The tropical fowl mite, Ornithonyssus bursa (Acari: Macronyssidae): environmental and host factors associated with its occurrence in Argentine passerine communities.

The tropical fowl mite, Ornithonyssus bursa, is a common avian parasite found on diverse bird species worldwide. In the Neotropical region, O. bursa is present in wild birds, but it may also infect poultry and bite humans. Little is known about the ecology and epidemiology of this parasite. We conducted a thorough longitudinal study in passerine assemblages from central Argentina, gathering data from six reproductive seasons, with the aim of identifying factors that have a role in driving the occurrence and distribution of O. bursa in its natural hosts. We focused on the brood and microhabitat levels, accounting for potential confounders of higher levels. The results hereby presented contribute to our knowledge on the eco-epidemiology of O. bursa in natural hosts of the Neotropical region. Among the many variables assessed, nest material and host species appeared to be the most important correlates of O. bursa prevalence. Nonetheless, supplementary analyses showed that host species is a stronger predictor than nest material. Moreover, mite burden (parasite intensity) was found to depend on host species, but not on nest material. The association with species depended on nestling age, suggesting that resistance builds up as the nestling develop, but at a different pace depending on the bird species. Brood size was inversely correlated with intensity of parasitism, suggesting a dilution of the parasite burden on each nestling.

3D MRI segmentation and 3D circumferential resection margin evaluation for a standard rectal cancer assessment.

AIM: Recent studies focused on rectal cancer suggested that a 3D imaging segmentation obtained from MRI data could contribute in the definition of the circumferential resection margin (CRM) and in the assessment of the tumor regression following neo-adjuvant treatments. Here, we propose a method for defining and visualizing the circumferential margins using 3D MRI segmentation; this methodology was tested in a clinical study comparing 3D CRM assessment vs standard MRI imaging. PATIENTS AND METHODS: MRI scans performed before neo-adjuvant treatments were selected and reviewed. 3D mesorectal/tumor segmentations were obtained using Digital Imaging and COmmunications in Medicine (DICOM) data; CRMs were calculated using 3D volumes plus a color scale for the closest distances. RESULTS: 3D reconstructions were possible in all selected cases and 3D images implemented by the color scale were positive for immediate CRM visualization. Statistical analyses comparing standard radiology disclosed that the degree of consistency, the reliability of ratings, the correlation and precision were optimal considering the overall cases, but lower in the CRM>0 mm sub-group. CONCLUSIONS: This new method is not inferior comparing standard radiology; moreover, the imaging segmentation we obtained was highly promising and could be helpful in defining a standard CRM measurement, thus it could improve clinical practice.

The insulin gene is transcribed in the human thymus and transcription levels correlated with allelic variation at the INS VNTR-IDDM2 susceptibility locus for type 1 diabetes.

Type 1, or insulin-dependent diabetes mellitus (IDDM) is an autoimmune disease associated with loss of tolerance to several pancreatic islet cell molecules, including insulin, glutamic acid decarboxylase (GAD), ICA69 and the tyrosine phosphatase IA-2 (refs 1-3). Among several predisposing loci, IDDM2 maps to the insulin gene (INS) VNTR (variable number of tandem repeats) minisatellite on chromosome 11p15 (refs 4-9). Allelic variation at this VNTR locus correlates with steady-state levels of INS mRNA in pancreas and transfected rodent cell lines, but it is difficult to reconcile the association of lower INS mRNA levels in the pancreas with class III VNTRs that are dominantly protective from IDDM. We show that during fetal development and childhood, mRNAs for insulin and other islet cell autoantigens (GAD, ICA69, IA-2) are expressed at low levels in the human thymus. Critically, we also detect proinsulin and insulin protein. VNTR alleles correlate with differential INS mRNA expression in the thymus where, in contrast to the pancreas, protective class III VNTRs are associated with higher steady-state levels of INS mRNA expression. This finding provides a plausible explanation for the dominant protective effect of class III VNTRs, and suggests that diabetes susceptibility and resistance associated with IDDM2 may derive from the VNTR influence on INS transcription in the thymus. Higher levels of (pro)insulin in the thymus may promote negative selection (deletion) of insulin-specific T-lymphocytes which play a critical role in the pathogenesis of type-1 diabetes.

Misdiagnosis in fibromyalgia: a multicentre study.

BACKGROUND: Fibromyalgia (FM) is the second most common cause of visits to rheumatologists after osteoarthritis, and may be difficult to diagnose in many patients. It is associated with various rheumatic disorders such as rheumatoid arthritis, spondyloarthropathies (SpA) and connective tissue disease (CTD), and a late diagnosis or misdiagnosis is a common and underestimated problem. OBJECTIVES: The aim of this study was to investigate the 'underdiagnosis' of FM, and which rheumatic diseases tend to be confused with it. METHODS: The following data were collected at baseline: symptoms, disease duration, physical examination findings, previous and current investigations and management, laboratory tests, tender point count, tender and swollen joint counts, and spinal pain. The clinimetric evaluation included the Fibromyalgia Impact Questionnaire (FIQ) and Fibromyalgia Assessment Status (FAS). RESULTS: The study population consisted of 427 outpatients (418 females and 9 males; mean age 49.3 years; mean disease duration 8.5 years). Fifty-seven patients (13.3%) had been previously misdiagnosed as having other musculoskeletal disorders (MSDs); 370 patients had been previous correctly diagnosed as having FM, or were diagnosed as having it during the course of the study. The FM and MSD groups were comparable in terms of demographic data and referral patterns. Disease duration was longer and the erythrocyte sedimentation rate was higher in the MSD patients, who also had less severe FIQ and lower pain visual analogue scale scores. Moreover, the FIQ and FAS scores correlated in the MS group. CONCLUSIONS: The findings of this study suggest that, although FM is a wellknown clinical entity, differential diagnosis with SpA, CTD and inflammatory arthritis can still be a challenge for rheumatologists and general practitioners.

Islet cell autoantigen 69 kD (ICA69). Molecular cloning and characterization of a novel diabetes-associated autoantigen.

We have identified a novel 69-kD peptide autoantigen (ICA69) associated with insulin-dependent diabetes mellitus (IDDM) by screening a human islet lambda gt11 cDNA expression library with cytoplasmic islet cell antibody positive sera from relatives of IDDM patients who progressed to the overt disease. The deduced open reading frame of the ICA69 cDNA predicts a 483-amino acid protein. ICA69 shows no nucleotide or amino acid sequence relation to any known sequence in GenBank, except for two short regions of similarity with BSA. The ICA69 cDNA probe hybridizes with a 2-kb mRNA in poly(A+) RNA from human pancreas, brain, heart, thyroid, and kidney, but not with skeletal muscle, placenta, spleen, or ovary. Expression of ICA69 was also detected in beta cells and cell lines, as well as in tumoral tissue of islet cell origin. The native ICA69 molecule migrates to 69 kD in SDS-PAGE as detected with specific antibodies. Serum samples from relatives of IDDM patients specifically reacted with affinity-purified recombinant ICA69 on Western blotting. The structural gene for ICA69 was designated ICA1. A homologue in the mouse, designated Ica-1 was mapped to the proximal end of chromosome 6 (within 6 cM of the Met protooncogene). ICA69 adds a novel autoantigen to the family of identified islet target molecules, and by the manner of its identification and characterization large amounts of antigen are available for development of quantitative, convenient predictive assays for autoantibodies and analysis of the role of this molecule in diabetes autoimmunity, as well as its physiologic function.

Platelet-leukocyte interactions in hemodialysis patients: culprit or bystander?

The formation of circulating platelet-leukocyte complexes has been observed in a variety of conditions and may be pathophysiologically significant. Platelet-leukocyte interactions in fact facilitate metabolic cooperation and mutual activation, which may be of relevance in many biological processes including inflammation, atherogenesis and hemostasis. During hemodialysis procedure, the series of reactions that can occur upon blood contact with the foreign membrane surface may involve a variety of changes affecting almost every cellular and plasmatic component of the blood. This article reviews the evidence for abnormal interactions between circulating platelets and leukocytes in uremic patients undergoing maintenance hemodialysis and the pathophysiologic implications which may stem from such interactions.

Gene expression of islet cell antigen p69 in human, mouse, and rat.

Based on the detection of specific antibodies and T-cell sensitization in patients with IDDM, islet cell antigen p69 (ICAp69) has been suggested to be a target antigen of diabetic autoimmunity. The biological function, tissue expression, and developmental kinetics of ICAp69 are largely unknown. We analyzed ICAp69 expression at the gene transcription and protein level in human and rodent tissues. By using template-calibrated quantitative reverse transcriptase polymerase chain reaction (RT-PCR), high levels of ICAp69 mRNA were found in human pancreatic islets and brain. In mouse and rat, ICAp69 gene expression peaked in islet cell lines followed by testis, islets, and brain. ICAp69 mRNA was found at low levels in other organs by RT-PCR but not by Northern blot analysis. In mice, ICAp69 transcription becomes detectable in fetal life, and fetal and adult gene expression patterns are similar. Western blot analysis of human and mouse tissues showed high expression of ICAp69 in brain, testis, pancreatic tissue, and islet cell lines. In these organs, ICAp69 immunoreactivity is predominately localized at the blood brain barrier (capillary endothelium), at the blood testis barrier (Sertoli cells and spermatids), and in pancreatic islets (beta-cells). The subcellular localization of ICAp69 to endoplasmic reticulum, Golgi complex, and vesicles by immune electron microscopy suggests a role of this neuroendocrine molecule in cellular protein traffic and processing.off

Evidence of islet cell autoimmunity in elderly patients with type 2 diabetes.

In light of an occurring growth of elderly people affected by type 2 diabetes and recent observations indicating that type 2 diabetes may be a disease of the innate immune system, we evaluated whether signs of islet cell autoimmunity are associated with an abnormal glucose control, the presence of insulin requirement, or an activation of the acute-phase response in older individuals with type 2 diabetes. GAD65 and IA-2 autoantibodies along with the acute-phase response markers fibrinogen and C-reactive protein were tested in 196 serum samples from patients with type 2 diabetes and in 94 nondiabetic control subjects over the age of 65 years from the Pittsburgh cohort of the Cardiovascular Health Study. Of the diabetic patients, 12% (24 of 196) had autoantibodies against GAD65 and/or IA-2, a prevalence significantly higher than that found in nondiabetic individuals (1 of 94, 1.1%; P = 0.001). Type 2 diabetic patients who were positive for GAD65 and/or IA-2 autoantibodies (Ab+), as compared with those negative for these autoantibodies (Ab-), had an abnormal oral glucose tolerance test (OGTT) (P = 0.03) before and a higher frequency of oral hypoglycemic treatment (P = 0.003) at the time of autoantibody testing. No differences were seen in the percentage of insulin requirement in the two groups. Moreover, a statistically significant increase in fibrinogen (P = 0.005) and C-reactive protein levels (P = 0.025) was found in type 2 diabetic patients with high levels of GAD65 and/or IA-2 autoantibodies as compared with Ab-patients and control subjects. In conclusion, in type 2 diabetic subjects > or =65 years old, the presence of islet cell autoimmunity is associated with an impairment of the acute-phase insulin secretion, as revealed by an OGTT. A pronounced activation of the acute-phase response, found to be associated with islet cell autoimmunity, may in part explain this defect in insulin secretion. These findings not only have direct implications for adequate classification and treatment of diabetes in the elderly, but also for understanding the autoimmune/inflammatory mechanisms involved in the pathogenesis of hyperglycemia.

ICA512 autoantibody radioassay.

As part of a general program of screening islet expression libraries we have identified a clone from a lambda gt11 human islet expression library that reacts with human diabetic sera and, upon sequencing, was determined to be the neuroendocrine islet autoantigen ICA512 (islet cell antigen 512). In the current communication, we describe the development of a radioassay for autoantibodies to ICA512 (ICA512AA) using in vitro transcribed and translated protein for production of labeled antigen. Our initial results indicate that this radioassay is significantly more sensitive than the enzyme-linked immunosorbant assay, which uses a COOH-terminal fragment of ICA512. The ICA512AA radioassay uses a 96-well format with membrane separation of antibody bound from free antigen and should be readily adaptable to automated large-scale screening. Only 7 microliters of serum is required for triplicate determinations. In order to determine the specificity and sensitivity of this assay and estimate its positive predictive value, we have studied 42 new-onset diabetic patients, 33 first-degree relatives of diabetic patients followed to diabetes, 694 islet cell antibody-negative (ICA-) relatives, and 205 normal control subjects. Thirty-eight percent of new-onset patients and 48% of relatives followed to diabetes express autoantibodies to ICA512 exceeding the 99th percentile of the normal control subjects. In contrast, only 1.4% of ICA- first-degree relatives were positive for ICA512 autoantibodies.(ABSTRACT TRUNCATED AT 250 WORDS)

Late progression to diabetes and evidence for chronic beta-cell autoimmunity in identical twins of patients with type I diabetes.

Previous studies suggest that after 6 years of discordance, identical twin pairs rarely become concordant for type I diabetes. With up to 39 years of follow-up from the onset of diabetes in the index twin, we determined how many discordant twins have evidence of beta-cell autoimmunity and how many develop overt diabetes. We longitudinally followed 23 pairs of identical twins (or triplets) that were selected from a total group of 30 pairs because they were discordant for type I diabetes when first ascertained. Seven developed diabetes after 3, 3, 7, 8, 9, 31 and 36 years of discordance. By survival analysis, the concordance after 10 years from the onset of diabetes in the index twin was estimated as 23% (95% confidence interval, 5-40%), increasing to 38% (95% confidence interval, 8-69%) after 31 years. Among 16 twins remaining nondiabetic at last follow-up (8-39 years of discordance), 12 were assessed with serial intravenous glucose tolerance tests and a total of 407 measurements by radioassay of antibodies against three defined autoantigens (glutamic acid decarboxylase, insulin, and the recently cloned molecule ICA512). Two-thirds (8 of 12) had evidence of beta-cell autoimmunity (persistently positive autoantibody levels) and/or first-phase insulin release less than the 1st percentile of control subjects. In summary, identical twins may develop diabetes after a prolonged period of discordance and approximately two-thirds of long-term discordant twins have evidence of persistent beta-cell autoimmunity and/or beta-cell damage. The concordance for beta-cell autoimmunity, therefore, is much higher than for overt diabetes.(ABSTRACT TRUNCATED AT 250 WORDS)

Prediction of type I diabetes in first-degree relatives using a combination of insulin, GAD, and ICA512bdc/IA-2 autoantibodies.

Islet cell antibodies (ICAs) are predictive of type I diabetes in first-degree relatives, but this immunohistochemical assay has proven difficult to standardize. As an alternative, we assessed the use of radioassays for antibodies against three molecularly characterized islet autoantigens, including ICA512bdc (amino acid residues 256-979 of the IA-2 molecule, incorporating the intracellular domain). We measured insulin autoantibodies (IAAs), GAD autoantibodies (GAAs), and ICA512bdc autoantibodies (ICA512bdcAAs) by radioassay, in addition to ICAs, in 882 first-degree relatives of patients with type I diabetes, 50 of whom later developed diabetes with a median follow-up of 2.0 years (maximum 11.3 years). The cutoff for each radioassay was determined by testing >200 control subjects. When autoantibody frequencies among the relatives were analyzed according to relationship to the proband, the offspring of diabetic fathers had a higher frequency of ICA5I2bdcAAs (P = 0.008), IAAs (P = 0.0001) and GAAs (P = 0.0001) than the offspring of diabetic mothers. ICA512bdcAAs and IAAs both showed a significant association with HLA-DR4-DQ8 (P = 0.0005). Among relatives developing diabetes, 98% had one or more of IAAs, GAAs, or ICA512bdcAAs, and 80% had two or more of these autoantibodies, compared with none of the control subjects. Using survival analysis to allow for different lengths of follow-up, there was a significant increase in the risk of diabetes with the number of these autoantibodies present, comparing zero, one, two, and three autoantibodies (P < 0.0001, log-rank test), and by Cox regression analysis, this was independent of ICAs and age. For relatives with two or more of these autoantibodies, the risk of diabetes within 3 years was 39% (95% CI, 27-52) and the risk within 5 years was 68% (95% CI, 52-84). Relatives with all three autoantibodies had a risk within 5 years estimated to be 100%. The presence of low first-phase insulin release further increased the risk for relatives with one or two autoantibodies. We conclude that the presence of two or more autoantibodies (out of IAAs, GAAs, and ICA512bdcAAs) is highly predictive of the development of type I diabetes among relatives.

Viral elements in autoimmunity of type I diabetes.

The involvement of viruses in the etiology of insulin-dependent diabetes was hypothesized more than 15 years ago based on solid scientific evidence, however; the true existence and the real nature of this involvement still eludes our efforts. This may be due to the frequently long interval between viral exposure and diabetes onset that makes the direct cause-effect relationship difficult to prove.

Islet cell autoimmunity in white and black children and adolescents with IDDM.

OBJECTIVE: To compare the frequency of islet cell antibodies (ICA) and antibodies to GAD65 and IA-2(ICA512) between black and white children and adolescents at the diagnosis of IDDM in a large consecutive series of cases from Children's Hospital of Pittsburgh. RESEARCH DESIGN AND METHODS: ICA and antibodies to GAD65 and IA-2 were measured in 437 white and black children and adolescents who were diagnosed with IDDM at < 19 years of age at Children's Hospital of Pittsburgh from January 1983 to December 1985, from January to December 1989, and from January 1996 to December 1997. RESULTS: The prevalence of ICA(H), GAD65, and IA-2 antibodies was significantly lower in blacks than whites at onset of the disease. In contrast, the prevalence of ICA(R) alone was higher in blacks. None of the antibodies were present in 12% of the blacks compared with 4% in whites. The same pattern was seen in both sexes. The prevalence of antibodies in white patients with onset of IDDM at <11 years of age was no different than in those who developed IDDM during adolescence. In contrast, black patients showed a significantly lower prevalence of almost all antibodies in the adolescent group. CONCLUSIONS: Black adolescents were more likely to not have antibodies, suggesting either that they have a nonautoimmune type of diabetes or that antibodies are not being detected by these assays.

Identification of 3 beta-hydroxysteroid dehydrogenase as a novel target of steroid cell autoantibodies: association of autoantibodies with endocrine autoimmune disease.

Autoantibodies directed against steroid hormone-producing cells (SCA) detectable by immunofluorescence are typically found in a small proportion of patients with premature ovarian failure (POF) as well as in other endocrine autoimmune diseases. The SCA pattern stains cells in the outer zones of the adrenal cortex, ovary, and testis. To identify the molecular target of SCA, an adrenal complementary DNA expression library was screened using SCA-positive serum, and the steroid enzyme 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) was identified. Only 1 of 48 (2%) patients with idiopathic POF, not pre-selected for the presence of other autoimmune diseases, had SCA by immunofluorescence, whereas 10 of 48 (21%) had anti-3 beta HSD autoantibodies detectable by immunoblot using recombinant human enzyme compared with 6 of 115 (5%) control subjects (P = 0.002). Absorption of SCA-positive serum with recombinant human 3 beta HSD abolished the immunofluorescence pattern. We also examined the prevalence of anti-3 beta HSD autoantibodies in other endocrine autoimmune diseases. Two of 112 (2%) diabetic patients, but none of the thyroid or Addisonian patients, had SCA by immunofluorescence. Twenty-six (23%) diabetic subjects (P < 0.001 vs. controls), 3 of 18 thyroid patients (P > 0.05 vs. controls), and none of 4 Addisonian patients had anti-3 beta HSD autoantibodies. 3 beta HSD is the first steroid cell autoantigen defined at the molecular level to be associated with idiopathic POF occurring in the absence of other polyglandular diseases. Autoantibodies to 3 beta HSD in patients with other organ-specific autoimmune diseases indicate that the enzyme behaves as a typical target of polyendocrine autoimmunity. Anti-3 beta HSD autoantibodies in patients with POF may provide a marker of those subjects whose ovarian failure is autoimmune in origin and, as recent studies suggest, may be salvageable with glucocorticoid treatment.

Sequence analysis of the diabetes-protective human leukocyte antigen-DQB1*0602 allele in unaffected, islet cell antibody-positive first degree relatives and in rare patients with type 1 diabetes.

The human leukocyte antigen (HLA)-DQA1*0102/DQB1*0602/DRB1*1501 (DR2) haplotype confers strong protection from type 1 diabetes. Growing evidence suggests that such protection may be mostly encoded by the DQB1*0602 allele, and we reported that even first degree relatives with islet cell antibodies (ICA) have an extremely low diabetes risk if they carry DQB1*0602. Recently, novel variants of the DQB1*0602 and *0603 alleles were reported in four patients with type 1 diabetes originally typed as DQB1*0602 with conventional techniques. One inference from this observation is that DQB1*0602 may confer absolute protection and may never occur in type 1 diabetes. By this hypothesis, all patients typed as DQB1*0602 positive with conventional techniques should carry one of the above diabetes-permissive variants instead of the protective DQB1*0602. Such variants could also occur in ICA/DQB1*0602-positive relatives, with the implication that their diabetes risk could be significantly higher than previously estimated. We therefore sequenced the DQB1*0602 and DQA1*0102 alleles in all ICA/DQB1*0602-positive relatives (n = 8) previously described and in six rare patients with type 1 diabetes and DQB1*0602. We found that all relatives and patients carry the known DQB1*0602 and DQA1*0102 sequences, and none of them has the mtDNA A3243G mutation associated with late-onset diabetes in ICA-positive individuals. These findings suggest that diabetes-permissive DQB1*0602/3 variants may be very rare. Thus, although the protective effect associated with DQB1*0602 is extremely powerful, it is not absolute. Nonetheless, the development of diabetes in individuals with DQB1*0602 remains extremely unlikely, even in the presence of ICA, as confirmed by our further evaluation of ICA/DQB1*0602-positive relatives, none of whom has yet developed diabetes.

The use of colony stimulating factors in elderly patients with cancer.

Hematological toxicity is the most common and the most frequent fatal complication of chemotherapy. It is observed with increased frequency with age, it is a significant independent predictor of the development of febrile neutropenia, and may contribute to a reluctance to administer chemotherapy in the elderly patient population. The authors analyze published data on effectiveness and results of the use of colony stimulating factors for preventing and treating elderly patients affected by tumors during chemotherapy.

Geriatric oncology: a clinical approach to the older patient with cancer.

Due to the ageing of the population and the sharp increase in life expectancy, cancer in the older person has become an increasingly common problem in the Western world. Although several authors have stressed that elderly cancer patients deserve special attention as a target group for research efforts, older aged patients are still less likely to be offered participation in clinical trials. The cellular and molecular mechanisms regulating the physiological process of ageing and senescence are far from understood, although inflammation is likely to play an important role, at least in some cancers. In addition, the relationship between ageing and cancer risk is also far from understood. One of the most intriguing aspects of ageing is how different the ageing process is from person to person; the basis for this variation is largely unknown. Population-based studies and longitudinal surveys have shown that comorbidity and physical and mental functioning are important risk factors; thus, a meaningful assessment of comorbidity and disability should be implemented in clinical practice. Modern geriatrics is targeted towards patients with multiple problems. Such patients are not simply old, but are geriatric patients because of interacting psychosocial and physical problems. As a consequence, the health status of old persons cannot be evaluated by merely describing the single disease, and/or by measuring the response, or survival after treatment. Conversely, it is necessary to conduct a more comprehensive investigation of the 'functional status' of the aged person. A geriatric consultation provides a variety of relevant information and enables the healthcare team to manage the complexity of health care in the elderly; this process is referred to as the Comprehensive Geriatric Assessment (CGA). The use of CGA is now being introduced into oncological practice. The definition of frailty is still controversial and represents a major issue of debate in clinical geriatrics. As the frail population increases, clinical trials in frail persons are needed. The usefulness of these trials requires a consensus as to the definition of frailty. Clearly, the management of older persons with cancer requires the acquisition of special skills in the evaluation of the older person and in the recognition and management of emergencies as well as experience in geriatric case management.

Autonomic function and autoantibodies to autonomic nervous structures, glutamic acid decarboxylase and islet tyrosine phosphatase in adolescent patients with IDDM.

Recent studies have linked autoimmunity to nervous tissue structures and diabetic autonomic neuropathy, but data on the early stage of IDDM and on the natural history of this association are not available. For this reason, we investigated autonomic nervous function, and the presence of autoantibodies to sympathetic and parasympathetic nervous structures, to glutamic acid decarboxylase (GAD) and tyrosine phosphatase (IA-2/ICA512) in 85 adolescents with insulin-dependent diabetes mellitus (IDDM) (mean age 14.7+/-1.6 yr, mean duration of diabetes 6.8+/-3.5 yr), and 45 age and sex-matched healthy subjects. Nervous tissues autoantibodies were detected using an indirect immunofluorescent complement-fixation technique, with monkey adrenal gland, rabbit cervical ganglia and vagus nerve as substrates. GAD and IA-2/ICA512 autoantibodies were detected by radioimmunoprecipitation assay. Seven patients (8%) had anti-vagus nerve autoantibodies, 7 other patients (8%) had anti-cervical ganglia autoantibodies, while all controls were negative (P < 0.05). Anti-adrenal medulla antibodies were detected in 16 patients (19%) and in 2 control subjects (P<0.02). None of the patients had autonomic symptoms. When patients were divided according to the presence or absence of autoantibodies, values of the cardiovascular tests (deep breathing, 30:15 ratio, Valsalva ratio) were similar in the two groups and similar to those in healthy subjects. However, when considered together, patients positive for one or more autoantibody showed a trend for lower values of deep breathing test and 30:15 ratio test, compared with healthy control subjects, which failed to reach conventional significance values (P=0.17 and P=0.07, respectively). No correlation was found between cardiovascular parameters and metabolic control or diabetes duration. There was no association between autoimmunity to nervous tissue structures and presence of GAD and IA-2/ICA512 Ab, and no correlation between these two autoantibodies and values of cardiovascular tests. Our data indicate that autonomic dysfunction is not a characteristic of young diabetic patients, but that autoantibodies against autonomic nervous structures are present during the first 1 to 15 yr of diabetes. GAD and tyrosine phosphatase appear to be excluded as target autoantigens within autonomic structures. Follow-up studies are required to evaluate future autonomic dysfunction and symptoms in these patients, and to establish whether the subtle autonomic dysfunction detected and/or the nervous tissue autoantibodies, are predictive of the development of this complication.

Identification of a novel serine protease-like molecule in human brain.

Proteolysis of the amyloid beta protein precursor (APP) is a key event in the development of Alzheimer's disease. In our search for proteases that can cleave APP and liberate the amino terminus of the amyloidogenic beta protein, we characterized a calcium-dependent serine protease (CASP) which is present in reactive astrocytes and cross-reacts with anti-cathepsin G antibodies. We wanted to take advantage of this cross-reactivity to clone the cDNA of CASP and eventually evaluate its tissue distribution. Screening of two human fetal brain cDNA libraries with anti-cathepsin G antibodies led to the identification of a cDNA coding for a novel protein whose only homology to known proteins is to the active site of trypsin-type serine proteases. We called this protein the novel serine protease (NSP). NSP exists in at least three differentially spliced forms, one of which is expressed predominantly in brain and testis. Immunohistochemistry and immunoprecipitation with antibodies generated against NSP show that it is expressed and secreted by a variety of cells and that, in brain, it is found primarily in cerebrovascular smooth muscle cells and reactive astrocytes.

Reduced thymic expression of islet antigen contributes to loss of self-tolerance.

Type 1 diabetes (T1DM) results from a failure of central and peripheral tolerance to islet cell antigens. ICA69 belongs to a group of molecules expressed predominantly in neuroendocrine tissues (including pancreatic islets), which are targets of autoimmune responses in T1DM. These molecules are also expressed in the thymus and peripheral lymphoid organs by dendritic cells. The aim of the present study was to evaluate possible variation in thymic ICA69 expression, comparing diabetes-resistant controls to T1DM-prone NOD mice. Thymic tissue was retrieved from 3- to 6-week-old female B6, NOD-H2(b), and NOD mice. Paraffin-embedded sections were stained with an ICA69-specific antibody in an immunoperoxidase assay. ICA69 staining of thymic sections from B6 and NOD.H2(b) showed strong and continual staining, yet the sections from the NOD mice showed significantly reduced staining for ICA69. Corroboration of the reduced level of ICA69 in the thymus of NOD mice has been obtained via analysis for the expression of ICA69 versus other candidate autoantigens (glutamic acid decarboxylase 65, glutamic acid decarboxylase 67, and insulin 2) in the thymus. Real-time PCR analysis, using cDNA generated from the thymus, displayed that the expression of GAD65, GAD67, and INS2 were equivalent when comparing NOD at any age to B6, BALB/cJ, and ALR/LtJ. In marked contrast, the level of ICA69 in the thymus of the NOD mice examined was significantly reduced when compared to the controls. In fact, the real-time PCR analysis strongly suggested that ICA69 was not expressed in the thymus of NOD mice. These findings support the hypothesis that the level of thymic ICA69 expression may be of importance in regulating self-tolerance in T1DM.