Comprehensive Analysis of Mannan-Binding Lectin (MBL) Status in a Cohort of Healthy Donors as Determined by MBL Protein Concentration, MBL Complement Pathway Function and MBL and MASP-2 Genotypes

Abstract

RATIONALE: Comprehensive analysis of MBL status is critical for evaluating overall immune function in patients with unexplained recurrent infections and will likely be important for characterizing patients prior to administering recombinant human MBL (rhMBL) therapy.METHODS: A cohort of 112 healthy adult donors was evaluated. MBL protein concentrations were determined using both a double-antibody and mannan-binding ELISA. Functional activity of the MBL complement pathway was evaluated with a C4b-deposition assay. MBL and MASP-2 genotypes were determined with real-time PCR.RESULTS: Significant correlation was observed between the two MBL protein ELISAs (r=0.96). Mean MBL concentrations for the entire cohort were 1705 ng/mL (SD = 1745, 98%CI = 384), as measured by the double-antibody and 2055 ng/mL (SD = 1999, 98%CI = 439) by mannan-binding assay. MBL concentrations obtained with either MBL protein ELISA correlated well with C4b-deposition (r=0.8). Genotypic analysis revealed that donors without MBL2 structural polymorphisms had the highest MBL concentrations, with the exception of some donors with LX/LX promoter genotypes. Donors who were either heterozygous (A/O), compound heterozygous (O/O) or homozygous (O/O) for MBL2 structural polymorphisms had either intermediate or profoundly reduced concentrations of circulating MBL. C4b-deposition was significantly reduced in individuals with the MBL genotypes LXPA/LYPB, LXPA/LYQC, LYPB/HYPD and LYPB/LYPB. The presence of variant MASP-2 polymorphisms was associated with reduced C4b-deposition in some donors.CONCLUSIONS: Either of the two MBL protein assays in combination with genotyping is informative in evaluating an individual's MBL status. Determining MBL pathway function will be important for patients who have reduced MBL protein concentrations due to A/O genotypes or when MASP-2 deficiency is suspected. RATIONALE: Comprehensive analysis of MBL status is critical for evaluating overall immune function in patients with unexplained recurrent infections and will likely be important for characterizing patients prior to administering recombinant human MBL (rhMBL) therapy. METHODS: A cohort of 112 healthy adult donors was evaluated. MBL protein concentrations were determined using both a double-antibody and mannan-binding ELISA. Functional activity of the MBL complement pathway was evaluated with a C4b-deposition assay. MBL and MASP-2 genotypes were determined with real-time PCR. RESULTS: Significant correlation was observed between the two MBL protein ELISAs (r=0.96). Mean MBL concentrations for the entire cohort were 1705 ng/mL (SD = 1745, 98%CI = 384), as measured by the double-antibody and 2055 ng/mL (SD = 1999, 98%CI = 439) by mannan-binding assay. MBL concentrations obtained with either MBL protein ELISA correlated well with C4b-deposition (r=0.8). Genotypic analysis revealed that donors without MBL2 structural polymorphisms had the highest MBL concentrations, with the exception of some donors with LX/LX promoter genotypes. Donors who were either heterozygous (A/O), compound heterozygous (O/O) or homozygous (O/O) for MBL2 structural polymorphisms had either intermediate or profoundly reduced concentrations of circulating MBL. C4b-deposition was significantly reduced in individuals with the MBL genotypes LXPA/LYPB, LXPA/LYQC, LYPB/HYPD and LYPB/LYPB. The presence of variant MASP-2 polymorphisms was associated with reduced C4b-deposition in some donors. CONCLUSIONS: Either of the two MBL protein assays in combination with genotyping is informative in evaluating an individual's MBL status. Determining MBL pathway function will be important for patients who have reduced MBL protein concentrations due to A/O genotypes or when MASP-2 deficiency is suspected.