Homozygous C3 Research Articles

672 - Complete response to eculizumab in atypical hemolytic uremic syndrome with homozygous C3 mutation.

672 - Complete response to eculizumab in atypical hemolytic uremic syndrome with homozygous C3 mutation.

A Single Nucleotide C3 Polymorphism Associates With Clinical Outcome After Lung Transplantation.

Background: Development of chronic rejection is still a severe problem and causes high mortality rates after lung transplantation (LTx). Complement activation is important in the development of acute rejection (AR) and bronchiolitis obliterans syndrome, with C3 as a key complement factor.Methods: We investigated a single nucleotide polymorphism (SNP) in the C3 gene (rs2230199) in relation to long-term outcome after LTx in 144 patient-donor pairs. In addition, we looked at local production of donor C3 by analyzing bronchoalveolar lavage fluid (BALF) of 6 LTx patients using isoelectric focusing (IEF).Results: We demonstrated the presence of C3 in BALF and showed that this is produced by the donor lung based on the genotype of SNP rs2230199. We also analyzed donor and patient SNP configurations and observed a significant association between the SNP configuration in patients and episodes of AR during 4-years follow-up. Survival analysis showed a lower AR-free survival in homozygous C3 slow patients (p = 0.005). Furthermore, we found a significant association between the SNP configuration in donors and BOS development. Patients receiving a graft from a donor with at least one C3 fast variant for rs2230199 had an inferior BOS-free survival (p = 0.044).Conclusions: In conclusion, our data indicate local C3 production by donor lung cells. In addition, a single C3 SNP present in recipients affects short-term outcome after LTx, while this SNP in donors has an opposite effect on long-term outcome after LTx. These results could contribute to an improved risk stratification after transplantation.

Hyperfunctional complement C3 promotes C5-dependent atypical hemolytic uremic syndrome in mice

Atypical hemolytic uremic syndrome (aHUS) is frequently associated in humans with loss-of-function mutations in complement-regulating proteins or gain-of-function mutations in complement-activating proteins. Thus, aHUS provides an archetypal complement-mediated disease with which to model new therapeutic strategies and treatments. Herein, we show that, when transferred to mice, an aHUS-associated gain-of-function change (D1115N) to the complement-activation protein C3 results in aHUS. Homozygous C3 p.D1115N (C3KI) mice developed spontaneous chronic thrombotic microangiopathy together with hematuria, thrombocytopenia, elevated creatinine, and evidence of hemolysis. Mice with active disease had reduced plasma C3 with C3 fragment and C9 deposition within the kidney. Therapeutic blockade or genetic deletion of C5, a protein downstream of C3 in the complement cascade, protected homozygous C3KI mice from thrombotic microangiopathy and aHUS. Thus, our data provide in vivo modeling evidence that gain-of-function changes in complement C3 drive aHUS. They also show that long-term C5 deficiency is not accompanied by development of other renal complications (such as C3 glomerulopathy) despite sustained dysregulation of C3. Our results suggest that this preclinical model will allow testing of novel complement inhibitors with the aim of developing precisely targeted therapeutics that could have application in many complement-mediated diseases.

Complement C3 levels in chronic hepatitis C patients from India

Background and Aims: Complement is a collective term applied to a group of serum proteins which participate in antigen antibody reactions leading to irreversible cell damage and eventually lysis. The serum concentration of the third component of complement, C3, closely reflects the total complement activity. Individuals affected by homozygous C3 deficiency suffer from recurrent pyogenic infections such as pneumonia, septicemia, otitis media, and meningitis, and the absence of C3 is frequently lethal. This study aims to find out if there is any relation between chronic hepatitis C infection and levels of C3 in serum. Methods: The study included 132 chronic hepatitis C patients diagnosed as per American Association for Study of Liver Disease (AASLD) practice guidelines. Control groups comprised of 81 healthy blood donors who did not have any history of liver disorders. C3 levels in serum were determined utilizing commercially available kit. Results: Out of 132 chronic hepatitis C patients included, 81 were males and 51 were females with mean age± S.D. of 35.11±11.40. C3 level of the healthy group was 88.5±25.3 mg/dl whereas the C3 level of the chronic hepatitis C group was 56±18 mg/dl and was statistically significant (P <0.0005) indicating that in HCV infection there is lowering of complement system C3 components. Conclusion: The significant difference between the chronic hepatitis C patient and healthy control C3 level indicates need for including it as one of the factors that should be monitored during chronic hepatitis C infection and treatment.

Consumption of high ω-3 fatty acid diet suppressed prostate tumorigenesis in C3(1) Tag mice

Prostate cancer incidence and mortality are high in the Western world and high ω-6/ω-3 PUFA in the Western diet may be a contributing factor. We investigated whether changing from a diet that approximates ω-6 fat content of the Western diet to a high ω-3 fat diet at adulthood might reduce prostate cancer risk. Female SV 129 mice that had consumed a high ω-6 diet containing corn oil for 2 weeks were bred with homozygous C3(1)Tag transgenic male mice. All male offspring were weaned to the corn oil diet (CO) until postpuberty when half of the male offspring were transferred to a high ω-3 diet containing canola oil and fish oil concentrate (FS). High ω-3 diet increased ω-3 and decreased ω-6 fat content of mice tissues. Average weights of prostate and genitourinary bloc were significantly lower in mice consuming high ω-3 diet at adulthood (CO-FS) than mice fed a lifetime high ω-6 diet (CO–CO). There was slower progression of tumorigenesis in dorsalateral prostate of CO-FS than in CO–CO mice. CO-FS mice had slightly lower plasma testosterone level at 24 and 40 weeks, significantly lower estradiol level at 40 weeks and significantly less expressed androgen receptor (AR) in the dorsalateral prostate at 40 weeks than CO–CO mice. Consumption of high ω-3 diet lowered the expression of genes expected to increase proliferation and decrease apoptosis in dorsalateral prostate. Our results suggest that consumption of high ω-3 diet slows down prostate tumorigenesis by lowering estradiol, testosterone and AR levels, promoting apoptosis and suppressing cell proliferation in C3(1)Tag mice.

Abstract 1816: Effects of high ω-3 fatty acid diet on prostate tumorigenesis in C3(1) Tag mice

Abstract Epidemiological studies have shown that intake of diet high in omega 6 polyunsaturated fatty acid (ω-6 PUFA) increases the risk of prostate cancer while the diet high in omega-3 polyunsaturated fatty acid (ω-3 PUFA) decreases the risk. Omega-6 PUFA are abundant in corn oil and sunflower oil while omega-3 PUFA are abundant in fish oil, canola oil and walnut. Both ω-6 and ω-3 play an important role in prostate cancer risk. Prostate cancer incidence and mortality are high in the Western world and high ω-6/ ω-3 PUFA in the Western diet may be a contributing factor. The aim of this study was to investigate whether a change from a diet that approximates the ω-6 fatty acid content of the Western diet to a high ω-3 fatty acid diet at adulthood might reduce prostate cancer risk. Female SV 129 mice that had consumed a high ω-6 diet containing 10% w/w corn oil (containing 50% ω-6 and 1% ω-3) for two weeks were bred with homozygous C3(1) Tag transgenic male mice. All male offspring (hemizygous) were weaned to the corn oil diet and consumed this diet until post puberty (7 weeks). Since the aim of the study was to simulate a diet change at adulthood, half of the male offspring were transferred to a high ω-3 diet containing 5%w/w canola oil and 5% w/w fish oil concentrate (containing 1%w/w ω-6 and 35%w/w ω-3). The male hemizygous mice were euthanized at age 24wks and 40wks (end of reproduction). Gas chromatography was used to assess the fatty acid composition in the prostate, liver and fat tissues. Enzyme-linked immunoassay was used to assess the levels of testosterone and estradiol in blood plasma. RT2 PCR and WB analyses were used to assess mRNA gene and protein expression respectively in the dorsalateral (DL) prostate. The results demonstrated that high ω-3 diet increased the ALA, EPA and DHA and decreased the LA and AA content of the prostate, liver and fat tissues. The average weights of prostate and genitourinary bloc (GU block) at 24 wks and 40 wks were significantly lower in mice that consumed the high ω-3 diet at adulthood (CO-FS) than in mice that consumed the high ω-6 diet (CO-CO) throughout life. Compared to mice in CO-CO group, CO-FS mice had lower plasma testosterone level at 24wks and 40wks, and significantly lower estradiol level at 40wks. Gene expression analyses showed that consumption of high ω-3 diet altered expression of genes that would be expected to slow proliferation and increase apoptosis in the DL prostate. Our findings suggest that if adult consume high ω-3 (but low in ω-6) diet, it may be useful in the suppression and prevention of prostate cancer. Further ongoing investigation include pathological evaluation of DL prostate from the two diet groups and immunohistochemical analyses to assess the expression of AR, ER and proliferation in the mice DL prostate. TUNEL assay will be used to assess apoptosis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1816. doi:10.1158/1538-7445.AM2011-1816

Maternal consumption of canola oil suppressed mammary gland tumorigenesis in C3(1) TAg mice offspring

BackgroundMaternal consumption of a diet high in omega 6 polyunsaturated fats (n-6 PUFA) has been shown to increase risk whereas a diet high in omega 3 polyunsaturated fats (n-3 PUFA) from fish oil has been shown to decrease risk for mammary gland cancer in female offspring of rats. The aim of this study was to determine whether increasing n-3 PUFA and reducing n-6 PUFA by using canola oil instead of corn oil in the maternal diet might reduce the risk for breast cancer in female offspring.MethodsFemale SV 129 mice were divided into two groups and placed on diets containing either 10% w/w corn oil (which is 50% n-6 PUFA, control diet) or 10% w/w canola oil (which is 20% n-6 PUFA, 10% n-3 PUFA, test diet). After two weeks on the diets the females were bred with homozygous C3(1) TAg transgenic mice. Mother mice consumed the assigned diet throughout gestation and nursing of the offspring. After weaning, all female offspring were maintained on the control diet.ResultsCompared to offspring of mothers fed the corn oil diet (CO/CO group), offspring of mothers fed the canola oil diet (CA/CO group) had significantly fewer mammary glands with tumors throughout the experiment. At 130 days of age, the CA/CO group had significantly fewer tumors per mouse (multiplicity); the tumor incidence (fraction of mice with any tumor) and the total tumor weight (per mouse that developed tumor) was less than one half that of the CO/CO group. At 170 days of age, the total tumor weight per mouse was significantly less in the CA/CO group and if a tumor developed the rate of tumor growth rate was half that of CO/CO group. These results indicate that maternal consumption of canola oil was associated with delayed appearance of mammary gland tumors and slowed growth of the tumors that developed.ConclusionsSubstituting canola oil for corn oil is an easy dietary change for people to make; such a change to the maternal diet may decrease risk for breast cancer in the daughter.

An association between homozygous C3 deficiency and low levels of anti-pneumococcal capsular polysaccharide antibodies.

Inherited deficiencies of complement components are associated with an increased risk of infection by encapsulated, high grade bacterial pathogens such as Streptococcus pneumoniae, Haemophilus influenzae type b and Neisseria meningitidis. Hence, the levels of antibodies to bacterial capsular polysaccharide antigens were measured using ELISA in 65 patients with inherited deficiencies covering the classical, alternative and terminal components of the complement cascade. Three of the four C3-deficient individuals studied were found to be almost totally deficient in specific anti-pneumococcal capsular polysaccharide (PCP) antibodies. These individuals had a history of recurrent pneumococcal sepsis. While single individuals with C1r, C2 and C1Inh deficiency were found to have low anti-PCP antibody levels, no other group of complement deficiency had significantly reduced anti-PCP antibody levels compared with 100 controls. Antibody levels to the other two polysaccharides were not significantly lower in the patient groups. These findings suggest that C3 may be able to provide a stimulatory signal to promote the production of anti-PCP antibodies.

Homozygous C3 Deficiency Associated with IgA Nephropathy

A 23-year-old male patient with homozygous C3 deficiency who developed asymptomatic proteinuria and hematuria was reported. Renal biopsy disclosed typical IgA nephropathy with deposition of early- and late-complement components except for C3 deposition. C9 and membrane attack complex were detected in the glomeruli despite the absence of C3. It was suggested that there might be some unknown complement activation mechanism which does not require C3 component.

Structure and expression of the C3 gene.

To map the multiple interactive sites on the C3 polypeptide, it is advantageous to combine the approaches of protein chemistry, nucleic acid technology, and molecular biology. This review summarizes the currently known molecular properties of mouse liver C3 mRNA, cloned C3 cDNA, and genomic DNA. Original data communicated have specified the amino acid sequence of the 215 amino-terminal residues of mature mouse C3 beta. Southern blot analysis of liver DNA indicated that the mouse genome contains only one type of C3 gene, that murine and human C3 sequences strongly cross-hybridize, and that the human C3 gene is not somatically rearranged. Included are descriptions of the first human C3 genomic DNA clones, their preparation, and their use to map the human C3 gene to chromosome 19 in linkage with the myotonic dystrophy (DM) locus. After a brief survey of reports describing inherited human C3 deficiencies, we discuss a Dutch family and their three members with total homozygous C3 deficiency who were the subjects of a recent publication. The restricted synthesis of C3 in major and minor producer tissues is discussed and it is proposed that the C3 gene provides a good model system for studying the molecular basis of tissue-specific gene expression. Data are presented documenting the production of C3 in two established mouse macrophage-like cell lines and two rat hepatoma cell lines in tissue cultures. A short account covers the extensive literature on regulation of C3 serum concentrations in acute and chronic inflammation and the very incomplete picture that presently depicts hormonal regulation of C3 synthesis. The final experiment reported demonstrates that nucleic acid hybridization with cloned cDNA probes is a sensitive assay for quantitative determinations of C3 mRNA. With the help of cloned cDNA and genomic DNA, researchers can address questions concerning the functional topography of the C3 polypeptide, the gene's structure, and the molecular nature of inherited C3 deficiencies in humans.

Complete absence of the third component of complement in a patient with repeated infections

A 10-year-old girl with complete absence of the third component of complement (C3) and repeated infections is reported. No serum C3 could be detected by either immunochemical or hemolytic assays. Fourteen out of the twenty-seven relatives, including both parents, had half-normal immunochemical and hemolytic C3 levels, suggesting the inheritance pattern of autosomal codominance for homozygous C3 deficiency. Restoration of the total hemolytic complement activity (CH 50) by purified C3 and the lack of anticomplement activity in the patient's serum indicate a defect in C3 synthesis. The ability of the patient's serum to mediate opsonization or generation of chemotactic factors was markedly impaired. The immune adherence of the patient's serum was normal and the reaction could be abolished by preincubation with anti-C4 antiserum.

Familial partial deficiency of the third component of complement (C3) and the hypocomplementemic cutaneous vasculitis syndrome

Familial hypocomplementemia of the third component of complement (C3) was found in four members of a family. The propositus had cutaneous vasculitis, hypocomplementemia, arthralgia, proteinuria and thrombocytopenia. The combination of clinical, laboratory and pathologic findings resembled the “hypocomplementemic cutaneous vasculitis syndrome” (HCVS) or the “SLE-like syndrome” but serum C3 concentration was 35 to 57 per cent of normal in the propositus and in three relatives. Results of Clq precipitins, cryoglobulins and serologic tests for systemic lupus erythematosus were negative. Proteinuria (815 mg/day) but no hematuria was present. Analysis of the C3 phenotypes in this family showed that three hypocomplementemic members were apparent homozygous C3 slow but one was heterozygous C3 fast-slow. Metabolic studies with 125-Iodinated C3 in the clinically normal mother showed a 50 per cent reduction in C3 synthesis which was consistent with hypocomplementemia documented by serum protein assay. The occurrence of an immune complex-like disease (with characteristics of the HCVS) in a patient with a familial deficiency of C3 suggests that the preexisting C3 deficiency may predispose such persons to certain diseases.

Homozygous C3 deficiency: Detection of C3 by radioimmunoassay

A 5 10 12 - year-old girl with recurrent infections was found to have homozygous C3 deficiency. Family studies demonstrated that the pattern of inheritance was consistent with that of an autosomal codominant trait. C3 levels have ranged from 5 mg/100 ml (electroimmunoassay) to 1.7 μg/100 ml (radioimmunoassay) 2 years later. C3 was also detected in two previous homozygous-deficient patients by radioimmunoassay. These findings suggest that an intact structural gene for C3 is present in these patients. In samples of the patient's serum with a C3 level less than 3 mg/100 ml, properdin factor B conversion did not occur upon incubation with zymosan. Immune adherence was normal. The vasopermeability response following intradermal injection of C 1s was normal. The patient's serum incubated with zymosan did not produce a chemotactic response of normal leukocytes. Phagocytosis of endotoxin-paraffin oil-oil red O particles by normal leukocytes was not enhanced following incubation with the patient's serum. Humoral antibody response and lymphocyte number and function were normal.