Research of anti-GAD and anti-IA2 autoantibodies by ELISA test in a series of Moroccan pediatric patients with diabetes type 1.

INTRODUCTION: Type I diabetes (T1D) is an autoimmune disease with a prediabetic, asymptomatic period characterized by the selective destruction of insulin-producing ß cells. During the pre-clinical phase, various auto-antibodies are generated against several beta cell antigens such as anti glutamate acid decarboxylase (Anti-GAD), anti tyrosine phosphatase (Anti-IA2). Today, the coupled detection of Anti-IA2 with that of Anti-GAD proves its great importance in the diagnosis and prediction of type 1 diabetes. The combined positivity for both antibodies has a specificity and a positive predictive value of 100%. OBJECTIVES: In this work, we evaluate the diagnostic value of anti-GAD and anti-IA2 antibodies in a series based on 78 Moroccan subjects initially under 16, suspected T1D. RESULTS AND DISCUSSION: Our series consists mainly of 74% of newly diagnosed patients for T1D and 26% of confirmed diagnostic patients, of whom 52% are females. The mean age of diagnosis is 7 ± 4 years, the mean of HbA1c at the time of diagnosis is 11.63 ± 2.16%, and the percentage of family history in our series is 69%. The proportion of positive results for anti-IA2 antibodies and anti-GAD antibodies are, respectively, 76.92% and 62.82%, and 52.56% of patients are positive for both auto-antibodies. This study confirms that anti-GAD and anti-IA2 auto-antibodies assays can detect patients early and the autoantibodies can persist several years after diagnosis of type 1 diabetes. CONCLUSION: This study confirmed the diagnosis and classification of T1D (type 1A) in 87.18% of patients, and we reported that the prevalence of anti-GAD and anti-IA2 is higher in girls than in boys.

Probucol promotes reverse cholesterol transport in heterozygous familial hypercholesterolemia. Effects on apolipoprotein AI-containing lipoprotein particles.

In order to investigate the effect of Probucol therapy on reverse cholesterol transport, apo AI-containing lipoprotein particles were isolated and characterized, and their cholesterol effluxing capacity and LCAT activity were assayed in four familial hypercholesterolemia patients before and after 12 weeks of Probucol therapy. Four major subpopulations of apo A-containing lipoprotein particles are separated before and after drug treatment; LpAI, LpAI:AII, LpAIV, LpAI:AIV:AII. Probucol reduces both total plasma and LDL-cholesterol (-17 and -14%, respectively). Apo B decreases slightly (-7.6%). Plasma HDL-cholesterol and apo AI decrease by 36.6 and 34.7%. LpA-I showed a marked decrease (-46%). Moreover, plasma LCAT and CETP activities were markedly increased under Probucol treatment. Analysis of lipoprotein particles showed that Probucol induces a decrease of protein content and an increase of cholesterol and triglycerides contents. Interestingly, Probucol induces an enhancement of LCAT activity in LpAI (4.5-fold). This drug induces a trend toward greater cholesterol efflux from cholesterol-preloaded adipose cells promoted by Lp AI and Lp AIV but not by Lp AI:AII. This study confirms the hypothesis, in addition to the lowering LDL-cholesterol levels and antioxidant effects of Probucol, that HDL reduction was not an atherogenic change in HDL system but may cause an antiatherogenic action by accelerating cholesterol transport through HDL system, promoting reverse cholesterol transport from peripheral tissues.

[Diagnostic strategy in amyloidosis]. / Stratégie diagnostique des amyloses.

INTRODUCTION: The diagnosis of amyloidosis is a four-step strategy requiring: 1) a high degree of suspicion, as the clinical presentation of the disease is very polymorphous; 2) the biochemical nature of the disease; 3) the evaluation of the disease spread and 4) how it evolves. EXEGESIS: Simple and noninvasive biopsies usually make it possible to diagnose amyloidosis. Available treatments of generalized amyloidosis require an exact characterization of the nature of amyloid deposits, which is based on the clinical context, immunohistochemical analysis of amyloid deposits, and genetic testing. CONCLUSION: Management of amyloidosis should be improved by better characterization of amyloid deposits. This would result in epidemiological data which are currently lacking.

[Treatments for amyloidosis beyond symptomatic care]. / Traitements des amyloses en dehors du traitement symptomatique.

INTRODUCTION: Amyloid syndromes are pathogenetically different, each of the various amyloid diseases requiring specific treatment. Unfortunately, those treatments are often preventive and symptomatic, some efficient therapies being limited to particular types of amyloidosis. CURRENT KNOWLEDGE AND KEY POINTS: Colchicine is effective in the prevention of amyloidosis due to familial Mediterranean fever but is less or not effective in other situations. Cytotoxic agents are useful in the treatment of AL amyloidosis with or without hemopoietic stem cell transplantation. Liver transplantation is indicated for familial polyneuropathy and kidney transplantation for dialysis-related beta 2 microglobulin amyloidosis. FUTURE PROSPECTS AND PROJECTS: In vivo binding of serum amyloid P (SAP) (component shared by all amyloid deposits) to amyloid fibril, is a new avenue in the therapeutic approach. Development of radiolabeled SAP scintigraphy allows assessment of the disease outcome and evaluation of treatment-related effects. The various treatments that were assessed until now with the objective of curing the disease are reviewed.

[Mechanisms of amyloidosis and the proteins involved]. / Mécanismes de l'amylose et protéines impliquées.

INTRODUCTION: Recent data in amyloid research have shed light on the amyloid substance and have broadened our knowledge on the mechanism of amyloid deposition. CURRENT KNOWLEDGE AND KEY POINTS: Despite uniform physical properties relating to the presence of beta-pleates, amyloid deposits are chemically heterogeneous and have different origins; additional types will probably be described in the future. Immunohistochemical techniques using specific antisera for each of the major protein present in fibrils could help greatly to subclassify these disorders. In most circumstances, a circulating precursor protein may result from overproduction of either intact or aberrant molecule, a reduction in its degradation or excretion, or genetic abnormalities associated with variant proteins. The cleavage of protein precursor molecules of the protein component of amyloid fibrils characterizes amyloidogenesis, though it is not necessary for some amyloidosis forms. This review summarizes advances in the understanding of the nature of amyloid substances, the mechanism of amyloid deposition and the principal pathogenic hypothesis. FUTURE PROSPECTS AND PROJECTS: SAP component is common in all amyloidosis and may be the target for future therapy.

Beneficial effect on serum apo AI, apo B and Lp AI levels of Ramadan fasting.

In order to investigate for the first time in Morocco the effect of fasting in Ramadan, the ninth lunar month of the muslim year, on lipoprotein metabolism, we determined the levels of serum apolipoproteins; apolipoprotein AI (apo AI), apo B, apo AIV and those of lipoprotein particles; apo AI-containing lipoprotein particles (Lp AI) and also apo AI and apo AII containing lipoprotein particles (Lp AI:AII) in a group of 32 healthy, volunteer adult males. Determination of all these parameters was carried out on each week of the month of Ramadan and the results are compared with the pre-fasting and the post-fasting values. Ramadan fasting reduces significantly serum apo B (P < 0.05), while serum apo AI is significantly increased (P < 0.05) compared with the pre-fasting period. The increase of apo AI occurred on day 29 of Ramadan by 11.8%. Serum apo AIV was unchanged during the fasting period indicating that food intake during Ramadan is not based on lipid diet. The observed diet pattern during Ramadan showed an increase of total energy intake based on carbohydrates (+1.4% of total energy), proteins (+0.4% of total energy) but not on fat (-0.7% of total energy), compared with a usual diet used in the rest of the year. The fat diet is high in monounsaturated (P < 0.05) and polyunsaturated fatty acid in contrast to saturated fatty acid which decreased (P < 0.05) during Ramadan. On the other hand, analysis of serum Lp AI and Lp AI:AII showed that the levels of Lp AI:AII were unchanged but those of Lp AI were significantly increased (P < 0.01) at the end of Ramadan. These findings show that feeding behaviour that occurs during Ramadan beneficially affects serum apolipoprotein metabolism and may contribute to prevention of cardiovascular diseases.

Apolipoproteins and lipoprotein particles in Moroccan patients with previous myocardial infarction.

We investigated for the first time in the Moroccan population the relationship between lipoprotein particles and the progression of coronary atherosclerosis. Plasma lipid variables, including total cholesterol, triglycerides, high-density lipoprotein-cholesterol, low-density lipoprotein-cholesterol, apolipoproteins AI and B, Lp AI, Lp AI: AII, and Lp(a) were measured in 40 Moroccan adults who suffered a verified myocardial infarction before the age of 50 years. The results were compared with a healthy control group. Plasma total cholesterol, triglyceride, and Lp AI: AII levels of patients did not differ significantly from control subjects. Patients had lower plasma high-density lipoprotein-cholesterol (P < 0.05), apo AI (P < 0.05), and Lp AI (P < 0.001) than control subjects, suggesting that the cholesterol reverse transport system is altered in patients with previous myocardial infarction. However, patients had higher plasma low-density lipoprotein-cholesterol (P < 0.001), apo B (P < 0.001), and Lp(a) (P < 0.001). In all patients the best predictor of cardiovascular risk was the independent risk factor Lp(a) plasma level, and the Lp AI plasma level. In this study, the increased coronary atherosclerosis risk with elevated plasma levels of apo B and Lp(a), and with reduced Lp AI, was substantially modified by smoking habits, but not by family history of myocardial infarction.

Fasting during Ramadan induces a marked increase in high-density lipoprotein cholesterol and decrease in low-density lipoprotein cholesterol.

We demonstrated for the first time in a Moroccan population that fasting during Ramadan, the ninth lunar month of the Muslims' year, affected lipid and lipoprotein metabolism in a group of 32 healthy adult male volunteers. This investigation was conducted to study the changes in serum total cholesterol, triglycerides, cholesterol in high-density lipoprotein (HDL) and low-density lipoprotein (LDL), glucose, and body weight during Ramadan. The results showed a significant decrease (7.9%, p < 0.001) in serum total cholesterol concentration during Ramadan as compared with the prefasting period. Also, we obtained a significant decrease of serum triglyceride concentration (30%, p < 0.001) during Ramadan fasting as compared to the period before Ramadan. The reduction of both serum triglycerides and total cholesterol was maintained 1 month after Ramadan. By the end of Ramadan, serum HDL cholesterol had markedly increased (14.3%, p < 0.001) and remained elevated 1 month after Ramadan in contrast to LDL cholesterol which showed a significant decrease (11.7%, p < 0.0001) also maintained 1 month after Ramadan. Mean body weight declined by 2.6% (p < 0.01) on day 29 of Ramadan, whereas during Ramadan, the diet pattern used by our subjects showed an increase of total energy intake due to carbohydrates (+ 1.4% of total energy), proteins (+ 0.4% of total energy) but not fat (-0.7% of total energy) compared to a usual diet used throughout the rest of the year. Moreover, the fat diet is high in monounsaturated (p < 0.05) and polyunsaturated fatty acid in contrast to saturated fatty acid which significantly (p < 0.05) decreased during Ramadan. These findings suggest that feeding behavior that occurs during Ramadan beneficially affects plasma lipids and lipoproteins.

Apolipoprotein AIV of human interstitial fluid is associated with apolipoprotein AI-containing but not with AII-containing particles.

Apolipoproteins and lipoprotein particles from human interstitial fluid and plasma were analyzed. The interstitial fluid was enriched in apolipoproteins AI, AII, and AIV compared with apo B, apo CIII, and apo E, LpAI was found to contain apo AIV which was absent from LpAI: AII. Moreover, the bulk of lecithin-cholesterol acyl-transferase was present in LpAI. The concentration range of these particles was in agreement with those required in vitro for cholesterol efflux. Thus the interstitial fluid contains particles in which two agonists but no antagonists of cholesterol efflux are associated with lecithin-cholesterol acyltransferase activity. This supports apolipoprotein AI- and/or AIV-containing particles playing a critical role in the first step of reverse cholesterol transport.

[Systemic amyloidosis in the elderly: diagnostic value of the test of subcutaneous abdominal fat and the labial salivary glands. Prospective study in 100 aged patients]. / L'amylose systémique du sujet âgé: valeur diagnostique de l'examen de la graisse sous-cutanée abdominale et des glandes salivaires accessoires. Etude prospective chez 100 patients âgés.

Primary and secondary amyloidosis are not uncommon in aging but the diagnosis is rarely made on account of the risk of bleeding in the site of biopsies and the difficulty to distinguish senile from systemic amyloidosis on the biopsy samples. We have studied the frequency of amyloid deposition in the abdominal fat aspirate (AFA), the labial salivary gland (LSG), the temporal arteries (four cases), bone marrow (two cases), digestive tract (four cases) in 100 elderly patients (aged 80 or greater). AFA was positive in 15 percent of the patients and LSG in 5%; both samples were positive in 4%. Four cases of systemic amyloidosis were found (two of the AL and two of the AA type). Sensitivity of AFA was 75%, specificity was 87% and the positive predictive value was 20%. The values were respectively 100%, 99%, 100% for LSG. In 11 patients whose AFA biopsies samples were singly positive, amyloid deposits were found in temporal arteries in four of four cases. We conclude that AFA is too sensitive for the diagnosis of systemic amyloidosis in aging. The responsibility of senile amyloid deposition on AFA should require further investigations. LSG biopsies seem to be a more reliable test for the diagnosis of primary and secondary amyloidosis in elderly.

[Value of the assay of protein SAA and urinary antitrypsin activity in osteoarticular infections]. / Intérêt du dosage de la protéine SAA et de l'activité antitrypsique urinaire dans les infections ostéo-articulaires.

Erythrocyte sedimentation rate is normal in 20% to 25% of patients with discitis due to common pathogens. We evaluated serum amyloid A (SAA) protein and urinary trypsin inhibitory activity in osteoarticular infections comparatively with erythrocyte sedimentation rate and serum C-reactive protein in 20 patients including 14 with discitis due to common pathogens and 6 with septic arthritis. Assays were performed on D0, D8, D15, D30, and D60 after initiation of antimicrobial therapy. On D0, all four markers were significantly higher in patients with septic arthritis than in patients with discitis. C-reactive protein levels exhibited the fastest kinetics with a return to normal values within 15 days in both conditions. Urinary trypsin inhibitory activity was only slightly elevated in patients with discitis and returned to normal within 30 days in both conditions. Serum amyloid A levels required 30 to 60 days to return to normal. Erythrocyte sedimentation rate exhibited the slowest kinetics, with normal values being achieved only after 60 days. Although simple, rapid, and inexpensive, urinary trypsin inhibitory activity determination exhibits poor sensitivity. Serum amyloid A assay is not routinely available but may be a valuable parameter for monitoring patients whose erythrocyte sedimentation rate and C-reactive protein level are normal (as in 2 of our patients with discitis).

Labial salivary gland biopsy is a reliable test for the diagnosis of primary and secondary amyloidosis. A prospective clinical and immunohistologic study in 59 patients.

OBJECTIVE: Reports of the detection of amyloidosis by labial salivary gland (LSG) biopsy have been mostly anecdotal. The aim of this study was to assess the value of this method in the diagnosis of amyloidosis. METHODS: LSG biopsy tissues were studied with a combination method using Congo red stain and immunohistologic characterization using an antibody directed against the serum amyloid P (SAP) component. Electron microscopy was performed in all cases. In a prospective study, we evaluated 30 patients with biopsy-proven AA or AL amyloidosis. We compared these patients with a control group of 29 age-matched patients without clinical or biologic evidence of amyloid disease (14 had rheumatoid arthritis and 15 had plasma cell dyscrasia). RESULTS: In 26 of the 30 patients with known systemic amyloidosis, amyloid deposits were identified on LSG biopsy (sensitivity of 86%). In 1 of the remaining patients, amyloid deposits were identified on LSG biopsy and systemic amyloidosis was confirmed by abdominal fat biopsy and 123I-labeled SAP scintigraphy. CONCLUSION: This study emphasizes the high sensitivity of LSG biopsy in the diagnosis of amyloidosis, even in the absence of oral symptoms.

Iodine-123-labelled serum amyloid P component scintigraphy in amyloidosis.

This study describes the results of scintigraphy with iodine-123-labelled serum amyloid P component (SAP) as a means of establishing the distribution of organ involvement in amyloidosis. The significance of 123I-SAP scans obtained in 15 patients with biopsy-proven AA or AL amyloidosis is discussed. Biopsy-proven amyloidosis was typically confirmed by scintigraphy, though such confirmation was not obtained in the kidneys in six patients with histological proof of extensive renal amyloid deposition. This lack of uptake may have been due to the accumulation of a major part of the 123I-SAP in the spleen and/or liver. Twenty-four hour whole-body retention of 123I-SAP was higher in patients with amyloidosis than in controls. Twenty-four hour tracer accumulation of the radioactivity in the extravascular compartment was notably greater in patients than in controls and appeared to be a good diagnostic criterion. We conclude that 123I-SAP scintigraphy may be helpful for the evaluation of organ involvement not only in patients with biopsy-proven amyloidosis but also when a biopsy cannot be performed or when a strong suspicion of amyloidosis exists in spite of repeated negative biopsies.

Quantification of beta 2-microglobulin and albumin in plasma and peritoneal dialysis fluid by a noncompetitive immunoenzymometric assay.

An enzyme-linked immunosorbent assay (ELISA) was developed for measuring beta 2-microglobulin (beta 2m) and albumin in continuous ambulatory peritoneal dialysis (CAPD) fluid. Plasma concentrations of beta 2m were twofold greater in hemodialysis patients (41.3 +/- 13.3 mg/L) than in CAPD patients (23.6 +/- 5.5 mg/L) matched for duration of treatment. Measurement of beta 2m in CAPD fluid showed a substantial loss of this protein, approximately 31% of total body beta 2m, compared with a 5% loss of a protein of middle molecular mass (albumin). Because of the molecular sieving effects of the peritoneal membrane, peritoneal clearance of beta 2m was sixfold greater than that of albumin. Whether beta 2m losses prevent or delay the incidence of dialysis-induced amyloidosis in these patients remains to be established.

Thrombin cleavage of apolipoprotein Bh of rabbit LDL: structural comparisons with human apolipoprotein B-100.

Rabbit plasma low density lipoprotein (LDL) contains one major apolipoprotein of apparent molecular weight of 320 kDa, designated apolipoprotein (apo) Bh, while another component termed apoB1 of apparent molecular weight of 220 kDa is found in chylomicrons. The fragments generated by thrombin digestion of the protein moieties of rabbit and human LDL were separated by polyacrylamide gradient gel electrophoresis and compared. As in the human species, the enzyme produced limited cleavage patterns of rabbit LDL apoB. Within the first 2 h, two fragments (Tr1 and Tr2, with apparent molecular weights 280,000 and 44,000, respectively) appeared. Longer incubations led to the production of two additional peptides, Tr3 and Tr4 (apparent molecular weights 180,000 and 96,000, respectively). Ten monoclonal antibodies, developed against rabbit LDL and designated P01 to P10, were found to react with rabbit apoB. Some also cross-reacted with human apoB. Epitope mapping, performed with these antibodies, showed that Tr3 and Tr4 were derived from the further degradation of Tr1. The rabbit is one of the most frequently used animals in atherosclerosis research. Its LDL receptor has been characterized and there exists a strain of homozygous LDL receptor-deficient rabbits referred to as WHHL rabbits. Despite this, little has been done to characterize the structure of rabbit apoB; only a short region has been sequenced and shown to be the carboxyl-terminal region, the rabbit apoB1. The molecular weight of human apoB (550,000) is much larger than rabbit apoBh. In both species, a primary and secondary thrombin cleavage occur, but the size of the fragments produced is very different between the two species. Identification of the thrombolytic fragments of the rabbit apoB have afforded the opportunity to compare the structures of both apoB species.

Quantification of serum amyloid P by enzyme-linked immunosorbent assay.

This enzyme-linked immunosorbent assay procedure for quantifying serum amyloid P (SAP) in human plasma makes use of affinity-purified polyclonal antibodies to SAP in a "sandwich"-type format. The procedure is sensitive, reproducible, simple, and easily automatable. Results correlate well with those by a rocket immunoelectrophoresis method performed with the same antibodies. Sera from apparently normal individuals had a mean SAP content of 44.17 mg/L and increased with age.

A sensitive immunochemiluminescence assay for human serum amyloid A protein.

We describe an immunochemiluminescence assay for human plasma serum amyloid A protein (SAA) in which specific rabbit polyclonal antibodies against synthetic peptides are used. The detection of the antigen-antibody reaction at 425 nm is based on a brief emission of light by a luminophor component (signal) in response to chemical energy. The working range of the assay covers plasma SAA concentrations from 5 to 100 micrograms/L. The lower detection limit is 5 micrograms/L, the within- and between-assay CVs are less than 12%. Bilirubin, cholesterol and triglyceride in final concentrations of up to 220 mumol/L, 8.1 mmol/L and 2.68 mmol/L, respectively, do not interfere with the assay. Results were correlated with those obtained by the enzyme-linked immunosorbent assay using the same antibodies (r = 0.95; p less than 0.001; n = 50). This method is inexpensive, simple and easily automated.

Serum amyloid A concentrations in giant-cell arteritis and polymyalgia rheumatica: a useful test in the management of the disease.

A prospective clinical study of 23 patients with giant-cell arteritis (GCA) and/or polymyalgia rheumatica (PMR) was undertaken in order to assess the behaviour of the non-specific markers of the disease activity, the erythrocyte sedimentation rate (ESR) and other acute phase markers, particularly the C-reactive protein (CPR) and serum amyloid A apolipoprotein (apo SAA) levels during induction of disease remission by prednisone therapy, and possible further recurrence of GCA and/or PMR. The apo SAA measurement is more sensitive than the CRP measurement in determining disease activity (97% and 61%, respectively). The specificity of apo SAA is greater than ESR in the determination of inactive disease (86% and 77%, respectively). In some cases with clinically active disease the ESR and CRP were normal, whereas the apo SAA was always elevated. We conclude that the apo SAA measurement in combination with clinical data and other laboratory parameters may be useful in the management of GCA and/or PMR.

Variations in apolipoproteins serum amyloid A, A-I, A-II, and C-III in severely head-injured patients.

In five severely head-injured patients we determined the plasma concentrations of apolipoproteins serum amyloid A, A-I, A-II, C-III, and B, prealbumin and C-reactive protein on day 1, 5, 10 and 15 after head injury where possible. A dramatic increase in apolipoprotein serum amyloid A up to a mean plasma level of 0.764 g/l was accompanied by a considerable decrease in apolipoprotein A-I, apolipoprotein A-II and apolipoprotein C-III concentrations. The variations observed by immunological methods were confirmed by two-dimensional gel electrophoresis performed on plasma and different lipoprotein fractions. In addition to its association with high density lipoproteins, apolipoprotein serum amyloid A was also found with lipoproteins of low and very low density. Two-dimensional electrophoresis also showed the presence of several different serum amyloid A-peptides not seen in plasmas from healthy subjects. We propose that apolipoprotein serum amyloid A may be responsible for the decrease of the main HDL apolipoproteins in head-injured patients.

[Serum amyloid A apolipoprotein (apo SAA). Implications in inflammation and in lipoprotein modifications]. / L'apolipoprotéine sérique amyloïde A (apo SAA). Implication dans l'inflammation et dans les modifications des lipoprotéines.

The measurement of serum amyloid A apolipoprotein (apo SAA) during acute phase inflammation offers a high interest because of its specificity, sensitivity and early increase of its levels, compared to other acute phase proteins. Furthermore apo SAA is transported in serum in association with lipoproteins, in particular with their denser subpopulation, HDL3 thus inducing their modification. The decrease in Lp AI:AII concentrations in inflammatory diseases is the consequence of the decrease in HDL3. In general the HDL3 composition was changed with a displacement of apo AI by SAA. Another interest to this protein is its relationship with amyloidosis. Apo SAA is the presumed precursor of amyloid A protein, which can be deposited in various tissues, leading to secondary amyloidosis.