Role of Sirt1 in innate immune mechanisms against Mycobacterium tuberculosis via the inhibition of TAK1 activation

Highlights

• Sirt1 expression was low in PBMCs from clinical TB samples and Mtb-infected murine peritoneal macrophages.

• Sirt1 interacted with TAK1 and inhibited its phosphorylation and ubiquitination.

• Sirt1^+/− mice were more susceptible to Mtb infection than WT mice.

• Resveratrol alleviated Mtb infection through anti-inflammatory mechanisms.

Abstract

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) has an alarming mortality rate, and the fast-growing drug resistance of Mtb necessitates the discovery and application of new therapeutic agents. Although Sirt1 is a key regulator in metabolic, cardiovascular and other age-related diseases, its role in the regulation of antibacterial immunity is unclear. Here, we have reported that Sirt1 expression was decreased, and the decreased Sirt1 level was correlated with increased pro-flammatory cytokines and TAK1 phosphorylation in active TB patients. Our mechanistic experiments showed that Sirt1 direclty interacted with TAK1 and suppressed its activation. Further, Sirt1^+/− macrophages infected with Mtb exhibited enhanced activation of TAK1, MAPKs and NF-κB, as well as simultaneously elevated expression of pro-inflammatory cytokines. Moreover, Sirt1^+/− mice exhibited overt inflammation, as indicated by the significant abundance of pro-inflammatory cytokines, and were more susceptible to Mtb infection than wild-type mice. Overall, the findings indicate that inhibition of Sirt1 expression by Mtb infection enhances TAK1 activation, and this in turn enhances the secretion of IL-6 and TNF-α via activation of the p65/p38/JNK/ERK signaling pathways. Treatment with resveratrol, which is known that one of its multiple effects is Sirt1 activation, in Mtb-infected macrophages inhibited the activation of TAK1, MAPK and NF-κB pathways, and the pro-inflammatory cytokine levels. Consitently, mice treated with resveratrol were more resistant to Mtb infection. The potent therapeutic effects of resveratrol against Mtb infection indicate that Sirt1 could be a novel therapeutic target for the treatment of TB.

Introduction

Tuberculosis (TB), which is one of the world's deadliest infectious diseases and is caused by Mycobacterium tuberculosis (Mtb), has become a global health issue, and the incidence of TB multidrug-resistant and extensively drug-resistant strains of Mtb has been on the increase [1,2]. Recently, host-directed therapies (HDTs) have emerged as a promising adjunct to standard therapy; these therapies typically target microbial cell autophagy, antimicrobial peptide production, limit the overt inflammation and pathological impairment that occur in the lung. The candidates for such interventions are biological agents or already approved drugs that can be re-purposed to interfere with biologically relevant cellular checkpoints [3,4]. A better understanding of the immunopathogenesis of TB and identification of the involved critical host factors could help in the establishment of novel HDTs that can be offered as an adjunct to the current treatment regimens.

The pro-inflammatory cytokines tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) play a protective role that includes the control of bacterial numbers and granuloma formation during TB infection [[5], [6], [7], [8]]. The biosynthesis of these cytokines involves the activation of the nuclear factor kappaB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling cascades, which include extracellular signal-regulated kinases (ERKs), c-Jun N-terminal kinases (JNKs) and p38 [[9], [10], [11]]. These signaling cascades are activated by transforming growth factor-β-activated kinase 1 (TAK1), which in turn is activated by auto-phosphorylation at key serine/threonine sites (Ser-192, Thr-178, Thr-187, and Thr-184) of the kinase activation loop and TAK1 complex K63 ubiquitination [[12], [13], [14], [15]]. At the start of this chain of activation are Toll-like receptors (TLRs), which can recognize a variety of molecular patterns, especially those of invading bacteria, and recruit various adaptor proteins that activate all the downstream signaling cascades that eventually lead to the synthesis of pro-inflammatory cytokines. Although activation of TLR-induced inflammatory response is essential for host defence against Mtb, overt activated inflammatory response may lead to tissue damage and TB progression. Therefore, a balanced inflammatory response is critical for the control of mycobacteria infection and resolution of the inflammation.

Sirtuins, or silent information regulator proteins, are a conserved family of seven proteins that regulate distinct biological pathways in mammals [16]. The sirtuin Sirt1, which is an NAD-dependent de-acetylase, plays crucial roles in aging, metabolism, apoptosis, inflammation and cell cycle regulation [17,18]. Considerable evidence has shown that Sirt1 plays a crucial role in inflammatory and other diseases, including metabolic, cardiovascular, age-related, and neuro-degenerative diseases, as well as cancer [[19], [20], [21], [22]]. However, it is unclear whether Sirt1 plays a role in the pathogenesis of TB. Therefore, one of the aims of the present study is to investigate the potential role of Sirt1 in TB, as well as the involvement of MAPK and NF-κB signaling pathways in the Sirt1-mediated pathogenesis of this disease, especially how to regulate NF-κB by unknown mechanism.

Resveratrol, a naturally occurring polyphenolic compound that is highly enriched in grapes, red wine, peanuts, and a wide variety of other food sources, is known to have anti-inflammatory, anti-oxidant, anti-viral, neuro-protective, chemo-preventive and chemotherapeutic properties [[23], [24], [25]]. Importantly, one of resveratrol's multiple effects is Sirt1 activation [26] and additionally is an inhibitor of NF-κB transcription [[27], [28], [29]]. Resveratrol has emerged in recent years as a compound that confers strong protection against metabolic, cardiovascular and other age-related complications, including neuro-degenerative diseases and cancer. Therefore, another aim of the present study was to determine whether resveratrol has therapeutic effects in TB.

The findings of the present study indicate that Sirt1 plays a role in the pathogenesis of TB. Further, the underlying mechanism seems to involve TAK1 activation and the subsequent production of inflammatory cytokines via the MAPK and NF-κB pathways. Finally, resveratrol is found to have therapeutic effects in vivo in infected mice.