Single-cell transcriptomic profiles reveal changes associated with BCG-induced trained immunity and protective effects in circulating monocytes.

Bacillus Calmette-Guérin (BCG) vaccine is one of the most widely used vaccines worldwide. In addition to protection against tuberculosis, BCG confers a degree of non-specific protection against other infections by enhancing secondary immune responses to heterologous pathogens, termed "trained immunity." To better understand BCG-induced immune reprogramming, we perform single-cell transcriptomic measurements before and after BCG vaccination using secondary immune stimulation with bacterial lipopolysaccharide (LPS). We find that BCG reduces systemic inflammation and identify 75 genes with altered LPS responses, including inflammatory mediators such as CCL3 and CCL4 that have a heightened response. Co-expression analysis reveals that gene modules containing these cytokines lose coordination after BCG. Other modules exhibit increased coordination, including several humanin nuclear isoforms that we confirm induce trained immunity in vitro. Our results link in vivo BCG administration to single-cell transcriptomic changes, validated in human genetics experiments, and highlight genes that are putatively responsible for non-specific protective effects of BCG.

Kong L ，Moorlag SJCFM ，Lefkovith A ，Li B ，Matzaraki V ，van Emst L ，Kang HA ，Latorre I ，Jaeger M ，Joosten LAB ，Netea MG ，Xavier RJ ... -  《Cell Reports》

BCG vaccination in humans inhibits systemic inflammation in a sex-dependent manner.

Koeken VA ，de Bree LCJ ，Mourits VP ，Moorlag SJ ，Walk J ，Cirovic B ，Arts RJ ，Jaeger M ，Dijkstra H ，Lemmers H ，Joosten LA ，Benn CS ，van Crevel R ，Netea MG ... -  《-》

Seasonal variation in BCG-induced trained immunity.

The Bacille Calmette-Guerin (BCG) vaccine is a well-established inducer of innate immune memory (also termed trained immunity), causing increased cytokine production upon heterologous secondary stimulation. Innate immune responses are known to be influenced by season, but whether seasons impact induction of trained immunity is not known. To explore the influence of season on innate immune memory induced by the BCG vaccine, we vaccinated healthy volunteers with BCG either during winter or spring. Three months later, we measured the ex vivo cytokine responses against heterologous stimuli, analyzed gene expressions and epigenetic signatures of the immune cells, and compared these with the baseline before vaccination. BCG vaccination during winter induced a stronger increase in the production of pro-inflammatory cytokines by peripheral blood mononuclear cells (PBMCs) upon stimulation with different bacterial and fungal stimuli, compared to BCG vaccination in spring. In contrast, winter BCG vaccination resulted in lower IFNγ release in PBMCs compared to spring BCG vaccination. Furthermore, NK cells of the winter-vaccinated people had a greater pro-inflammatory cytokine and IFNγ production capacity upon heterologous stimulation. BCG had only minor effects on the transcriptome of monocytes 3 months later. In contrast, we identified season-dependent epigenetic changes in monocytes and NK cells induced by vaccination, partly explaining the higher immune cell reactivity in the winter BCG vaccination group. These results suggest that BCG vaccination during winter is more prone to induce a robust trained immunity response by activating and reprogramming the immune cells, especially NK cells. (Dutch clinical trial registry no. NL58219.091.16).

Kilic G ，Debisarun PA ，Alaswad A ，Baltissen MP ，Lamers LA ，de Bree LCJ ，Benn CS ，Aaby P ，Dijkstra H ，Lemmers H ，Martens JHA ，Domínguez-Andrés J ，van Crevel R ，Li Y ，Xu CJ ，Netea MG ... -  《-》

Human Newborn Monocytes Demonstrate Distinct BCG-Induced Primary and Trained Innate Cytokine Production and Metabolic Activation In Vitro.

Newborns exhibit distinct immune responses and are at high risk of infection. Neonatal immunization with BCG, the live attenuated vaccine against tuberculosis (TB), is associated with broad protection against a range of unrelated pathogens, possibly reflecting vaccine-induced training of innate immune cells ("innate memory"). However, little is known regarding the impact of age on BCG-induced innate responses. Establish an age-specific human monocyte in vitro training platform to characterize and compare BCG-induced primary and memory cytokine responses and immunometabolic shifts. Human neonatal and adult CD33-selected monocytes were stimulated for 24h with RPMI (control) or BCG (Danish strain) in 10% autologous serum, washed and cultured for 5 additional days, prior to re-stimulation with the TLR4 agonist LPS for another 24h. Supernatants were collected at Day 1 (D1) to measure primary innate responses and at Day 7 (D7) to assess memory innate responses by ELISA and multiplex cytokine and chemokine assays. Lactate, a signature metabolite increased during trained immunity, was measured by colorimetric assay. Cytokine production by human monocytes differed significantly by age at D1 (primary, BCG 1:750 and 1:100 vol/vol, p<0.0001) and D7 (innate memory response, BCG 1:100 vol/vol, p<0.05). Compared to RPMI control, newborn monocytes demonstrated greater TNF (1:100, 1:10 vol/vol, p<0.01) and IL-12p40 (1:100 vol/vol, p<0.05) production than adult monocytes (1:100, p<0.05). At D7, while BCG-trained adult monocytes, as previously reported, demonstrated enhanced LPS-induced TNF production, BCG-trained newborn monocytes demonstrated tolerization, as evidenced by significantly diminished subsequent LPS-induced TNF (RPMI vs. BCG 1:10, p <0.01), IL-10 and CCL5 production (p<0.05). With the exception of IL-1RA production by newborn monocytes, BCG-induced monocyte production of D1 cytokines/chemokines was inversely correlated with D7 LPS-induced TNF in both age groups (p<0.0001). Compared to BCG-trained adult monocytes, newborn monocytes demonstrated markedly impaired BCG-induced production of lactate, a metabolite implicated in immune training in adults. BCG-induced human monocyte primary- and memory-innate cytokine responses were age-dependent and accompanied by distinct immunometabolic shifts that impact both glycolysis and training. Our results suggest that immune ontogeny may shape innate responses to live attenuated vaccines, suggesting age-specific approaches to leverage innate training for broad protection against infection.

Angelidou A ，Diray-Arce J ，Conti MG ，Netea MG ，Blok BA ，Liu M ，Sanchez-Schmitz G ，Ozonoff A ，van Haren SD ，Levy O ... -  《-》

Circadian rhythm influences induction of trained immunity by BCG vaccination.

de Bree LCJ ，Mourits VP ，Koeken VA ，Moorlag SJ ，Janssen R ，Folkman L ，Barreca D ，Krausgruber T ，Fife-Gernedl V ，Novakovic B ，Arts RJ ，Dijkstra H ，Lemmers H ，Bock C ，Joosten LA ，van Crevel R ，Benn CS ，Netea MG ... -  《-》