Genetic,molecular and functional analyses of complement factor I deficiency

Complete deficiency of complement inhibitor factor I (FI) results in secondary complement deficiency due to uncontrolled spontaneous alternative pathway activation leading to susceptibility to infections. Current genetic examination of two patients with near complete FI deficiency and three patients with no detectable serum FI and also close family members revealed homozygous or compound heterozygous mutations in several domains of FI. These mutations were introduced into recombinant FI and the resulting proteins were purified for functional studies, while transient transfection was used to analyze expression and secretion. The G170V mutation resulted in a protein that was not expressed, whereas the mutations Q232K, C237Y, S250L, I339M and H400L affected secretion. Furthermore, the C237Y and the S250L mutants did not degrade C4b and C3b as efficiently as the WT. The truncated Q336x mutant could be expressed, in vitro, but was not functional because it lacks the serine protease domain. Furthermore, this truncated FI was not detected in serum of the patient. Structural investigations using molecular modeling were performed to predict the potential impact the mutations have on FI structure. This is the first study that investigates, at the functional level, the consequences of molecular defects identified in patients with full FI deficiency.     

Deficiency of human complement factor I associated with lowered factor H

Deficiencies of factor I and/or factor H result in an increased consumption of C3 and higher susceptibility to recurrent infections. Here we describe a case of human factor I deficiency and lowered factor H levels. C3 concentration was 50% lower than normal, the classical pathway-dependent hemolytic activity was reduced to almost 30% of normal, and alternative pathway-dependent activity was completely absent. The killing by peripheral leukocytes of Candida albicans treated with deficient serum and the production of complement-dependent chemotactic factors were reduced in the proband's serum when compared with normal serum. Finally, we observed that C3 antigen present in the proband's serum has a different electrophoretic mobility than native C3 (most likely C3b), confirming the deregulation of complement activation due to the lack of regulatory proteins factors I and H. The impaired complement system described in this case, the first of its kind described in a Chile, explains the higher susceptibility to infections found in the proband.     

Pharmacological modulation of platelet-activating factor (PAF) release from rabbit leucocytes. I. Role of cAMP

Basophil-rich rabbit leucocytes sensitized by anti-horseradish peroxidase antibodies released platelet-activating factor (PAF) and histamine upon exposure to the specific antigen. This release was preceded and accompanied by a sharp decrease in the intracellular concentration of cyclic AMP. Isoproterenol, a beta-adrenergic agent, and theophylline, a phosphodiesterase inhibitor, used individually or in combination, increased the intracellular concentration of cyclic AMP and inhibited the release of both PAF and histamine. Propranolol, a beta-adrenergic blocking agent, suppressed the effect of isoproterenol on cyclic AMP level and mediator release. Dibutyryl cyclic AMP, an alkylated derivative of cyclic AMP, inhibited PAF and histamine release. These results indicate that cyclic AMP, which is known to control the release of other mediators of immediate hypersensitivity, also regulates the release of PAF. Histamine and PAF followed one another closely in all of our release or inhibition experiments, bringing more evidence for the basophil origin of PAF.     

Molecular heterogeneity in deficiency of complement protein C2 type I.

Deficiency of the complement protein C2 (C2D), one of the most common genetic deficiencies of the complement system, is associated with rheumatological disorders and increased susceptibility to infection. Two types of C2D have been recognized, each in the context of specific major histocompatibility complex (MHC) haplotypes; type I, a deletion, frameshift and premature stop codon resulting in absence of detectable C2 protein synthesis, and type II, missense mutations resulting in a block in secretion of C2 proteins. Analysis of C2 expression in a child with C2 deficiency, a MHC haplotype different from those associated with type I or II C2D, and recurrent infections revealed additional molecular heterogeneity among C2 deficient patients. No detectable C2 protein was synthesized in the child's fibroblasts under conditions supporting C2 synthesis and secretion in normals and the child's C2 mRNA was reduced to 42% of normal. Nucleotide sequencing of RT-PCR fibroblast mRNA and genomic DNA revealed a type I C2 deficiency (28 base-pair deletion) on one allele and a previously unrecognized two base-pair deletion in exon 2 on the other. Expression of the closely linked factor B gene was markedly decreased (Bf mRNA 25% of normal), though Bf was up-regulated appropriately by interferon-gamma and the flanking sequence containing the Bf promoter was normal in this C2-deficient patient. Moreover, the concentration of Bf protein was normal in the patient's plasma.     

Primary pneumococcal peritonitis can be the first presentation of a familial complement factor I deficiency1

Primary pneumococcal peritonitis is a rare infection that has been described in women but has not been previously linked with immunodeficiency. The complement system plays a central role in immune defence against Streptococcus pneumoniae and, in order to evade complement attack, pneumococci have evolved a large number of mechanisms that limit complement-mediated opsonization and subsequent phagocytosis. We investigated an apparently immunocompetent woman with primary pneumococcal peritonitis and identified a family with deficiency for complement factor I. Primary pneumococcal peritonitis should be considered a possible primary immunodeficiency presentation.     

Recurrent infections in partial complement factor I deficiency: evaluation of three generations of a Brazilian family

We report here on the evaluation of a factor I-deficient Brazilian family (three generations, 39 members) with strong consanguinity. The complete factor I-deficient patients (n = 3) presented recurrent respiratory infections, skin infections and meningitis; one of them died after sepsis. They presented an impaired total haemolytic activity (CH50), low C3, low factor H and undetectable C3dg/C3d. Partial factor I deficiency was detected in 16 family members (normal low cut-off value was 25 microg/ml). Respiratory infections were the most common clinical occurrence among partial factor I-deficient relatives. Two of them were submitted to nephrectomy following recurrent urinary tract infections. An additional two heterozygous relatives presented with arthritis and rheumatic fever. Apparently, patients with partial factor I deficiency are also at higher risk for recurrent infections. Vaccination against capsulated bacteria and the eventual use of prophylactic antibiotics should be considered individually in this patient group.     

Familial deficiency of complement factor 7: association with bacterial meningitis. Apropos of 3 recent cases

Three recent cases of inherited deficiency of the seventh component of complement (C7) associated with recurrent infectious meningitis are described. Two cases were associated with meningococcal meningitis, the third is the first case report of C7 inherited deficiency associated with Haemophilus parainfluenzae meningitis. Family studies are consistent with inheritance and non-HLA-linked, autosomal codominant trait of the C7 deficiency. The three patients have remained well, following antibiotic treatment.     

Functional deficiency of complement factor D in a monozygous twin

An adult twin with recurrent bacterial infections was found to have a partial functional deficiency of complement factor D. Full restoration of alternative pathway activity and zymosan- or cobra venom factor-induced consumption of C3 and B was found after reconstitution of patient's serum with purified D. Family studies revealed normal D levels in the mother, a brother and another sister. After gel filtration of patient's sera only little D activity could be detected in the fractions, and trypsin activation of the fractions also did not uncover detectable precursor D activity.     

Decreased immunoreactive beta-endorphin in mononuclear leucocytes from patients with rheumatic diseases.

The neuroendocrine polypeptide hormone beta-endorphin (beta-END), which is released from various tissues including the anterior pituitary gland and cells of the immune system, has recently been implicated as having an immunoregulatory role. We used a radioimmunoassay to measure beta-END levels in circulating mononuclear leucocytes from normal subjects and patients with various rheumatic diseases. Levels of beta-END in leucocytes from patients were lower than in leucocytes from healthy subjects (P less than 0.001). Whereas levels of beta-END in leucocytes from patients with the various rheumatic disorders were not significantly different, an inverse correlation was found between beta-END levels in leucocytes and expression of rheumatoid factor (P less than 0.025) and erythrocyte sedimentation rate (P less than 0.025). This study demonstrates decreased content of beta-END in cells of the immune system related to parameters of inflammatory activity in rheumatic diseases.     

In vitro biosynthesis of complement factor I by human endothelial cells.

We have studied the secretion of the complement regulatory protein factor I by human umbilical vein endothelial cells (HUVEC). Northern and Western blot analysis and biosynthetic labeling experiments indicate that HUVEC secrete factor I at very low levels in basal conditions and that this secretion is significantly enhanced by interferon-gamma. Analysis of the proteolytic inactivation of C3b by HUVEC supernatants show that factor I is secreted in a functional form and can promote the specific proteolytic inactivation of C3b to iC3b. Together with previous studies establishing the secretion of complement factor H by HUVEC, this work demonstrates that the endothelial cell is able to secrete in its environment two complement regulatory proteins, factor I and factor H, which can mediate the degradation of C3b to iC3b. The secretion of factor I by HUVEC provides a useful in vitro model to analyze the modulation of this secretion and may be relevant to the local deposition of iC3b at the surface of the endothelium during the inflammatory reaction.     

Three cases of factor I deficiency: the effect of treatment with plasma.

Three patients with congenital factor I deficiency associated with different clinical manifestations are described. Case 1 had one single episode of meningococcal disease, case 2 experienced four episodes of meningococcal disease and several other severe infections, whereas case 3, without known predisposition for infections, died from a subacute immune-complex mediated syndrome, resembling polyarteritis nodosa. Family studies in cases 1 and 2 revealed healthy individuals with factor I concentrations below the lower reference limit, indicating heterozygous carriers. The pedigree analyses were consistent with autosomal codominant inheritance. The estimated minimal frequency of the deficient gene was 0.002. Pedigree analysis was not performed in case 3 but the father and sister was found to be probable heterozygous carriers. Cases 2 and 3 were treated with infusions of freshly frozen plasma (FFP) (40 and 27 ml/kg bodyweight) during acute illness and the immunochemical complement profile was monitored. Following plasma infusion factor I was cleared from the circulation with a half-life of 29-45 h. The plasma infusions induced generation of C3d and C4d, increase in native factor B and C3 concentrations and disappearance of Ba split products. Native C3 and C4 increased to normal concentrations and remained normal till 16 days after the plasma infusions, whereas native factor B decreased to preinfusion levels 8 days after plasma infusion. It is concluded, that congenital factor I deficiency can present with different clinical manifestations and may be more prevalent than hitherto anticipated. Furthermore, infusion of blood products containing small amounts of functional factor I can partly normalize the complement profile, with a more prolonged effect on C3 and C4 than on factor B metabolism.     

Variation near complement factor I is associated with risk of advanced AMD

A case-control association study for advanced age-related macular degeneration was conducted to explore several regions of interest identified by linkage. This analysis identified a single nucleotide polymorphism just 3' of complement factor I on chromosome 4 showing significant association (P<10(-7)). Sequencing was performed on coding exons in linkage disequilibrium with the detected association. No obvious functional variation was discovered that could be the proximate cause of the association, suggesting a noncoding regulatory mechanism.     

Studies of the third component of complement in synovial fluid from arthritic patients. I. Immunochemical quantitation and relation to total complement

In a study of forty-one patients, most of whom had longstanding rheumatoid arthritis, a comparison was made in serum and synovial fluid between total complement activity (CH₅₀) and C3, measured immunochemically. Nineteen out of twenty-seven patients with rheumatoid arthritis and four out of four cases of systemic lupus erythematosus (SLE) or SLE-like syndromes had a depressed CH₅₀ value in the synovial fluid, while ten cases with a non-rheumatoid form of arthritis all had normal activity.

The total C activity in serum was normal or increased in most cases. While no correlation was found between CH₅₀ and C3 in serum, there was a statistically highly significant correlation in synovial fluid, indicating that C3 determinations can be used to reveal alterations in C activity of synovial fluid.

     

Growth factor deficiency in patients with ataxia telangiectasia

One prominent feature of patients with the autosomal recessive disease ataxia telangiectasia (AT) is somatic growth retardation. Due to their essential roles in development we examined levels of insulin-like growth factor-I (IGF-I) as well as its main binding protein (IGFBP-3) in a group of AT patients. Growth status of 19 patients was assessed by body mass index (BMI) and nutritional protocols. As suspected, BMI was low in AT patients despite adequate nutrition. Serum levels of IGF-I were found to be below the 3rd percentile in 9 (56%) out of 16 patients and of IGFBP-3 in 13 (81%) out of 16 patients. Our observations demonstrate that IGF-I and IGFBP-3 levels reflect the impaired growth status in patients with AT.     

A new family with congenital factor XIII deficiency showing a deficit of both subunit A and B. Type I factor XIII deficiency

In this study we present a new case of Factor XIII deficiency. The proposita, a 34 year old woman, showed a deficiency of both subunit a and subunit b, and a moderate bleeding tendency. Because of the concomitant decrease of subunits a and b the proposita is considered to be an example of Type I disease. Factor XIII levels were less than 10% both as activity and antigen. Several family members showed intermediate levels of both subunit a and b and were asymptomatic. They were considered to be heterozygotes. The hereditary pattern is autosomal incompletely recessive. Type I disease appears much less frequent than Type II.