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

Abstract

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 µg/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.

Introduction

The complement system is composed of a series of plasma proteins and plays a critical role in host defence and inflammation [1,2]. In addition to opsonization, other complement biological functions are mediated by peptides derived from the third (C3) and fifth (C5) components, the so-called anaphylatoxins C5a and C3a, and by the terminal membrane attack complex, C5b-9. Complement may be activated via the classical, the alternative or the MBL (mannan binding lectin) pathways. All of them lead to cleavage of C3 into C3a and C3b. C3b serves the assembly of further C3 and C5 molecules on the activating particle, to enable better clearance of pathogens and elimination of immune complexes. Factor I (C3b inactivator), is a serine protease that binds its target molecules C3b and C4b with the aid of co-factor molecules such as C4 binding protein (C4bp), factor H, complement receptor 1 (CR1, CD35) and membrane co-factor protein (MCP, CD46). C3b cleavage by factor I is considered a major regulatory step in limiting the potentially deleterious cytotoxic and proinflammatory activity of the complement system [3–5].

Factor I is composed of two disulphide-linked polypeptide chains with molecular weights of 50 000 and 38 000 Da. It is encoded by a gene on chromosome 4q, and synthesized as a single-chain precursor, which undergoes intracellular proteolytic processing [6]. One polypeptide contains a cysteine-rich domain similar to those found in the low-density lipoprotein receptor and complement components C6 and C7. The other polypeptide carries serine protease activity, which converts C3b and C4b to their inactive haemolytic forms, iC3b and iC4b, respectively. Factor I action requires the formation of bimolecular complexes of C3b and C4b with their respective co-factors factor H, C4b binding protein (C4bp) and membrane co-factor protein (MCP, CD46) or complement receptor 1 (CR1, CD35) [3–5].

Primary deficiencies of the complement system are rare disorders, comprising about 6% of immunodeficiencies [7–10]. Most of human complement deficiencies are due to autosomal co-dominant inheritance. Factor I deficiency causes a permanent, uncontrolled activation of the alternative pathway resulting in increased turnover of C3 and consumption of factor B, factor H and properdin. Persistent activation of C3 leads to deposition of C3b on bystander cells even in the absence of specific allo- or autoantibodies [11]. As a consequence, immune complex elimination may be impaired due to the inability of immune complex-fixed C3b to be converted to iC3b, a requirement for its subsequent binding to CR3 [12,13].

Patients with factor I deficiency present typically with systemic pyogenic infections, particularly with encapsulated bacteria such as Neisseria meningitidis, Haemophilus influenzae and/or Streptococcus pneumoniae. Heterozygous individuals for a complement factor deficiency usually present half-normal plasma levels of the respective component [11,14]. However, there is sparse information in the literature about the occurrence of infections in partial complement deficiencies, in particular, in patients with partial factor I deficiency.

The aim of this study was to extend our previous investigation on a Brazilian family with factor I deficiency and consanguinity [15] to the clinical and biochemical analysis of 39 individuals from three generations.

Patients and methods

Complement analysis was performed in three generations (39 members) of a Caucasian Brazilian family. Strong consanguinity among family members is reflected by the fact that the parents of the complete factor I-deficient patients are second-degree cousins, and all the grandparents are first-degree cousins (Fig. 1). The study was approved by the State University of Campinas Medical School Ethics Committee, according to the Helsinki Convention and to Brazil Ministry of Health, resolution 196/96.

Fig. 1.

Pedigree of a Brazilian family with factor I deficiency. ^aTwenty siblings: eight males, 12 females; ^b11 siblings: five males, six females.

A detailed medical history regarding infections and autoimmunity was obtained from the patients and family members. Recurrent respiratory infections were considered for more than two episodes of pneumonia per year above age 5 years, according to the findings of Barata et al. (1996) in a Brazilian community [16] and WHO criteria [17]. According to the Brazilian study, a new upper recurrent respiratory infection was only considered after 7 days of asymptomatic period.

Serum and ethylenediamine tetraacetic acid (EDTA)-plasma were obtained from family members, frozen immediately and stored at −80°C until analysis. Functional activity of the classical complement pathway (CH₅₀) was measured by haemolytic titration according to described procedures [18]. Plasma C3 concentrations were determined by nephelometry. C3dg/C3d was measured by double-decker rocket immunoelectrophoresis, using rabbit anti-C3c in the lower gel and anti-rabbit anti-C3d antibodies (Dako, Hamburg, Germany) in the upper gel [19].

Plasma concentrations of complement factors H and I were assessed by enzyme-linked immunosorbent assay (ELISA) using polyclonal goat antihuman factor H IgG or monoclonal anti-human factor I IgG (Genzyme, Boston), respectively, as capture antibodies. Bound regulator molecules were detected by rabbit anti-H IgG (ICN Biochemical, Eschwege, Germany), or goat anti-I IgG (ICN Biochemical, Eschwege, Germany), respectively. The reactions were visualized by the appropriate peroxidase-conjugated third antibodies, using 2,2′-azino-bis-3-ethyl-benzothiazoline sulphonate as substrate [15]. Optical density was measured at λ= 405 nm on an EAR 340 AT Microplate Reader (SLT, Calbiochem, Germany). Purified factors H and I (Quidel, San Diego, CA, USA) were taken as standards. The normal range for factor I-values was 44–25 µg/ml. This range was obtained by ELISA assay from 100 healthy blood donors and includes ± 2 s.d. Levels below 25 µg/ml were considered to define heterozygous individuals, and were different to those observed by Ferreira de Paula in the Brazilian population due to distinct methodology [20]. Specific anti-pneumococcal antibodies against serotypes 1, 5, 6B, and 8 were evaluated by ELISA [21].

Results

Tables 1, 2 and 3, respectively, summarize the complement profile, relationship and principal clinical manifestations of normal, heterozygous and homozygous individuals of this family. Among 39 members of the same family, we detected three patients with complete factor I deficiency in addition to 16 individuals with factor I levels below the normal range (Fig. 2). The parents of the affected patients presented low values of 20 and 22 µg/ml, respectively. Total complement activity (CH₅₀) and C3 levels were below normal values in complete factor I-deficient patients only. One heterozygous member presented a CH₅₀ below normal values. No C3dg/C3d was detected in complete factor I-deficient patients and the C3 cleavage product was low in an additional 14 individuals. Factor H was below normal values only in complete factor I-deficient patients.

Table 1.

Complement levels, relationship, and clinical profile of a factor I-deficient Brazilian family: family members with normal concentrations.

n	Sex	Age (years)	C3 0·55–1·1 mg/ml	Factor H 360–680 µg/ml	CH50 80–140 %	C3d 35–70 mU/l	Factor I 25–44 µg/ml	Clinical manifestations	Relationship
1	F	70	1·65	580	130	43	131	Healthy	Grandmother
3	F	32	1·57	606	136	36	95	Healthy	Aunt
4	F	39	1·58	580	132	38	75	Healthy	Aunt
5	F	28	1·40	580	128	40	45	Healthy	Aunt
8	F	42	1·27	554	116	38	45	Healthy	Aunt
12	M	44	1·27	554	106	38	55	Healthy	Uncle
13	M	25	1·43	1108	118	50	43	Healthy	Uncle
14	F	5	1·26	580	122	45	86	Chronic urticaria	Aunt
15	M	12	1·50	668	118	30	97	Sporadic tonsillitis and a single pneumonia	Cousin
16	F	15	1·16	636	106	28	40	Recurrent tonsillitis (average two episodes per year requiring antibiotics)	Cousin
17	M	10	1·22	636	112	33	48	Healthy	Cousin
18	M	6	1·36	816	127	43	71	Healthy	Cousin
20	F	14	1·20	380	105	36	52	Sporadic tonsillitis	Cousin
24	F	21	1·24	408	125	28	29	Healthy	Cousin
25	F	13	1·22	436	112	28	46	Sporadic tonsillitis	Cousin
26	F	27	1·03	380	105	33	45	Sporadic tonsillitis	Cousin
27	F	23	1·00	436	108	31	37	Sporadic tonsillitis and otitis	Cousin
28	M	18	1·22	408	101	31	40	Sporadic otitis and pneumonia (single episode)	Cousin
29	M	17	1·30	408	104	31	30	Healthy	Cousin
33	F	29	1·37	520	150	36	43	Recurrent pneumonias (10 episodes requiringhospitalization), chronic sinusitis and renal tuberculosis	Sister

Table 2.

Complement levels, relationship, and clinical profile of a factor I-deficient Brazilian family: partial deficient members.

n	Sex	Age (years)	C3 0·55–1·1 mg/ml	Factor H 360–680 µg/ml	CH50 80–140 %	C3d 35–70 mU/l	Factor I 25–44 µg/ml	Clinical manifestations	Relationship
2	M	73	1·45	580	125	30	23	Recurrent tonsillitis (average four episodes per yearrequiring antibiotics) and a single episode of pneumoniawith pleural effusion requiring hospital treatment	Grandfather
6	F	36	1·12	504	65	45	20	Healthy, minor infections	Aunt
7	F	37	1·60	668	143	50	20	Recurrent urinary infections including several episodesof pyelonephritis requiring hospitalization and furthernephrectomy. A single episode of meningitis (N. menigitidis) requiring hospitalization	Aunt
9	F	36	1·23	528	113	45	18	Recurrent urinary infections including several episodesof pyelonephritis requiring hospitalization and furthernephrectomy	Aunt
10	F	29	1·33	606	132	45	22	Recurrent urinary infections and viral respiratory infections	Aunt
11	M	21	0·95	668	91	43	19	Chronic arthritis	Uncle
19	F	14	1·23	464	102	38	17	Recurrent tonsillitis (average three episodes per yearrequiring antibiotics)	Cousin
21	M	9	1·28	436	110	48	13	Recurrent tonsillitis (average two episodes per yearrequiring antibiotics) and a single episode ofpneumonia requiring hospitalization	Cousin
22	F	20	1·00	380	101	36	14	Rheumatic fever	Cousin
23	M	17	0·99	408	101	28	19	Healthy, minor infections	Cousin
30	F	13	1·40	492	112	31	23	Recurrent otitis (average two episodes per year requiringantibiotics) and tonsillitis (average two episodes peryear requiring antibiotics)	Cousin
31	M	53	1·18	520	118	33	20	A single episode of pneumonia requiring hospitalization	Father
32	F	46	1·11	520	122	33	22	Recurrent otitis (average two episodes per year requiringantibiotics), tonsillitis (average three episodes per yearrequiring antibiotics), and recurrent pyodermitis. Asingle episode of pneumonia requiring hospitalization	Mother
34	F	27	1·16	552	113	41	18	Recurrent urinary infections (average three episodesper year requiring antibiotics), tonsillitis (average twoepisodes per year requiring antibiotics), and a stronglocal reaction to chickenpox immunization	Sister
35	M	26	1·31	612	112	39	24	Recurrent otitis (average two episodes per yearrequiring antibiotics)	Brother
39	M	8	1·05	472	108	39	22	Healthy, minor infections	Brother

Table 3.

Complement levels, relationship, and clinical profile of a factor I-deficient Brazilian family: deficient members.

n	Sex	Age (years)	C3 0·55–1·1 mg/ml	Factor H 360–680 µg/ml	CH50 80–140 %	C3d 35–70 mU/l	Factor I 25–44 µg/ml	Clinical manifestations	Relationship
36	M	18	0·27	245	43	0	0	Severe meningitis, sepsis and death	Brother
37	F	12	0·28	240	33	0	0·31	Recurrent tonsillitis (average six episodes per yearrequiring antibiotics), recurrent otitis (average twoepisodes per year requiring antibiotics), sinusitis (average two episodes per year requiring antibiotics), several subcutaneous infections requiring hospitalization, pulmonary tuberculosis and meningitis also requiringhospitalization	Sister
38	F	11	0·28	212	32	0	0·23	Recurrent tonsillitis (average two episodes per yearrequiring antibiotics), recurrent otitis (average twoepisodes per year requiring antibiotics), and diarrhoea (four episodes, two of them requiring hospitalization)	Sister

Fig. 2.

Distribution of normal, heterozygous and homozygous deficient factor I plasma levels (µg/ml) in a Brazilian family according to a normal range of 44–25 µg/ml.

Complete factor I-deficient patients presented recurrent respiratory infections, meningitis and skin infections (Table 3). One of them had pulmonary tuberculosis and the other died after sepsis, as outlined in detail in our previous report [15].

Partial factor I deficiency was assumed in 16 family members (Table 2). One patient presented with chronic arthritis and another with rheumatic fever. Ten individuals presented with more frequent infections including tonsillitis, pneumonia, urinary tract infections, otitis and meningitis. Two of those individuals were submitted to nephrectomy after recurrent urinary tract infections. Only two individuals with partial factor I deficiency did not present recurrent infections.

Normal factor I concentrations were detected in 20 family members; 11 of them were healthy (Table 1). The others presented only minor infections with the exception of patient 33, sister of the factor I-deficient patients, who presented recurrent pneumonia, chronic sinusitis and renal tuberculosis (Table 1).

Considering the occurrence of more frequent infections or probable autoimmunity (arthritis and rheumatic fever), we found, respectively, 12 of 16 and two of 20 clinically affected individuals in heterozygous and normal factor I members of the family (χ²; P < 0·0001). In our study, individuals with both homozygous and heterozygous factor I deficiency received a 23-valent pneumococcal vaccine and the levels of specific antibodies were still adequate after 3 years of vaccination (>1·3) (Table 4).

Table 4.

Anti-pneumococcal IgG levels (mg/l) in members of a factor I-deficient Brazilian family.

Member	Anti-PS1	Anti-PS3	Anti-PS5	Anti-PS6	Anti-PS9	Anti-PS14
31
″Pre	0·8	1·3	1·7	0·1	0·4	0·5
″Post	3·7	2·8	5·8	1·9	1·8	6·7
32
″Pre	3·5	2·5	2·5	1·4	0·3	0·2
″Post	8·9	6·9	8·9	7·9	5·9	1·4
33
″Pre	0·3	0·8	2·8	4·5	1·2	3·1
″Post	2·3	3·6	3·6	5·3	3·2	2·8
34
″Pre	3·3	2·1	1·1	2·3	0·2	2·4
″Post	9·2	9·3	5·4	5·5	2·7	5·8
35
″Pre	1·3	1·5	1·9	2·4	3·2	2·7
″Post	4·5	6·4	2·4	2·6	2·3	9·4
37
″Pre	2·4	2·0	1·2	3·5	9·3	3·2
″Post	8·0	9·5	6·8	10·7	14·5	4·1
38
″Pre	5·6	9·0	4·8	8·7	5·2	9·7
″Post	15·9	18·8	15·8	12·9	15·9	21·2
39
″Pre	0·4	1·4	1·2	3·2	1·8	1·1
″Post	1·8	3·8	5·7	9·7	5·4	5·2

Discussion

Factor I is a proteolytic enzyme that regulates the multiple biological activities of C3b. Deficiency of this regulator causes a permanent activation of the alternative complement pathway leading to uncontrolled formation of the fluid phase C3 convertase (C3bBb) with subsequent depletion of C3 and Factor B. In consequence, C3 is converted to C3b without further degradation to iC3b and C3dg/d [11,22–25].

Alper et al. [26] showed that a patient with (type I essential) hypercatabolism of C3 was homozygous for an inherited deficiency of factor I. Amadei et al. [27] recently described two sisters with complement factor I deficiency from one family, bringing the total number of cases reported to 37 individuals of 27 families [3,11–15,27–41]. Using factor I cDNA clones in Southern analysis of somatic cell hybrids, Goldberger [6] mapped the factor I gene to chromosome 4. Shiang et al. [42], used portions of the long arm of chromosome 4 and mapped factor I to 4q23–q25. The molecular basis of factor I deficiency was determined in few cases [41,43], revealing distinct mutation sites.

Complete factor I deficiency is associated usually with increased susceptibility to bacterial infections. As in primary C3 deficiency, some individuals lacking factor I show clinical similarities to those with agammaglobulinaemia [44]. Respiratory infections, meningitis and arthritis are usual clinical consequences of factor I deficiency [14,45,46]. Factor I deficiency has also been associated with a reduced generation of complement-derived chemotactic factors. Important consequences are impaired recruitment of phagocytic cells to the site of infection and subsequent reduced inflammatory response [41].

In our study, the strong consanguinity is the major factor for the distribution of this primary immunodeficiency. Factor I plasma levels below 25 µg/ml established in our laboratory by ELISA assay led to the assumption that 16 family members are heterozygous for factor I deficiency. In contrast to the findings of complete factor I deficiency, partial factor I deficiency was reported hitherto not to be associated with clinical consequences. In the present family we observed frequent episodes of infections, although the functional capacity of the complement system (as judged from its haemolytic activity) appeared not to be diminished significantly, as reported previously [29,32].

Meningitis caused by N. meningitidis was reported in a total of 14 patients with factor I deficiency [45] in addition to the two complete factor I-deficient patients of this study. Infection caused by N. meningitidis has been observed in 50% of factor I-deficient patients, in 30% of factor H-deficient patients and in 18% of C3-deficient patients [14].

We also observed the occurrence of meningitis in one partial factor I-deficient individual. Bonnin et al. [40] reported a factor I-deficient patient with recurrent aseptic Mollaret meningitis, which is self-limited, causes no neurological damage and has an unknown aetiology. Although renal involvement has been observed in factor H (haemolytic uraemic syndrome [47] and C3-deficient patients [48], pyelonephritis was reported previously only in factor I deficiency by Reutter-Simon et al. 1994 (Reutter-Simon, personal communication). Sadallah et al. [49] reported the loss of renal function in one factor I-deficient patient, caused clearly by progressive glomerulonephritis. Two of the partial factor I-deficient relatives described here were submitted to nephrectomy after recurrent urinary tract infections at the age of 36 and 40 years, respectively. The prognosis of pyelonephritis is related closely to the number of risk factors [50,51], e.g. occurrence of urinary tract infections. In pyelonephritis, partial factor I deficiency could be a triggering factor for bad prognosis.

It is interesting that case 33, the sister of the complete factor I-deficient patients, presented normal factor I levels but also suffered from recurrent pneumonia, chronic sinusitis and renal tuberculosis. Considering the strong consanguinity of this family, other unrecognized immunological defects need to be considered in this particular case.

In the homozygous-deficient patients of this study, infections started early in life. Although in complete factor I-deficient patients the initial clinical manifestations often occur in childhood at a mean age of 2 years, some patients present symptoms later in life [33,40], due probably to other compensatory mechanisms [14].

Factor I deficiency partially impairs the clearance of immune complexes by phagocytes, increasing the risk for the development of immune complex-mediated diseases [12,32]. Teisner et al. [32] observed the presence of circulating immune complexes in patients with factor I deficiency, but without clinical evidence for autoimmune diseases. In our study, two partial factor I-deficient relatives presented diseases with probable autoimmune mechanism: chronic arthritis and rheumatic fever. Low C3 levels may lead to decreased solubility of autoantibodies and to the development of immune complex disease. As a consequence, vasculitis and other related abnormalities may occur, as described for our previous reported C3-deficient patient [48].

Wahn et al. [30] and later Rasmussen et al. [33] reported low levels of factor H in factor I-deficient patients to be associated with an altered electrophoretic mobility of the regulator from alpha to beta pattern. In our previous study, we could demonstrate that a tight complex of factor H with C3b, normally dissolved by the action of factor I leads to a change in factor H electrophoretic mobility [15]. This finding was confirmed recently by Naked et al. [41]. A high number of relatives in that family had factor H-values around 50% of normal.

Impairment of antibody response has not been described in factor I-deficient patients, except for impaired antibody response to thymus-dependent antigen [52]. Although impaired antibody responses to polysaccharides have not been definitely documented in complement-deficient patients, it is likely to occur because conjugates of pneumococcal polysaccharide with C3d have been shown to promote antibody responses [53]. Reduction of IgG subclasses in C3-deficient patients has also been documented previously [54]. Therefore, the immunization of factor I-deficient patients for N. meningitidis, S. pneumoniae and H. influenzae has been recommended. Indeed, our patients with complete or partial factor I deficiency showed a normal response to pneumococcal vaccine. Drogari-Apiranthitou et al. [55] reported a good antibody response to meningococcal vaccine in patients with late complement (C5–C8) components as well as in those with C3 deficiency. Our complete and partial factor I-deficient patients were all immunized, except one of the homozygous patients who died after fatal meningococcal meningitis. All presented a good response. In addition to immunizations, our factor I-deficient patients receive penicillin on a regular basis, which proved an adequate control of infections.

Patients with factor I deficiency have been commonly treated with antibiotics during infectious episodes. If the infections are severe, repetitive infusion of fresh plasma may be of benefit as suggested previously [30,31,34,56]. However, the risk of reactions and/or the transmission of infectious agents have to be taken into consideration.

Although factor I deficiency is a rare disease its identification appears to be of importance, as even partial deficiencies may increase the risk for severe infections and autoimmunity. We conclude that patients with factor I deficiency are at high risk for recurrent infections and should receive vaccination against capsulated bacteria and prophylactic antibiotics on a regular basis. Apparently, patients with partial factor I deficiency are also at higher risk for recurrent infections. Although few infections could be associated with encapsulated bacteria in our study, we propose that available vaccination against encapsulated bacteria and the eventual use of prophylactic antibiotics should be considered individually for patients in the heterozygous group.