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Pathogenic profiles and molecular signatures of antinuclear autoantibodies rescued from NZM2410 lupus mice.

Liang Z, Xie C, Chen C, Kreska D, Hsu K, Li L, Zhou XJ, Mohan C - J. Exp. Med. (2004)

Bottom Line: Most intriguingly, the CDR3 regions of the ANAs exhibited alternating arginine/lysine peaks at H96, H98, and H100, with neutral troughs at H95, H97, and H99.To summarize, glomerular-binding anti-dsDNA antibodies appear to be the most pathogenic variety of lupus autoantibodies.The presence of an alternating charge pattern in their HC CDR3 regions appears to be a prominent hallmark of ANAs.

View Article: PubMed Central - PubMed

Affiliation: Simmons Arthritis Research Center, University of Texas Southwestern Medical School, Dallas 75390, USA.

ABSTRACT
Two outstanding questions concerning antinuclear antibodies (ANAs) in lupus involve their pathogenic potential and their molecular signatures. To address these questions, a panel of 56 antinuclear and 47 nonnuclear binding monoclonal antibodies was rescued from four seropositive NZM2410 lupus mice. The monoclonals varied in their reactivity to nucleosomes, ssDNA, dsDNA, and glomerular substrate. A large fraction of the antibodies demonstrated apparent polyreactivity (to DNA, histones, and glomerular antigens) due to bound, DNase-1 sensitive nuclear antigenic bridges. Although nephrophilic immunoglobulin (Ig) M and IgG antibodies were the most pathogenic, the dsDNA-binding antibodies were modestly so; in contrast, antinucleosome antibodies were clearly not pathogenic. Compared with the nonnuclear antigen-binding monoclonal antibodies rescued from the same mice, ANAs exhibited increased utilization of VH5/7183 genes and highly cationic heavy chain (HC) CDR3 regions. Most intriguingly, the CDR3 regions of the ANAs exhibited alternating arginine/lysine peaks at H96, H98, and H100, with neutral troughs at H95, H97, and H99. To summarize, glomerular-binding anti-dsDNA antibodies appear to be the most pathogenic variety of lupus autoantibodies. The presence of an alternating charge pattern in their HC CDR3 regions appears to be a prominent hallmark of ANAs.

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Renal pathogenicity of NZM2410-derived IgG ANAs. (A) Seven representative NZM2410-derived IgG mAbs were tested for in vivo pathogenicity as detailed in Materials and Methods. Although ZB4D8, ZDB4, and ZB1A7 were IgG1, IgG2b, and IgG3 in isotype, respectively, all other Abs were IgG2a in isotype. The specificity profiles of these Abs that have been tabulated below the figure were obtained from Table I. It should be noted that “+” in this figure simply indicates that the mAb reacts with the respective Ag, independent of the strength of reactivity. Both the beginning (D0, white dots), and ending (D10, black dots) 24-h urine protein levels (measured using metabolic cages) are depicted. Where the D10 proteinuria levels were found to be significantly higher than the corresponding D0 levels (using the Student's t test), the p-values are listed. (B) The 24-h urine protein excretion profiles of the 10 mice injected with IgG anti-dsDNA/glomerular dual binding Abs, ZB1B11 and ZB4D8, are depicted. 24-h urine protein measurements were performed on D0, D3, D7, and D10 after mAb administration. (C) The BUN levels measured on D0 and D10 after administration of the dsDNA/glomerular dual binding Abs, ZB4D8 and ZB1B11, are depicted. Depicted below each column of dots are the p-values calculated when the D0 and D10 BUN values were compared (using the Student's t test), and found to be significantly different. The dotted line refers to the mean D10 BUN level noted in all the other experimental groups of mice, combined.
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fig3: Renal pathogenicity of NZM2410-derived IgG ANAs. (A) Seven representative NZM2410-derived IgG mAbs were tested for in vivo pathogenicity as detailed in Materials and Methods. Although ZB4D8, ZDB4, and ZB1A7 were IgG1, IgG2b, and IgG3 in isotype, respectively, all other Abs were IgG2a in isotype. The specificity profiles of these Abs that have been tabulated below the figure were obtained from Table I. It should be noted that “+” in this figure simply indicates that the mAb reacts with the respective Ag, independent of the strength of reactivity. Both the beginning (D0, white dots), and ending (D10, black dots) 24-h urine protein levels (measured using metabolic cages) are depicted. Where the D10 proteinuria levels were found to be significantly higher than the corresponding D0 levels (using the Student's t test), the p-values are listed. (B) The 24-h urine protein excretion profiles of the 10 mice injected with IgG anti-dsDNA/glomerular dual binding Abs, ZB1B11 and ZB4D8, are depicted. 24-h urine protein measurements were performed on D0, D3, D7, and D10 after mAb administration. (C) The BUN levels measured on D0 and D10 after administration of the dsDNA/glomerular dual binding Abs, ZB4D8 and ZB1B11, are depicted. Depicted below each column of dots are the p-values calculated when the D0 and D10 BUN values were compared (using the Student's t test), and found to be significantly different. The dotted line refers to the mean D10 BUN level noted in all the other experimental groups of mice, combined.

Mentions: As evident from Fig. 3 A, uninjected NZW mice typically excreted 0.4–1.0 mg urinary protein daily, with occasional mice excreting >1 mg/d. Clearly, the control non-ANA Ab as well as the IgG ANAs with reactivity to histone–DNA nucleosomal complexes and/or ssDNA (but not dsDNA) did not compromise renal function. In contrast, dsDNA-reactive IgG ANAs that lacked glomerular reactivity (i.e., ZB5D3 and ZA7H10) caused significantly increased proteinuria on D10, in the range of 1–1.5 mg/d. Interestingly, with both the histone/DNA–reactive Abs (ZDB4 and ZB1A7) and the dsDNA-reactive Abs (ZB5D3 and ZA7H10), concomitant reactivity to ssDNA did not affect the pathogenic potential of these Abs.


Pathogenic profiles and molecular signatures of antinuclear autoantibodies rescued from NZM2410 lupus mice.

Liang Z, Xie C, Chen C, Kreska D, Hsu K, Li L, Zhou XJ, Mohan C - J. Exp. Med. (2004)

Renal pathogenicity of NZM2410-derived IgG ANAs. (A) Seven representative NZM2410-derived IgG mAbs were tested for in vivo pathogenicity as detailed in Materials and Methods. Although ZB4D8, ZDB4, and ZB1A7 were IgG1, IgG2b, and IgG3 in isotype, respectively, all other Abs were IgG2a in isotype. The specificity profiles of these Abs that have been tabulated below the figure were obtained from Table I. It should be noted that “+” in this figure simply indicates that the mAb reacts with the respective Ag, independent of the strength of reactivity. Both the beginning (D0, white dots), and ending (D10, black dots) 24-h urine protein levels (measured using metabolic cages) are depicted. Where the D10 proteinuria levels were found to be significantly higher than the corresponding D0 levels (using the Student's t test), the p-values are listed. (B) The 24-h urine protein excretion profiles of the 10 mice injected with IgG anti-dsDNA/glomerular dual binding Abs, ZB1B11 and ZB4D8, are depicted. 24-h urine protein measurements were performed on D0, D3, D7, and D10 after mAb administration. (C) The BUN levels measured on D0 and D10 after administration of the dsDNA/glomerular dual binding Abs, ZB4D8 and ZB1B11, are depicted. Depicted below each column of dots are the p-values calculated when the D0 and D10 BUN values were compared (using the Student's t test), and found to be significantly different. The dotted line refers to the mean D10 BUN level noted in all the other experimental groups of mice, combined.
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Related In: Results  -  Collection

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fig3: Renal pathogenicity of NZM2410-derived IgG ANAs. (A) Seven representative NZM2410-derived IgG mAbs were tested for in vivo pathogenicity as detailed in Materials and Methods. Although ZB4D8, ZDB4, and ZB1A7 were IgG1, IgG2b, and IgG3 in isotype, respectively, all other Abs were IgG2a in isotype. The specificity profiles of these Abs that have been tabulated below the figure were obtained from Table I. It should be noted that “+” in this figure simply indicates that the mAb reacts with the respective Ag, independent of the strength of reactivity. Both the beginning (D0, white dots), and ending (D10, black dots) 24-h urine protein levels (measured using metabolic cages) are depicted. Where the D10 proteinuria levels were found to be significantly higher than the corresponding D0 levels (using the Student's t test), the p-values are listed. (B) The 24-h urine protein excretion profiles of the 10 mice injected with IgG anti-dsDNA/glomerular dual binding Abs, ZB1B11 and ZB4D8, are depicted. 24-h urine protein measurements were performed on D0, D3, D7, and D10 after mAb administration. (C) The BUN levels measured on D0 and D10 after administration of the dsDNA/glomerular dual binding Abs, ZB4D8 and ZB1B11, are depicted. Depicted below each column of dots are the p-values calculated when the D0 and D10 BUN values were compared (using the Student's t test), and found to be significantly different. The dotted line refers to the mean D10 BUN level noted in all the other experimental groups of mice, combined.
Mentions: As evident from Fig. 3 A, uninjected NZW mice typically excreted 0.4–1.0 mg urinary protein daily, with occasional mice excreting >1 mg/d. Clearly, the control non-ANA Ab as well as the IgG ANAs with reactivity to histone–DNA nucleosomal complexes and/or ssDNA (but not dsDNA) did not compromise renal function. In contrast, dsDNA-reactive IgG ANAs that lacked glomerular reactivity (i.e., ZB5D3 and ZA7H10) caused significantly increased proteinuria on D10, in the range of 1–1.5 mg/d. Interestingly, with both the histone/DNA–reactive Abs (ZDB4 and ZB1A7) and the dsDNA-reactive Abs (ZB5D3 and ZA7H10), concomitant reactivity to ssDNA did not affect the pathogenic potential of these Abs.

Bottom Line: Most intriguingly, the CDR3 regions of the ANAs exhibited alternating arginine/lysine peaks at H96, H98, and H100, with neutral troughs at H95, H97, and H99.To summarize, glomerular-binding anti-dsDNA antibodies appear to be the most pathogenic variety of lupus autoantibodies.The presence of an alternating charge pattern in their HC CDR3 regions appears to be a prominent hallmark of ANAs.

View Article: PubMed Central - PubMed

Affiliation: Simmons Arthritis Research Center, University of Texas Southwestern Medical School, Dallas 75390, USA.

ABSTRACT
Two outstanding questions concerning antinuclear antibodies (ANAs) in lupus involve their pathogenic potential and their molecular signatures. To address these questions, a panel of 56 antinuclear and 47 nonnuclear binding monoclonal antibodies was rescued from four seropositive NZM2410 lupus mice. The monoclonals varied in their reactivity to nucleosomes, ssDNA, dsDNA, and glomerular substrate. A large fraction of the antibodies demonstrated apparent polyreactivity (to DNA, histones, and glomerular antigens) due to bound, DNase-1 sensitive nuclear antigenic bridges. Although nephrophilic immunoglobulin (Ig) M and IgG antibodies were the most pathogenic, the dsDNA-binding antibodies were modestly so; in contrast, antinucleosome antibodies were clearly not pathogenic. Compared with the nonnuclear antigen-binding monoclonal antibodies rescued from the same mice, ANAs exhibited increased utilization of VH5/7183 genes and highly cationic heavy chain (HC) CDR3 regions. Most intriguingly, the CDR3 regions of the ANAs exhibited alternating arginine/lysine peaks at H96, H98, and H100, with neutral troughs at H95, H97, and H99. To summarize, glomerular-binding anti-dsDNA antibodies appear to be the most pathogenic variety of lupus autoantibodies. The presence of an alternating charge pattern in their HC CDR3 regions appears to be a prominent hallmark of ANAs.

Show MeSH
Related in: MedlinePlus