Complement component 7 is associated with total- and cardiac death in chest-pain patients with suspected acute coronary syndrome.

BACKGROUND: Complement activation has been associated with atherosclerosis, atherosclerotic plaque destabilization and increased risk of cardiovascular events. Complement component 7 (CC7) binds to the C5bC6 complex which is part of the terminal complement complex (TCC/C5b-9). High-sensitivity C-reactive protein (hsCRP) is a sensitive marker of systemic inflammation and may reflect the increased inflammatory state associated with cardiovascular disease. AIM: To evaluate the associations between CC7 and total- and cardiac mortality in patients hospitalized with chest-pain of suspected coronary origin, and whether combining CC7 with hsCRP adds prognostic information. METHODS: Baseline levels of CC7 were related to 60-months survival in a prospective, observational study of 982 patients hospitalized with a suspected acute coronary syndrome (ACS) at 9 hospitals in Salta, Argentina. A cox regression model, adjusting for conventional cardiovascular risk factors, was fitted with all-cause mortality, cardiac death and sudden cardiac death (SCD) as the dependent variables. A similar Norwegian population of 871 patients was applied to test the reproducibility of results in relation to total death. RESULTS: At follow-up, 173 patients (17.7%) in the Argentinean cohort had died, of these 92 (9.4%) were classified as cardiac death and 59 (6.0%) as SCD. In the Norwegian population, a total of 254 patients (30%) died. In multivariable analysis, CC7 was significantly associated with 60-months all-cause mortality [hazard ratio (HR) 1.26 (95% confidence interval (CI), 1.07-1.47) and cardiac death [HR 1.28 (95% CI 1.02-1.60)], but not with SCD. CC7 was only weakly correlated with hsCRP (r = 0.10, p = 0.002), and there was no statistically significant interaction between the two biomarkers in relation to outcome. The significant association of CC7 with total death was reproduced in the Norwegian population. CONCLUSIONS: CC7 was significantly associated with all-cause mortality and cardiac death at 60-months follow-up in chest-pain patients with suspected ACS. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT01377402, NCT00521976.

Screening and identification of potential protein biomarkers for evaluating the efficacy of intensive therapy in pulmonary tuberculosis.

This research aimed to discover potential biomarkers for evaluating the therapeutic efficacy of intensive therapy in pulmonary tuberculosis (TB). Protein profiles in 2-months intensively treated TB patients, untreated TB patients, and healthy controls were investigated with iTRAQ-2DLC-MS/MS technique. 71 differential proteins were identified in 2-months intensively treated TB patients. Significant differences in complement component C7 (CO7), apolipoprotein A-IV (APOA4), apolipoprotein C-II (APOC2), and angiotensinogen (ANGT) were found by ELISA validation. CO7 and ANGT were also found significantly different in sputum negative patients, compared with sputum positive patients after intensive treatment. Clinical analysis showed that after 2-months intensive treatment several indicators were significantly changed, and the one-year cure rate of sputum negative patients were significantly higher than sputum positive patients. Diagnostic models consisting of APOC2, CO7 and APOA4 were established to distinguish intensively treated TB patients from untreated TB patients and healthy controls with the AUC value of 0.910 and 0.935. Meanwhile, ANGT and CO7 were combined to identify sputum negative and sputum positive TB patients after intensive treatment with 89.36% sensitivity, 71.43% specificity, and the AUC value of 0.853. The results showed that APOC2, CO7, APOA4, and ANGT may be potential biomarkers for evaluating the efficacy of intensive anti-TB therapy.

Comparing serum protein levels can aid in differentiating HPV-negative and -positive oropharyngeal squamous cell carcinoma patients.

BACKGROUND: The surrogate immunohistochemical marker, p16INK4a, is used in clinical practice to determine the high-risk human papillomavirus (HPV) status of oropharyngeal squamous cell carcinomas (OPSCC). With a specificity of 83%, this will misclassify some patients compared with direct HPV testing. Patients who are p16INK4a-positive but HPV DNA-negative, or RNA-negative, may be unsuitable for treatment de-escalation aimed at reducing treatment-related side effects. We aimed to identify cost-effective serum markers to improve decision making for patients at risk of misclassification by p16INK4a alone. METHODS: Serum proteins from pre-treatment samples of 36 patients with OPSCC were identified and quantified using label-free mass spectrometry-based proteomics. HPV-status was determined using p16INK4a/HPV DNA and E6/E7 mRNA. Serum protein expressions were compared between groups of patients according to HPV status, using the unpaired t-test with a Benjamini-Hochberg correction. ROC curves (AUC) were calculated with SPSS (v25). RESULTS: Of 174 serum proteins identified, complement component C7 (C7), apolipoprotein F (ApoF) and galectin-3-Binding Protein (LGALS3BP) significantly differed between HPV-positive and -negative tumors (AUC ranging from 0.84-0.87). ApoF levels were more than twice as high in the E6/E7 mRNA HPV-positive group than HPV-negative. CONCLUSIONS: Serum C7, ApoF and LGALS3BP levels discriminate between HPV-positive and HPV-negative OPSCC. Further studies are needed to validate these host immunity-related proteins as markers for HPV-associated OPSCC.

Proteomic screening of plasma identifies potential noninvasive biomarkers associated with significant/advanced fibrosis in patients with nonalcoholic fatty liver disease.

Noninvasive biomarkers are clinically useful for evaluating liver fibrosis stage in patients with nonalcoholic fatty liver disease (NAFLD). The aim of the present study was to compare plasma proteins in patients with early nonalcoholic steatohepatitis (NASH) (F0-F1) versus NASH with significant/advanced fibrosis (F2-F4) to determine whether candidate proteins could be used as potential noninvasive biomarkers. Nineteen biopsy-proven NAFLD patients including ten early NASH patients and nine NASH patients with significant/advanced fibrosis were enrolled in the present study. High-resolution proteomics screening of plasma was performed with the SCIEX TripleTOF 5600 System. Proteins were quantified using two different software platforms, Progenesis Qi and Scaffold Q+, respectively. Progenesis Qi analysis resulted in the discovery of 277 proteins compared with 235 proteins in Scaffold Q+. Five consensus proteins (i.e. Complement component C7; α-2-macroglobulin; Complement component C8 γ chain; Fibulin-1; α-1-antichymotrypsin) were identified. Complement component C7 was three-fold higher in the NASH group with significant/advanced fibrosis (F2-F4) compared with the early NASH (F0-F1) group (q-value = 3.6E-6). Complement component C7 and Fibulin-1 are positively correlated with liver stiffness (P=0.000, P=0.002, respectively); whereas, Complement component C8 γ chain is negatively correlated (P=0.009). High levels of Complement C7 are associated with NASH with significant/advanced fibrosis and Complement C7 is a perfect classifier of patients included in this pilot study. Further studies will be needed in a larger validation cohort to confirm the utility of complement proteins as biomarkers or mechanistic determinants of NASH with significant/advanced fibrosis.

Abnormalities of complement and its components in patients with acute leukemia, Hodgkin's disease, and sarcoma.

Whole complement and component titers were measured in patients with acute leukemia, Hodgkin's disease, and sarcoma. Serum samples were obtained from 42 consecutive patients and 11 healthy control subjects. Sera were frozen and maintained at -70 degrees until analyzed by hemolytic assay. Titers were normalized using a titer obtained from a single source of pooled human serum analyzed simultaneously with each patient sample to correct for day-to-day variation inherent in the assay technique. Significant elevations (p less than or equal to 0.05) of whole complement and C5, C8, and C9 were observed for each patient category, compared to controls. Forty-one of 42 patients had C9 titers greater than or equal to 2 S.D. above the mean titer for controls. Mean C3 and C7 titers were not elevated or depressed in any group. No clinical factors that correlated with abnormal complement or component titers were identified.

Familial late complement component (C6, C7) deficiency with chronic meningococcemia.

Two patients with chronic meningococcemia were found to lack hemolytic complement, one because of C6 deficiency, the other because of C7 deficiency. In both cases family studies were consistent with inheritance of the deficiencies as non-HLA-linked, autosomal co-dominant traits. Functional studies showed the deficient sera to support monocyte chemotaxis but not phagocytosis or lysis of meningococci. Both patients have remained well following antibiotic treatment.

Characterization of human complement components C6 and C7.

Human complement components C6 and C7 have been purified and characterized. Component C6 is a single-chain plasma protein of mol. wt 104,800 containing 3.8% carbohydrate and it has alanine as the amino terminal residue. Component C7 is a single-chain plasma protein of mol. wt 92,400 containing 6.4% carbohydrate and it has serine as the amino terminal residue. Primary sequence analysis failed to provide evidence of homology between these two proteins.

Isolation of mouse complement component C7.

Mouse complement component C7 was purified from serum by a sequential procedure of fractionation precipitation by ammonium sulfate, followed by DE-52 anion exchange chromatography. Protein G affinity column chromatography, Mono S cation exchange chromatography and Superdex 200 gel filtration. The final product contained a highly purified mouse C7 component showing a single band on SDS-PAGE at the apparent Mrs of 90 kDa and 100 kDa under non-reduced and reduced conditions respectively. The yield of C7, which was measured by the biological activity, was 7.0%

Complement levels in normal and inflamed aqueous humor.

C2, C6, and C7 were measured by hemolytic assay and Factor B and IgG were measured by radial immunodiffusion in samples of normal and inflamed aqueous humor. Normal aqueous humor was found to contain functional C2, C6, and C7, but the small ratios of aqueous humor to serum measurements suggested that there was relatively little of these complement components in normal aqueous humor when compared to serum. The mean values of C2, C6, C7, and Factor B in aqueous humor and the median ratios of aqueous humor to serum measurements for each complement component were higher in patients with inflamed aqueous humor than in patients with normal aqueous humor. A comparison of the ratios of IgG to each complement component in normal and inflamed aqueous humor suggested that levels of IgG and complement increased proportionately in inflamed aqueous humor. Factor B, a component of the alternative pathway, was not detected in normal aqueous humor but measured in five of six samples of aqueous humor from eyes with anterior chamber inflammation.

Hemolytic complement activity in normal human donor corneas.

Normal human donor corneas were minced into small fragments and eluted for 16 to 23 hours at 4 degrees C. The corneal eluates were then studied for hemolytic complement activity of C1, C4, C2, C3, C5, C6, and C7 with 50% hemolysis (CH50) of sensitized sheep RBCs. Sera from ten normal volunteers were also assayed for hemolytic complement activity in CH50 units per milliliter. For each complement component, the mean hemolytic activity in corneas was compared with the mean hemolytic activity in sera. These comparisons suggest that molecular weight may be a factor in determining the concentration of complement components in the cornea. The present study provides normal values of hemolytic complement activity for further studies of complement consumption in corneal diseases.

Immunoglobulins and complement in pleural effusions associated with bronchogenic carcinoma.

Levels of IgG, IgA, IgM, the total haemolytic complement (CH50), and the individual components C1q, C3, C4, C6, and C7 were measured in 29 pleural effusions. Of these, 18 were associated with carcinoma of the bronchus and 11 were non-malignant effusions including empyemas. The level of IgG was significantly lower in the malignant group when compared with non-malignant effusions. The usefulness of measurements of IgG with respect to malignant effusions associated with carcinoma of the bronchus requires an expanded study to show whether it has any real diagnostic value. There were no significant differences in other immunoglobulins, the CH50, and individual complement components between the two groups. The identification of total haemolytic activity in the majority of effusions in both groups indicates that all nine components of the classical pathway of complement, including macromolecules such as C1, can be present in pleural fluids.

Testing of hemolytic complement components in domestic animals.

Total complement (C) and its components were assayed in the serum of 8 species of domestic animals, using commercially prepared cellular intermediates of sheep erythrocytes and functionally pure guinea pig and human components of the C system. Testing was done according to methods recommended by the producer for testing human C components. The late-acting components (C6 throug C9) and C1 were detected in carnivorous (dog and cat) and omnivorous (swine) animals. Undetectable or low titers of C4, C2, C3, and C5 were present in large herbivorous animals (cattle, horse, sheep, and goat), indicating major differences in comparison with human or guinea pig components of C. Porcine serum contained an inhibiting substance which interfered with testing C2 and later-acting components at serum dilutions up to 1:100. All components except C2 were detected in chicken serum. The binding or activation (or both) of C4, C2, C3, and C5 is more species specific than is the binding or activation (or both) of other components. Requirements for species specificity between antibody and C1 were not detected. Presence of C1 inactivator was detected in bovine, caprine, equine, and ovine sera. The CH50 (50% hemolysis) titers of C components tested in pooled serum samples from the 8 species of clinically healthy domestic animals are presented.

[Immunological study in sickle cell disease patients: importance of the complement system]. / Etude immunologique du drepanocytaire homozygote: importance du systeme complementaire.

Ten Tunisian patients, with homozygote sickle cell disease and asplenia were studied to investigate and to determine possible immunological function defects. Obtained results directed us to an abnormality of the alternate complement pathway activation which is expressed by a decreased hémolytic activity, while the classic pathway is normal. Quantification of C3, C4, C5, C6, C7 and factor B by immunochemical assay were normal, whereas factor B functional activity was depressed to a mean level of about half of normal in eight patients, IgG was increased in one subject and IgA in two others. Numeration of Band T cells revealed slight decrease in proportion of CD3 and CD4 at one patient associated with an increase in B cells, but normal or increased absolute numbers of all cells population.

C7*9, a new frequent C7 allele detected by an allotype-specific monoclonal antibody.

Two sandwich ELISA for the quantitation of C7 in human plasma--one based on a monoclonal (MAb), the other performed with a polyclonal anti-C7 IgG--were established. Results from ELISA as well as from isoelectric focussing and immunoblotting with both antibodies revealed that the MAb is directed to an allotypic epitope on C7. About 6.7% of the plasma samples did not react with this MAb. Since these findings cannot be explained by the known alleles, the existence of a new C7 allele, C7*9, was suggested, whose product C7-9 is not detected by the MAb. A gene frequency of 0.21 C7*9 was assumed. When using conventional techniques C7*9 is included in the frequency of C7*1.