LONG-TERM EFFECTS OF BCG VACCINATION ON TELOMERE LENGTH AND TELOMERASE ACTIVITY

Vaccination with Bacillus Calmette-Guérin (BCG) induces long-term innate immune memory, also called trained immunity, characterized by metabolic and epigenetic changes leading to enhanced responsiveness upon exposure to heterologous pathogens. BCG vaccination was also shown to reduce systemic inflammation. Thus, BCG counteracts two significant immunological changes associated with aging: impaired responsiveness and elevated systemic inflammation. However, if and how BCG impacts other aging-related processes in immune cells, such as telomere shortening, remains unexplored. In this study, we investigated the transcriptional impact of BCG training on telomere maintenance-related genes through RNA sequencing and determined average telomere length from whole blood via RT-qPCR before and three months after BCG vaccination in two independent human cohorts. Trained immunity response was measured by ex-vivo cytokine production induced by a heterologous stimulus three months after vaccination compared to baseline. In addition, we examined the effects of BCG on telomerase activation using an in vitro trained immunity model. In vitro BCG training upregulated processes related to telomere maintenance and telomerase localization. In vivo, we observed shorter telomeres three months after BCG vaccination in both studies. Interestingly, the induction of a trained immunity response by BCG was correlated to the change in telomere length: more telomere shortening was observed in trained immunity non-responder individuals, particularly in males. Higher testosterone concentrations before vaccination were linked to more telomere loss. In vitro, BCG training of human monocytes activated the telomerase enzyme, predominantly in females, an effect that was blocked by exogenous testosterone treatment. Overall, this study reports sex-specific long-term impacts of BCG vaccination on telomerase activity and telomere maintenance. These data add to the arguments that BCG vaccination impacts aging mechanisms, which warrants more investigation. Pathway analysis of genes detected as differential in RNA sequencing data from in vitro BCG-trained and control adherent PBMCs of healthy donors.

卡介苗（Bacillus Calmette-Guérin, BCG）接种可诱导长期先天免疫记忆，又称训练免疫（trained immunity），其特征为代谢与表观遗传改变，使机体在遭遇异源病原体时的应答能力增强。研究证实，卡介苗接种还可降低全身性炎症水平。因此，卡介苗可抵消衰老相关的两大重要免疫学改变：应答能力受损与全身性炎症升高。然而，卡介苗是否以及如何影响免疫细胞内其他衰老相关进程（如端粒缩短（telomere shortening）），目前仍未得到探索。本研究纳入两独立人类队列，分别在卡介苗接种前及接种后3个月采集全血样本，通过RNA测序（RNA sequencing）分析卡介苗训练对端粒维持相关基因的转录调控影响，并通过实时定量聚合酶链反应（RT-qPCR）检测全血中的平均端粒长度。研究以接种后3个月时异源刺激诱导的离体细胞因子生成量与基线水平对比，以此评估训练免疫应答水平。此外，本研究利用体外训练免疫模型，探究了卡介苗对端粒酶激活（telomerase activation）的影响。体外实验结果显示，卡介苗训练可上调与端粒维持及端粒酶定位相关的生物学过程。体内实验中，两项队列均观察到卡介苗接种后3个月的端粒长度缩短。有趣的是，卡介苗诱导的训练免疫应答与端粒长度变化存在相关性：训练免疫无应答个体的端粒缩短更为显著，这一现象在男性群体中尤为突出。接种前睾酮浓度较高的个体，其端粒丢失程度也更为明显。体外实验中，卡介苗训练可激活人单核细胞的端粒酶活性，这一效应在女性个体中更为显著，而外源性睾酮处理可阻断该激活作用。综上，本研究揭示了卡介苗接种对端粒酶活性及端粒维持的性别特异性长期影响。上述研究结果进一步支持卡介苗接种可调控衰老机制这一观点，值得开展更深入的研究。本研究同时对健康供者体外卡介苗训练组与对照组贴壁外周血单个核细胞（PBMC）的RNA测序差异基因进行了通路分析。