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MZ B cells from 14-week-old WT C57BL/6 and CD55 KO mice

ABSTRACT: MZ B cells from 14-week-old WT C57BL/6 and CD55 KO mice

PROVIDER: PRJNA1238830 | ENA |

REPOSITORIES: ENA

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			Action	DRS
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	SRR32788719_1.fastq.gz	Fastqsanger.gz
	SRR32788719_2.fastq.gz	Fastqsanger.gz
	SRR32788722_1.fastq.gz	Fastqsanger.gz
	SRR32788722_2.fastq.gz	Fastqsanger.gz

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MZ B cells from 14-week-old WT C57BL/6 and CD55 KO mice

Project description:Marginal zone (MZ) B cells bridge innate and adaptive immunity by sensing bloodborne antigens and producing rapid antibody and cytokine responses. When unregulated, MZ B cells are associated with autoimmunity. CD55 is a membrane-bound complement regulator that interferes with complement activation and is another important component of innate and adaptive immunity. MZ B cells express low CD55 in both mice and humans, but the role of CD55 in MZ B cell function is unknown. Using germline knockout mice, we found that similar numbers of MZ B cells were initially established in 3-week-old CD55-deficient mice compared to wild-type (WT) mice. However, MZ B cells failed to accumulate as mice aged and underwent increased apoptosis independent of alternative complement activation. Following ex vivo stimulation of MZ B cells through Toll-like receptor 9 (TLR9), we observed increased IL-6 expression in CD55 KO MZ B cells. In addition, CD55 KO mice exhibited reduced total IgG response with in vivo administration of TLR9 agonist. These findings provide new insights to the role of CD55 in MZ B cell survival and B cell function.

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Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare the genes expression difference at transcriptome level; Methods: Total RNA was extracted from whole cells with the mirVana miRNA Isolation Kit according to the manufacturer’s protocol. RNA quality and integrity were evaluated with an Agilent 2100 Bioanalyzer. Samples with an RNA integrity number (RIN) ≥ 7 were considered to be of high quality and were processed further and subjected to subsequent analysis. Total RNA-seq libraries were generated using 4 μg of total RNA, which was analyzed using the TruSeq Stranded mRNA LTSample Prep Kit. These libraries were then sequenced using the Illumina sequencing platform (HiSeqTM 2500 or Illumina HiSeq X Ten), and 125-bp/150-bp paired-end reads were generated. Results: The raw reads containing adaptors and the low-quality reads from the raw data were removed using Trimmomatic to obtain clean reads. Transcriptome sequencing was conducted by OE Biotech Co., Ltd. (Shanghai, China), and clean reads were provided. The clean reads were mapped to the hg38 reference genome using hisat2 (version 2.1.0). The output BAM files were converted to SAM files using SAMtools 1.9. The final TPM values were obtained using Stringtie 1.3.5. Conclusions: To understand the mechanistic basis of GPI biosynthesis upregulation by the CD55 precursor, we performed RNA- sequencing (RNA-seq) of samples of parental PIGS-HRD1-DKO, PIGS-HRD1-CD55-TKO, and PIGS-HRD1-CD55-TKO stably overexpressed HA-CD55 stably overexpressing cells. Total RNA was extracted and analyzed. The expression profile of GPI biosynthesis -related genes was not significantly affected by CD55.

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