slow growing tumors. We conclude that the proteolytic activity of FAP participates in matrix degradation, but other functions of the protein stimulate increased tumor growth.

Repeat testing for congenital hypothyroidism in preterm infants is unnecessary with an appropriate thyroid stimulating hormone threshold.

BACKGROUND: Revised UK neonatal screening guidelines recommend that a second blood sample for assay of thyroid stimulating hormone (TSH) be taken when preterm infants reach a postmenstrual age of 36 weeks. OBJECTIVE: To examine the results of a regional screening programme to see whether a rise in TSH concentration was observed in some preterm infants between the first sample taken around 5 days after delivery and the second sample taken at around 36 weeks. METHODS: Whole-blood TSH concentrations in preterm infants born over a 2-year period (April 2005 to March 2007) were assessed, and the number of infants in whom there was a fall or rise to values below or above the local screening threshold (6 mU/l) was determined. RESULTS: Baseline TSH samples were obtained from 2238 preterm infants (median gestational age 32 weeks, range 21-35) with second samples obtained from 2039 (median gestational 32 weeks, range 23-35). In 19 infants, TSH concentrations fell from above to below the screening threshold, and in five infants values rose from below the screening threshold to 6-10 mU/l. However, TSH concentrations fell to <6 mU/l on a further blood spot in four of these infants, and the remaining infant had a serum TSH of 6.8 mU/l. Three infants had raised TSH concentrations on both occasions with unequivocal hypothyroidism (serum TSH >80 mU/l). The initial TSH concentration in one of these infants was 6-10 mU/l. CONCLUSIONS: No infant with a normal TSH concentration on first sampling had a TSH concentration that rose above 10 mU/l on second sampling, and no infants with a normal TSH concentration on first screening are receiving long-term thyroxine treatment. This study suggests that a second sample may not be necessary with a screening threshold of 6 mU/l.

Family study of fibromyalgia.

OBJECTIVE: To assess for familial aggregation of fibromyalgia (FM) and measures of tenderness and pain, and for familial coaggregation of FM and major mood disorder (major depressive disorder or bipolar disorder). METHODS: Probands meeting the American College of Rheumatology criteria for FM and control probands with rheumatoid arthritis (RA) and no lifetime diagnosis of FM were recruited from consecutive referrals to 2 community-based rheumatology practices. Probands were ages 40-55 years and had at least 1 first-degree relative age 18 years or older who was available for interview and examination. All probands and interviewed relatives underwent a dolorimeter tender point examination and a structured clinical interview. Interviewed relatives were asked about first-degree relatives who were not available for interview, using a structured family interview. Logistic and linear regression models, adjusting for the correlation of observation within families, were applied to study the aggregation and coaggregation effects. RESULTS: Information was collected for 533 relatives of 78 probands with FM and 272 relatives of 40 probands with RA. FM aggregated strongly in families: the odds ratio (OR) measuring the odds of FM in a relative of a proband with FM versus the odds of FM in a relative of a proband with RA was 8.5 (95% confidence interval [95% CI] 2.8-26, P = 0.0002). The number of tender points was significantly higher, and the total myalgic score was significantly lower in the relatives of probands with FM compared with the relatives of probands with RA. FM coaggregated significantly with major mood disorder: the OR measuring the odds of major mood disorder in a relative of a proband with FM versus the odds of major mood disorder in a relative of a proband with RA was 1.8 (95% CI 1.1-2.9, P = 0.013). CONCLUSION: FM and reduced pressure pain thresholds aggregate in families, and FM coaggregates with major mood disorder in families. These findings have important clinical and theoretical implications, including the possibility that genetic factors are involved in the etiology of FM and in pain sensitivity. In addition, mood disorders and FM may share some of these inherited factors.

Preoperative ICU tours: are they helpful?

BACKGROUND: Although preoperative education decreases the anxiety of patients and family members, the usefulness of a preoperative tour of the ICU has not been studied. In this study, the effect of an ICU tour on the anxiety levels of patients (n = 92) and family members (n = 91) before and after cardiac surgery was examined. METHODS: Using a pretest-posttest quasi-experimental design, patients and family members were assigned to a control group, which received preoperative teaching only (patients, n = 48; family members, n = 48), or to an experimental group, which received preoperative teaching with an ICU tour (patients, n = 44; family members, n = 43). Patients and family members completed two measures of anxiety, the State Trait Anxiety Inventory and a visual analog scale, before and after the intervention. After their first postoperative visit, family members completed the measures again. Repeated measures analysis of variance was used to compare anxiety levels after the intervention. In addition, patients completed the Patient Perception of the ICU Tour Questionnaire after transfer from the ICU. RESULTS: Although patients and family members had a decrease in anxiety after cardia surgery teaching, the decrease was not due to an ICU tour. However, the majority of patients who toured the ICU perceived the tour as beneficial and recommended a tour for future patients. CONCLUSIONS: ICU tours are included in many cardiac surgery educational programs. The majority of patients in this study perceived a benefit or a future benefit from an ICU tour, even though the tour did not significantly reduce the anxiety of the patients or family members.