Fine-mapping of immunodominant linear B-cell epitopes of
C.jejuni PEB1 antigen using short overlapping peptides- Journal of Third Military Medical University Vol. 41, Issue 4, Pages: 302-307(2019)
- 作者机构：
1.610083 成都，西部战区总医院：老年医学科
2.610083 成都，西部战区总医院：信息科
3.610083 成都，西部战区总医院：检验科
- 作者简介：
- 基金信息：the Notural Science Foundation for Young Scholars National Natural Science of China(31400128);the Training Project of Medical Young Scientists of PLA(孵化项目19QNP043)
DOI：10.16016/j.1000-5404.201809180
CLC： R378.99;R392-33;R392.13Received：26 September 2018，
Revised：2018-12-09，
Published：28 February 2019
- 稿件说明：
移动端阅览
Jian FENG, Yunming LI, Yugang LIU, et al. Fine-mapping of immunodominant linear B-cell epitopes of
C.jejuni PEB1 antigen using short overlapping peptides[J]. Journal of Third Military Medical University, 2019, 41(4): 302-307.Jian FENG, Yunming LI, Yugang LIU, et al. Fine-mapping of immunodominant linear B-cell epitopes of
C.jejuni PEB1 antigen using short overlapping peptides[J]. Journal of Third Military Medical University, 2019, 41(4): 302-307. DOI： 10.16016/j.1000-5404.201809180.
摘要
目的
2
利用表位预测分析技术筛选空肠弯曲菌(
C.jejuni
)黏附蛋白PEB1的B细胞免疫优势表位，并评价其免疫保护效果。
方法
2
采用氨基酸步移策略，合成18 mer氨基酸重叠肽段。ELISA系统筛选鉴定PEB1的B细胞免疫优势表位。采用KLH偶联免疫优势表位肽免疫BALB/c小鼠，ELISA测定优势表位肽诱导的IgG抗体效价。末次免疫后7 d，经口灌胃感染
C.jejuni
11168，在感染后的28 d，定量检测感染攻毒后各组小鼠的空肠组织
C.jejuni
的定植量以及qRT-PCR技术检测炎性细胞因子TNF-α的相对表达水平。通过HL-60细胞调理吞噬实验测定表位肽诱导产生抗体所介导的调理吞噬杀伤功能。免疫B细胞缺失小鼠，感染攻毒后检测肠组织
C.jejuni
的定植量。
结果
2
PEB1
55-72aa
、PEB1
97-114aa
、PEB1
211-228aa
均能与PEB1抗血清产生强烈IgG抗体反应。抗PEB1
55-72aa
、抗PEB1
97-114aa
和PEB1
211-228aa
血清均能与重组的PEB1产生较强的抗原抗体反应。与CFA/IFA组相比，免疫PEB1
55-72aa
、PEB1
97-114aa
、PEB1
211-228aa
后血清中的抗体能够显著增强抗体介导的HL-60细胞的调理吞噬作用(
P
<
0.01)，均显著降低
C.jejuni
在空肠组织中的定植量，同时也均显著降低炎性细胞因子TNF-α的相对表达水平(
P
<
0.01)。PEB1
55-72aa
-KLH+CFA/IFA组，PEB1
97-114aa
-KLH+CFA/IFA组，PEB1
211-228aa
-KLH+CFA/IFA组中，免疫B cell Knock out(B细胞缺失)小鼠攻毒后，
C.jejuni
在空肠组织中的定植量均显著高于WT(野生型)小鼠的定植量(
P
<
0.01)。
结论
2
成功鉴定出3个具有良好免疫原性和免疫保护作用的优势B细胞抗原表位(PEB1
55-72aa
、PEB1
97-114aa
、PEB1
211-228aa
)，可用于
C.jejuni
疫苗的后续开发研究。
Abstract
Objective
2
To map the immunodominant linear B-cell epitopes of
Campylobacter jejuni
(
C.jejuni
) PEB1 antigen and evaluate the protective immune responses elicited by these epitopes in a mouse model of oral infection with
C.jejuni
.
Methods
2
The B-cell immunodominant PEB1 epitopes were identified using synthetic overlapping peptide ELISA. B
ALB/c mice were immunized with the immunodominant PEB1 peptides conjugated with KLH plus CFA/IFA
and the IgG titers against these peptides were detected using ELISA. Seven days after the last immunization
the mice were orally infected with
C.jejuni
11168
and the bacterial burden and expression of tumor necrosis factor-α (TNF-α) mRNA in the jejunum were analyzed using qRT-PCR in 28 d after the challenge. We assessed the effect of the antibodies against the immunodominant PEB1 epitopes in mediating the opsonophagocytic killing of
C.jejuni
by HL-60 cells. We also assessed the bacterial burden in the jejunum following
C.jejuni
challenge in a B cell-knockout mice immunized with the peptides.
Results
2
The immunodominant peptides PEB1
55-72aa
PEB1
97-114aa
and PEB1
211-228aa
all induced strong IgG responses to PEB1 antiserum
and the antisera of these immunodominant peptides also showed strong IgG responses to recombinant PEB1. Compared with the antiserum of CFA/IFA
the antisera of these immunodominant peptides induced significantly enhanced opsonophagocytic activity of HL-60 cells (
P
<
0.01). Both the bacterial burdens and TNF-α mRNA expression level in the jejunum were significantly lowered in the mice immunized with the 3 immunodominant peptides in comparison with the control mice immunized with CFA/IFA (
P
<
0.01). In B cell-knockout mice
immunization with the 3 peptides did not provide immune protection against
C.jejuni
and the bacterial burden in the jejunum after
C.jejuni
challenge was significantly greater than that in immunized wild-type mice (
P
<
0.01).
Conclusion
2
We successfully identified 3 linear B-cell epitopes of
C.jejuni
PEB1 (PEB1
55-72aa
PEB1
97-114aa
and PEB1
211-228aa
)
which exhibit good immunogenicity and immuno
protective activities and may facilitate the future development of vaccines against
C.jejuni
infection.
关键词
Keywords
references
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