Immunological responses of healthy elderly population

We measured in vitro mitogen responses, delayed hypersensitivity skin tests, circulating immune complexes (CIC) and three autoantibodies in 279 healthy and 24 chronically ill individuals over 65, and in young controls. The elderly individuals had previously undergone a complete medical examination and laboratory screening tests, and were on no medications. Compared to the results of 180 young controls tested concurrently, the healthy elderly individuals had significantly depressed PHA responses and skin test responses. In addition, CIC and autoantibodies were increased in the healthy elderly group compared to young controls. There was no difference in PHA or skin test responses between the healthy and chronically ill elderly subjects, suggesting that the major determinant of depressed cellular immunity in the elderly is age per se and not age-associated diseases. Within the elderly population, aged 65-94, there was a significant decrease in PHA response with age. Previously it has been reported that correlations exist between measurements of cellular immune response (mitogen response and skin testing) and manifestations of autoimmunity (CIC or autoantibodies) in elderly subjects. However, in this well characterized healthy elderly population we could not verify an association between the cellular immune response and either autoantibodies or immune complexes. In addition, was no increased prevalence of autoantibodies in subjects with CIC.     

Soluble immune complexes in the sera of newly diagnosed insulin-dependent diabetics and in treated diabetics

Soluble immune complexes were detected by the Raji cell assay in seven out of thirteen newly diagnosed insulin-dependent diabetics, six of whom had islet cell antibodies (ICAb) in the serum. None of the others had ICAb. The titres for a wide range of viral antibodies were similar in these thirteen diabetics as in age- and sex-matched controls, except for antibodies to Coxsackie B4. Six had titres of Coxsackie B4 antibodies greater than or equal to 1:32, but only three of these had evidence of immune complexes in the serum, and these were not correlated with the titres of Coxsackie B4 antibodies. None of these diabetics had antibodies to insulin and none of their age- and sex-matched controls had evidence of immune complexes in the serum.

53% of thirty-two diabetics treated with insulin and 10% of fifty-two diabetics requiring oral hypoglycaemic agents (OHA) or diet had evidence of immune complexes in the serum, compared to 6% of control subjects. High titres of insulin antibodies correlated with evidence for immune complexes. There was a stronger tendency for immune complexes to occur in the presence of moderate titres of insulin antibodies when the age at onset of insulin-dependent diabetes was less than 30 years. Out of sixteen patients treated with heterologous insulin for 13 years or more, and who also had late diabetic complications, twelve had immune complexes in the serum.

     

Occurrence of anti-low density lipoprotein antibodies and circulating immune complexes in aged subjects

The incidence of circulating immune complexes, anti-low density lipoprotein (LDL) autoantibodies and the anti-LDL activity of immune complexes was studied in healthy young and aged controls and in patients with vascular diseases. Circulating immune complexes (CIC) frequently occurred both in the young or old patient groups and in the aged healthy control groups, whereas they could not be found in the young controls. Marked differences were found in the incidence of anti-LDL antibodies between the groups tested. In both young and aged control groups such antibodies were very rarely observed (4-5%). In contrast anti-LDL antibodies were present in 35-45% in the aged, or young patients. Similarly, no anti-LDL activity was found in CIC of the controls, whereas in the patients with vascular diseases a significant CIC-associated anti-LDL activity was detected. These results suggest that the presence of anti-LDL antibodies are associated with the arteriosclerotic manifestations, while that of circulating immune complexes is connected by the ageing process itself.     

Circulating immune complexes among diabetic children

Insulin dependent diabetes mellitus (IDDM) is an autoimmune disease associated with the presence of different types of autoantibodies. The presence of these antibodies and the corresponding antigens in the circulation leads to the formation of circulating immune complexes (CIC). CIC are known to persist in the blood for long periods of time. Such CIC following deposition in the small blood vessels have the potential to lead to microangiopathy with debilitating clinical consequences. The aim of our pilot study was to investigate whether a correlation exists between CIC and the development of microvascular complications in diabetic children. Isolation of a new glycoprotein complement inhibition factor (CIF) from the parasitic plant Cuscuta europea seed, which appears to bind specifically to complement component C3 has provided an unique tool for the measurement of immune complexes by means of ELISA-type techniques (CIF-ELISA). We studied the levels of CIC (IgG, IgM and IgA) in 58 diabetic children (mean age 12.28 +/- 4.04 years, diabetes duration 5.3 +/- 3.7 years), 29 of them had vascular complications (group 1) and the other 29 were without vascular complications (group 2). As controls, we studied sera samples from 21 healthy children (mean age 13.54 +/- 4.03 years). Sera from the diabetic patients showed statistically significant higher levels of CIC IgG (p = 0.03) than sera from the control group. In sera from group 1 values of CIC IgG showed statistically significant higher levels than controls (0.720 +/- 0.31 vs. 0.46 +/- 0.045; p = 0.011) Sera from 59% of the patients were positive for CIC IgG, 36% for CIC IgM and 9% for CIC IgA. Among 26 patients with microalbuminuria, sera from 17/26 (65%) were positive for CIC IgG, 8/26 (31%) for CIC IgM and 2/26 (8%) for CIC IgA. CIC IgG correlated with HbAlc (r = 0.51; p = 0.005) and microalbuminuria (r = 0.42, p = 0.033). CIC IgA correlated with age (r = 0.44, p = 0.03). CIC IgM correlated with the duration of diabetes (r = 0.63, p = 0.02). These findings suggest that elevated levels of CIC IgG are associated with the development of early diabetic nephropathy.     

Circulating immune complexes and cell-mediated immunity in patients with hepatitis B virus associated liver diseases

The prevalence of circulating immune complexes (CIC), their role and their relationship to cell-mediated immunity in patients with hepatitis B virus associated liver disease are still controversial. This study was designed to investigate the prevalence of CIC and their relationship to viral markers, to subsets of peripheral blood T lymphocytes and to suppressor cell activity in patients with hepatitis B virus associated liver diseases. CIC were positive in 29.3% of 41 healthy HBsAg carriers, 37.8% of 88 patients with hepatitis B virus associated liver diseases, and 15% of 41 healthy subjects by the platelet aggregation test (PAT). The prevalence of CIC in patients with acute hepatitis (40.0%) and in those with cirrhosis (61.5%) was significantly higher than in normal controls (p less than 0.05, p less than 0.005 respectively). There was no correlation between the titer of CIC and serum HBsAg titer or the status of HBeAg, and no significant decrease in the peripheral blood lymphocyte CD4/CD8 ratio in healthy HBsAg carriers (1.39 +/- 0.31) and in patients with liver diseases (1.40 +/- 0.54) compared to the normal controls (1.48 +/- 0.31). Concanavalin A induced suppressor cell activity on IgG producing cells was impaired in healthy HBsAg carriers (34.9%) (p less than 0.005) and in patients with liver diseases (25.3%) (p less than 0.0001), and this change was prominent in patients with chronic active hepatitis and cirrhosis (p less than 0.0001). And there was a significant reverse correlation between concanavalin A induced suppressor cell activity on IgG-producing cells and the titer of CIC in PAT positive patients with hepatitis B virus associated liver diseases. In conclusion, it was suggested that defective suppressor cell function may lead to an increased B cell activation and such activity may account for the presence of CIC.     

Cell-mediated immunity in diabetes mellitus

The quantified lymphocyte response to phytohaemagglutin (PHA) was investigated in fifty-six control subjects, classed according to age and sex, in twenty-nine insulin-dependent (IDD) and eight non-insulin-dependent diabetics. In nine cases one-way mixed lymphocyte cultures (MLC) were performed using mixed lymphocytes from IDD and paired controls. In the controls, a reduction of the response to PHA was observed with ageing; this was more marked in women than in men. The diabetics gave a greatly impaired response to PHA but only a slightly depressed response in MLC. These anomalies are not influenced by age but by the severity of the diabetes.     

Physiological aspects of circulating immune complexes in the normal population.

Circulating immune complexes (CIC) have been investigated in 100 normal subjects; the RIA-Raji and the C1q-BA conventional methods, as well as a new solid phase microassay utilizing purified C1q and the systematic search of cryoprecipitates were employed. CIC serum levels did not differ in regards to sex; in relation to age, values for C1q-BA were identical in subjects from 0 to 60 years and also in those beyond age 60; the differences encountered by RIA-Raji or by the C1q-SP microassay in these two main groups were not statistically significant. Cryoprecipitates were present in 100% of the 68 examined subjects. Immunoglobulins (G, A and M), anti-nucleic acid (DNA and Poly A) and CIC (by the three methods) were present in the cryoprecipitates while lymphocytotoxins, rheumatoid factor and C3 were undetectable; protein content of the cryoprecipitates increased significantly with age, reaching a normal superior limit of 0.52 mg/ml beyond age 30. These findings further support the role played by CIC in normal immune response and may help in the understanding of the physiopathology of clinical conditions associated with immune complexes.     

Factors affecting the insulin autoantibody ELISA

IgG antibodies to insulin are present in insulin-treated patients and are detected in the prodrome of untreated type I diabetes. Sporadic reports of autoantibodies to insulin suggest that they are also present in other disorders. To establish the incidence of insulin autoantibodies in other endocrine and autoimmune diseases an ELISA was used to examine sera from 529 subjects with no prior insulin therapy. These untreated patients included: normal controls (adults and children), newly-diagnosed type I diabetes, first-degree relatives of diabetics, type II diabetes, Graves' hyperthyroidism, and systemic lupus erythematosus. As a positive control group, 280 insulin-treated patients were studied. Measurement of IgG antibodies by direct binding to insulin coated plates was complicated by differences between adult and pediatric populations and by overlap of binding between treated and untreated subjects. Competitive inhibition with excess soluble human insulin overcame these problems and permitted identification of insulin specific binding. Using this approach insulin antibodies were most frequent in insulin-treated diabetics (98%) and in type I diabetics (37%) prior to treatment. The absolute numbers of subjects with insulin autoantibody in the other groups differed depending upon whether a cut-off for binding (mean + 2SD of controls) or for insulin inhibition of binding (45%) was used. Regardless of the criteria used there were subjects (2-24%) in all groups tested with circulating insulin-specific IgG autoantibody detected by ELISA. These low level antibodies detected in solid phase assays may be part of the normal immune repertoire.     

Pancreatic islet and other autoantibodies in juvenile and adult onset diabetics in Australia

The prevalence of serum antibodies to the cytoplasm of the pancreatic islet cell (PICA), and of thyroid microsomal (TMA), gastric parietal cell (GPCA) and anti-nuclear (ANA) antibodies was studied in 135 newly diagnosed diabetics presenting to a hospital for adults and 83 children with recent onset diabetes presenting to a children's hospital. The study also included another 144 diabetic children whose disease had been present longer, and 200 control children. There was a high prevalence (87%) of PICA in the children whose diabetes had just been diagnosed in comparison with control children (1%):(P less than .0001). Diabetic children also had a high prevalence (21%) of one or more of the other autoantibodies in comparison with the control children (9%):(P less than .001). Only 26% of the 58 insulin dependent adults had PICA but 33% had other autoantibodies. Two (3%) of the 77 adult diabetics who did not require insulin had PICA; 8% had other autoantibodies.     

Antisperm Antibodies and Circulating Immune Complexes of Vasectomized Men With and Without Coronary Events

ABSTRACT: We compared antisperm antibody and circulating immune complex (CIC) levels in serum samples from 101 vasectomized and 101 normal age-matched nonvasectomized men; 31 of each group had histories of coronary heart disease (CHD). Vasectomy and CHD status were treated as categorical independent variables in the two-way analysis of variance. Elevations of both systolic and diastolic blood pressures were significantly associated with age and body mass index but not vasectomy. Antisperm antibodies (immobilizing and agglutinating) were significantly associated with vasectomy (P ≤ .001); the incidences were similar in men with and without CHD. The CICs were significantly associated with vasectomy in a Staphylococcus aureus (FcSa) CIC assay (P ≤ .001) and a Raji cell CIC assay (P ≤ .05). A third CIC assay, the Clq binding asssay, did not reveal a difference between any subgroups. Generally, CICs occurred more frequently in the CHD group by the FcSa assay and particularly the Raji cell assay (P ≤ .001). In summary, vasectomized men had a higher incidence and higher levels of circulating antisperm autoantibodies and CICs than did age-matched controls.     

Circulating immune complexes in various thyroid diseases.

In a study of 171 patients with various thyroid diseases, circulating immune complexes (CIC), measured by a C1q solid phase radioassay, were detected in 26% of the patients as compared to 8% of the control subjects. CIC were found in 33--55% of the patients with a well defined thyroid autoimmune disorder (Hashimoto's goitre, asymptomatic thyroiditis, spontaneous myxoedema and Graves' disease) and also in the same proportion of patients with diffuse goitre. CIC were correlated to the presence of serum antibodies to microsomal thyroid antigen but not to their titre. No relationship was observed between CIC and the age or sex of the patients and the presence of exophthalmos, or between CIC and the different thyroid function tests or serum anti-thyroglobulin antibodies. CIC were found in untreated patients as well as in those treated with prednisone, methimazole or thyroxine.     

Autoantibodies and circulating immune complexes in sera from patients with hepatitis B virus-related chronic liver disease.

Sera from 56 patients with hepatitis B virus-related chronic liver disease (CLD) and 30 normal individuals as controls were examined by enzyme immunoassay (EIA) for the presence of autoantibodies directed against actin, tubulin, myosin, double-stranded (ds) DNA, polymerized human albumin, thyroglobulin, and trinitrophenyl coupled to bovine serum albumin (TNP-BSA). Patients with CLD had consistently elevated levels of IgG, IgA, and IgM antibodies directed against all the panel antigens. The percentage of patients with autoantibodies of the IgA class was particularly high: respectively, 88 and 78% of the patients had strikingly high levels of anti-actin and anti-TNP-BSA IgA autoantibodies. High amounts of IgA and IgG antibodies to polymerized albumin as well as IgG to thyroglobulin were also detected. Circulating immune complexes (CIC) were isolated from patients' sera and their autoantibody activities were tested on the same antigen panel. The autoantibodies thus detected were of the same class and possessed the same activities, although at higher values than those present in the homologous sera. These results indicate that, regardless of their origin, autoantibodies are present in high amounts in the sera of these patients. Moreover, autoantibodies participating in the formation of CIC might play a pathological role.     

Low pancreatic lipase in insulin-dependent diabetics.

Serum samples obtained from 20 insulin-dependent diabetics (IDD), 23 non-insulin-dependent diabetics (NIDD) and 30 controls were assayed for their pancreatic lipase activity, immunoreactive trypsin concentration and glycosylated haemoglobin (HbA1) respectively. The distribution of serum pancreatic lipase activity in normal subjects and diabetics was nonparametric. The median serum lipase activity in IDDs (86 U/l) was significantly lower that that in controls (131 U/l, p less than 0.002) and NIDDs (126 U/l, p less than 0.001). There was a significant correlation between serum pancreatic lipase activity and serum IRT concentration (r = 0.65, p less than 0.001). Neither pancreatic lipase activity nor IRT was related to HbA1 concentrations. These data show for the first time that serum pancreatic lipase activity is diminished in IDDs.     

Anti-C1q autoantibodies from active lupus nephritis patients could inhibit the clearance of apoptotic cells and complement classical pathway activation mediated by C1q in vitro

Anti-C1q antibodies are prevalent in patients with active lupus nephritis and were found to be closely associated with renal involvement and predictive for a flare of nephritis. However, the pathogenesis of anti-C1q antibodies involved in human lupus nephritis remains unclear. C1q, which plays a key role in apoptotic cell and immune complex removal, is a very important functional molecule in the pathogenesis of SLE. The aim of this study was to investigate the influence of anti-C1q autoantibodies from active lupus nephritis patients on the bio-functions of C1q in vitro. We purified IgG autoantibodies against C1q from lupus nephritis patients, and found that they could recognize C1q bound on early apoptotic cells at 30 μg/ml, and could significantly decrease the phagocytosis by macrophages of early apoptotic cells opsonized by 50 μg/ml C1q in comparison with normal IgG. Levels of circulating immune complexes of the ten patients were measured by a circulating immune complexes (CIC)-C1q Enzyme Immunoassay Kit. Anti-C1q autoantibodies affinity purified by microtiter plates could significantly inhibit the deposition of C3c on CIC-C1q in a dose dependent manner in comparison with IgG from 10 healthy blood donors. The binding of opsonized immune complexes to RBCs was significantly inhibited by anti-C1q autoantibodies purified by microtiter plates in a dose dependent manner. Our observations suggest that serum anti-C1q autoantibodies from active lupus nephritis patients could interfere with some biological function of C1q in vitro.     

Circulating Immune Complexes and Rheumatoid Factors in Patients with Farmer''s Lung

Circulating IgG immune complexes (CICs), IgM rheumatoid factors (RFs), complement level (C3 and C4), and IgG antibodies to Aspergillus fumigatus, Aspergillus umbrosus, Thermoactinomyces vulgaris, and Micropolyspora faeni were analysed in the sera of 14 patients with farmer's lung (FL), 10 in the acute and four in the subacute phase of the disease. Ten spouses of FL patients served as exposed healthy controls. C3 and C4 were measured fluoronephelometrically. C3 levels were above the normal range and C4 levels near the upper limit of the normal range in both patients and controls; no statistically significant difference between patients and controls were observed. CICs were determined by enzyme immunoassays (EIA) of conglutinin-binding (KgB) and Clq-binding (ClqB). CIC levels above the normal range were detected in 12 (KgB-EIA) and nine (ClqB-EIA) of the patients and three (KgB-EIA) and six (ClqB-EIA) of the controls. No statistically significant differences were found in the mean levels between patients and controls. In contrast, RF levels in the acute phase of FL were significantly higher in the patients (P less than 0.01) than in the controls. CICs correlated positively with most microbial IgG antibodies, but negatively with RFs. RFs also correlated negatively with microbial IgG antibodies, as well as with both C3 and C4. In FL, the increased RF level may, in the absence of increased CIC and decreased complement levels, represent an immune response (IgM anti-IgG autoantibodies) induced by local mechanisms of IgG immune complexes in the lungs.     

GAD65 Autoantibodies Increase the Predictability but not the Sensitivity of Islet Cell and Insulin Autoantibodies for Developing Insulin Dependent Diabetes Mellitus

The clinical onset of insulin-dependent diabetes (IDD) can be predicted by determination of autoantibodies to several pancreatic-islet cell antigens. Islet-cell autoantibodies (ICA) and insulin autoantibodies (AA) are most commonly used. We have developed a recombinant human glutamic acid decarboxylase (GAD₆₅) radioimmunoassay and measured autoantibodies to GAD₆₅ (GAD₆₅A) in the sera of 73 documented prediabetic individuals, 76 newly-diagnosed patients, 103 relatives of IDD probands at increased risk for the development of IDD because they were positive for ICA and/or IAA, 72 ICA and IAA negative relatives, and 207 healthy controls. Our data demostrated that GAD₆₅A are strongly associated with the currently established autoantibody markers of IDD. Their presence in prediabetic subjects with only ICA or IAA enhances their risk for progression to IDD, yet does not much enhance the screening sensitivity already available through conventional ICA and IAA for IDD prediction.     

Immune complex-mediated inhibition of lymphocyte Fc-gamma receptors in the plasma of patients with type 1 (insulin-dependent) diabetes mellitus: association with anti-ssDNA antibodies

Sera of 43 patients with type 1 (insulin-dependent) diabetes and from 42 normal subjects were examined for the presence of Fc-gamma receptor-blocking IgG immune complexes, detected by EA rosette inhibition, and for the presence of anti-ssDNA antibodies by ELISA. Forty-four percent of diabetic patients had levels of inhibition above the upper limit of normality in comparison to 7% of normal controls. Anti-ssDNA antibodies were found in the sera of 14 out of 43 diabetic patients. Ten out of 14 anti-ssDNA positive patients (71.5%) had inhibition levels above the upper limit of normality in comparison to 9 out of 29 (31%) of the anti-ssDNA negative population. The difference was statistically significant (P less than 0.005). Levels of EA rosette inhibition were found to be high in patients with duration of diabetes of less than 2 years and correlated with high prevalence of anti-ssDNA antibodies. The percentage of EA rosette inhibition was found not to correlate with the levels of C1q-binding immune complexes, suggesting that immune complexes detected by EA rosette inhibition may belong to a pool of noncomplement-fixing immune complexes. The possible role of immune complexes with autoantibodies anti-ssDNA in the mechanism triggering and perpetuating autoimmune phenomena in diabetes is discussed.     

Circulating immune complexes in patients with diabetes mellitus.

Circulating immune complexes (CIC) were detected by the solid phase C1q binding assay in 16% of 103 diabetic patients and by the fluid phase C1q binding assay in 31% of patients as compared to 5% of 58 control subjects for each assay. Plasma glucose determinations revealed that most patients were moderately hyperglycaemic (mean glucose = 264 mg/dl), and thus were not selected for tight metabolic control. All but six patients had elevated levels of plasma insulin, including both the insulin treated and diet treated subgroups. There was no correlation between the presence of CIC detected by either assay and plasma glucose, insulin, or the presence of microangiopathy. Multiple factors must contribute to the increase in CIC in both insulin deficient and insulin resistant diabetics. The role of these various factors remains to be defined.     

Circulating immune complexes in sera of patients infected with Echinococcus granulosus

Circulating immune complexes (CIC) were investigated by the C1q binding assay in sera of 23 patients infected with Echinococcus granulosus. For the 23 sera studied, nine were found to be positive in the test. When the samples were grouped according to the cyst localization, the highest rate of CIC positively was found in the group of sera from patients with pulmonary cyst; in this group, double diffusion (DD) and indirect haemagglutination (IHA) tests gave a low rate of positivity for antibodies directed against parasitic antigens. Rheumatoid factor, anti-nuclear and anti-mitochondrial autoantibodies were not detectable in patients' sera by indirect immunofluorescence technique (IIF). Anti-smooth muscle autoantibodies, detectable by IIF, were present in 56% of the sera and this positivity was higher in the hepatic (83%) than in the pulmonary form (40%).     

Circulating autoantibodies in patients with aspirin-intolerant asthma: an epiphenomenon related to airway inflammation

Several studies have suggested the involvement of an autoimmune mechanism in aspirin (ASA)-intolerant asthma. To test this hypothesis, we measured the levels of circulating autoantibodies, such as IgG and IgA to tissue transglutaminase (TGase), IgG to cytokeratins (CKs) 8, 18, and 19, Clq-binding immune complex (CIC), and antinuclear antibody (ANA), in the sera of 79 patients with ASA-intolerant asthma (Group I) and those of two control groups, consisting of 61 patients with ASA-tolerant asthma (Group II) and 88 healthy control subjects (Group III) by means of ELISA. Significantly higher prevalences of IgG antibodies to CK18 (13.9%) and CK19 (17.7%) were noted in Group I, as compared with Group III (p<0.05 for all) not with Group II. Regarding the prevalences of other autoantibodies, the levels of ANA (1.3%), IgG to TGase (3.8%), and CIC (24.7%) in Group I were not significantly different from those in Groups II and III. Significant correlations were found between positivities for the anti-CK18 and anti-CK19 autoantibodies and the PC(20) methacholine values in the analysis of asthma Groups I and II vs. normal controls, (p=0.001 and p=0.003, respectively). Further studies are needed to explore the potential involvement of an autoantibody-mediated mechanism in the clinical manifestation of bronchial asthma.