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The Global Self-Reactivity Profile of the Natural Antibody Repertoire Is Largely Independent of Germline D H Sequence

View Article: PubMed Central - PubMed

ABSTRACT

Natural antibodies (NAbs) are produced in the absence of exogenous antigenic stimulation and circulate in the blood of normal, healthy individuals. These antibodies have been shown to provide one of the first lines of defense against both bacterial and viral pathogens. Conservation of the NAb repertoire reactivity profile is observed both within and across species. One view holds that this conservation of NAb self-reactivities reflects the use of germline antibody sequence, whereas the opposing view holds that the self-reactivities reflect selection driven by key conserved self-antigens. In mice, B-1a B cells are a major source of NAbs. A significant fraction of the B-1a antibody repertoire is devoid of N nucleotides in H chain complementarity determining region 3 (CDR-H3) and, thus, completely germline encoded. To test the role of germline DH sequence on the self-reactivity profile of the NAb repertoire, we examined the composition and self-antigen specificity of NAbs produced by a panel of DH gene-targeted BALB/c mice, each strain of which expresses a polyclonal, altered CDR-H3 repertoire that differs from the wild-type norm. We found that in most cases the same key self-antigens were recognized by the NAbs created by each DH-altered strain. The differences in reactivity appeared to represent the genetic signature of the NAb repertoire of each mouse strain. These findings suggest that although germline CDR-H3 sequence may facilitate the production of certain NAbs, a core set of self-antigens are likely the main force driving the selection of Nab self-specificities.

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DH-altered B cells response to polyclonal stimuli. (A) Frequencies of response to LPS of PerC B cell subsets of each mouse strain were calculated by limiting dilution analysis. Presence of IgM in culture supernatants of 18, 6, 2, and 0.66 cells/culture on day 7 was assessed by ELISA. (B) IgM production by each PerC B cell subset in culture was measured by ELISA on culture supernatants of increasing number of cells/culture (370, 1111, 3333, and 10,000 cells/culture). PerC B-1a cells were sorted as B220loCD5+, B-1b as B220loCD5− Mac-1lo/+, and PerC B-2 cells were sorted as B220hiCD5−Mac-1−. All cultures received 30 μg/mL of LPS and 5 × 103 S17 feeder cells. Supernatants were collected on day 7.
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Figure 2: DH-altered B cells response to polyclonal stimuli. (A) Frequencies of response to LPS of PerC B cell subsets of each mouse strain were calculated by limiting dilution analysis. Presence of IgM in culture supernatants of 18, 6, 2, and 0.66 cells/culture on day 7 was assessed by ELISA. (B) IgM production by each PerC B cell subset in culture was measured by ELISA on culture supernatants of increasing number of cells/culture (370, 1111, 3333, and 10,000 cells/culture). PerC B-1a cells were sorted as B220loCD5+, B-1b as B220loCD5− Mac-1lo/+, and PerC B-2 cells were sorted as B220hiCD5−Mac-1−. All cultures received 30 μg/mL of LPS and 5 × 103 S17 feeder cells. Supernatants were collected on day 7.

Mentions: All three subsets from DH-altered PerC B cells demonstrated frequencies of response to LPS that were similar to their corresponding WT B cell subsets (Figure 2A). Production of IgM in culture was also similar (Figure 2B). Thus, DH alteration and restriction of the CDR-H3 repertoire did not affect the B cell response to polyclonal stimulus from LPS.


The Global Self-Reactivity Profile of the Natural Antibody Repertoire Is Largely Independent of Germline D H Sequence
DH-altered B cells response to polyclonal stimuli. (A) Frequencies of response to LPS of PerC B cell subsets of each mouse strain were calculated by limiting dilution analysis. Presence of IgM in culture supernatants of 18, 6, 2, and 0.66 cells/culture on day 7 was assessed by ELISA. (B) IgM production by each PerC B cell subset in culture was measured by ELISA on culture supernatants of increasing number of cells/culture (370, 1111, 3333, and 10,000 cells/culture). PerC B-1a cells were sorted as B220loCD5+, B-1b as B220loCD5− Mac-1lo/+, and PerC B-2 cells were sorted as B220hiCD5−Mac-1−. All cultures received 30 μg/mL of LPS and 5 × 103 S17 feeder cells. Supernatants were collected on day 7.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4979587&req=5

Figure 2: DH-altered B cells response to polyclonal stimuli. (A) Frequencies of response to LPS of PerC B cell subsets of each mouse strain were calculated by limiting dilution analysis. Presence of IgM in culture supernatants of 18, 6, 2, and 0.66 cells/culture on day 7 was assessed by ELISA. (B) IgM production by each PerC B cell subset in culture was measured by ELISA on culture supernatants of increasing number of cells/culture (370, 1111, 3333, and 10,000 cells/culture). PerC B-1a cells were sorted as B220loCD5+, B-1b as B220loCD5− Mac-1lo/+, and PerC B-2 cells were sorted as B220hiCD5−Mac-1−. All cultures received 30 μg/mL of LPS and 5 × 103 S17 feeder cells. Supernatants were collected on day 7.
Mentions: All three subsets from DH-altered PerC B cells demonstrated frequencies of response to LPS that were similar to their corresponding WT B cell subsets (Figure 2A). Production of IgM in culture was also similar (Figure 2B). Thus, DH alteration and restriction of the CDR-H3 repertoire did not affect the B cell response to polyclonal stimulus from LPS.

View Article: PubMed Central - PubMed

ABSTRACT

Natural antibodies (NAbs) are produced in the absence of exogenous antigenic stimulation and circulate in the blood of normal, healthy individuals. These antibodies have been shown to provide one of the first lines of defense against both bacterial and viral pathogens. Conservation of the NAb repertoire reactivity profile is observed both within and across species. One view holds that this conservation of NAb self-reactivities reflects the use of germline antibody sequence, whereas the opposing view holds that the self-reactivities reflect selection driven by key conserved self-antigens. In mice, B-1a B cells are a major source of NAbs. A significant fraction of the B-1a antibody repertoire is devoid of N nucleotides in H chain complementarity determining region 3 (CDR-H3) and, thus, completely germline encoded. To test the role of germline DH sequence on the self-reactivity profile of the NAb repertoire, we examined the composition and self-antigen specificity of NAbs produced by a panel of DH gene-targeted BALB/c mice, each strain of which expresses a polyclonal, altered CDR-H3 repertoire that differs from the wild-type norm. We found that in most cases the same key self-antigens were recognized by the NAbs created by each DH-altered strain. The differences in reactivity appeared to represent the genetic signature of the NAb repertoire of each mouse strain. These findings suggest that although germline CDR-H3 sequence may facilitate the production of certain NAbs, a core set of self-antigens are likely the main force driving the selection of Nab self-specificities.

No MeSH data available.


Related in: MedlinePlus