State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, the First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003, China.
J Transl Med. 2012 Sep 15;10:193. doi: 10.1186/1479-5876-10-193.
CD4(+)CD25(+) regulatory T cells (Tregs) play an important role in maintaining immunological tolerance to self and foreign antigens. T cell receptors (TCR) reflect the composition and function of T cells. It is not universally agreed that there is a relationship between CD4(+)CD25(+) Treg frequency and the severity of acute-on-chronic liver failure (ACLF). The repertoire of TCR beta chain variable (TCRBV) regions of peripheral Tregs in ACLF patients is not well understood.
Human PBMCs were separated and sorted into CD4(+)CD25(+) Treg subsets using density gradient centrifugation and magnetic activated cell sorting (MACS). The CD4(+)CD25(high) Treg frequency in peripheral blood of ACLF and chronic hepatitis B (CHB) patients was measured by flow cytometry. The molecular profiles of TCRBV CDR3 were determined using gene melting spectral pattern (GMSP) analysis. TCRBV gene families were cloned and sequenced when the GMSP profiles showed a single-peak.
CD4(+)CD25(high) Treg prevalence in peripheral blood of ACLF patients is increased significantly compared to healthy donors (HDs) (P < 0.01) and CHB patients (P < 0.01). The prevalence of CD4(+)CD25(high) Tregs in ACLF or CHB patients is positively correlated with HBV DNA load. The TCRBV11, BV13.1, BV18, BV20 are the most prevalent TCRBV in CD4(+)CD25(+) Tregs in ACLF and CHB patients. In addition, the CDR3 motifs were relatively conserved in these four TCRBV gene families.
The CD4(+)CD25(high) Tregs prevalence in peripheral blood is indicative of disease severity in ACLF or CHB patients. The relatively conserved TCRBV20 CDR3 motif "TGTGHSPLH" and TCRBV11 CDR3 motif "VYNEQ" may be used in helping diagnosis and treat patients with ACLF.
CD4(+)CD25(+)调节性 T 细胞（Tregs）在维持对自身和外来抗原的免疫耐受方面发挥着重要作用。T 细胞受体（TCR）反映了 T 细胞的组成和功能。CD4(+)CD25(+)Treg 频率与急性加重慢性肝衰竭（ACLF）的严重程度之间是否存在关系尚未得到普遍认可。ACLF 患者外周血 Tregs 的 TCRβ链可变（TCRBV）区库的情况尚不清楚。
使用密度梯度离心和磁激活细胞分选（MACS）分离和分选人 PBMC 成 CD4(+)CD25(+)Treg 亚群。通过流式细胞术测量 ACLF 和慢性乙型肝炎（CHB）患者外周血中 CD4(+)CD25(high)Treg 的频率。使用基因熔融光谱图（GMSP）分析确定 TCRBV CDR3 的分子谱。当 GMSP 谱显示单峰时，克隆和测序 TCRBV 基因家族。
与健康供体（HDs）（P<0.01）和 CHB 患者（P<0.01）相比，ACLF 患者外周血中 CD4(+)CD25(high)Treg 的发生率明显升高。ACLF 或 CHB 患者中 CD4(+)CD25(high)Tregs 的发生率与 HBV DNA 载量呈正相关。TCRBV11、BV13.1、BV18 和 BV20 是 ACLF 和 CHB 患者中 CD4(+)CD25(+)Tregs 中最常见的 TCRBV。此外，这四个 TCRBV 基因家族的 CDR3 基序相对保守。
外周血中 CD4(+)CD25(high)Tregs 的发生率提示 ACLF 或 CHB 患者疾病的严重程度。相对保守的 TCRBV20 CDR3 基序“TGTGHSPLH”和 TCRBV11 CDR3 基序“VYNEQ”可用于帮助诊断和治疗 ACLF 患者。
