Molecular Dissection of Pro-Fibrotic IL11 Signaling in Cardiac and Pulmonary Fibroblasts

Abstract

In fibroblasts, TGFβ1 stimulates IL11 upregulation that leads to an autocrine loop of IL11-dependent pro-fibrotic protein translation. The signaling pathways downstream of IL11, which acts via IL6ST, are contentious with both STAT3 and ERK implicated. Here we dissect IL11 signaling in fibroblasts and study IL11-dependent protein synthesis pathways in the context of approved anti-fibrotic drug mechanisms of action. We show that IL11-induced ERK activation drives fibrogenesis and while STAT3 phosphorylation (pSTAT3) is also seen, this appears unrelated to fibroblast activation. Ironically, recombinant human IL11, which has been used extensively in mouse experiments to infer STAT3 activity downstream of IL11, increases pSTAT3 in Il11ra1 null mouse fibroblasts. Unexpectedly, inhibition of STAT3 was found to induce severe proteotoxic ER stress, generalized fibroblast dysfunction and cell death. In contrast, inhibition of ERK prevented fibroblast activation in the absence of ER stress. IL11 stimulated an axis of ERK/mTOR/P70RSK protein translation and its selectivity for Collagen 1 synthesis was ascribed to an EPRS-regulated, ribosome stalling mechanism. Surprisingly, the anti-fibrotic drug nintedanib caused dose-dependent ER stress and lesser pSTAT3 expression. Pirfenidone had no effect on ER stress whereas anti-IL11 specifically inhibited the ERK/mTOR axis while reducing ER stress. These studies define the translation-specific signaling pathways downstream of IL11, intersect immune and metabolic signaling and reveal unappreciated effects of nintedanib.

IL11 stimulates ERK/P90RSK to inhibit LKB1/AMPK and activate mTOR in hepatocytes, the stroma and cancer cells

“…We have shown previously that TGFβ1 or IL11 stimulation of human cardiac fibroblasts over a 24 h time course causes disparate p-STAT3 profiles but common biphasic activation of ERK (Widjaja et al, 2021a). We examined whether this pattern was linked with phosphorylation of LKB1 (p-S428; putative inactive LKB1), AMPK (p-T172; active AMPK) or mTOR (S2448; active mTOR) ( Figure 1A and S1 ).…”

“…All cells were grown and maintained at 37°C and 5% CO 2 . The culture of primary human cardiac fibroblasts (HCFs), primary human hepatic stellate cells (HSCs), and primary human hepatocytes were described previously (Widjaja et al, 2019(Widjaja et al, , 2021a. Human lung epithelial carcinoma cell line (A549; CCL-185, ATCC) were grown and maintained in DMEM complete medium which contains 10% FBS (10500064, Gibco) and 1% P/S (15140122, Gibco).…”

“…ERK activation in the liver has previously been linked with diet-induced metabolic dysfunction (Jiao et al, 2013; Zheng et al, 2009b) and ERK is a important for activation of both fibroblasts (Du et al, 2008; Schafer et al, 2017) and hepatic stellate cells (HSCs) (Foglia et al, 2019; Widjaja et al, 2019). Intriguingly, IL11 was recently shown to activate rapamycin-sensitive ERK-dependent, mTOR-driven protein translation in fibroblasts (Widjaja et al, 2021a). This nascent discovery raises the question: could the IL11/ERK/mTORC1 axis be a shared mechanism for stromal cell activation and hepatocyte dysfunction and, if so, what is the full signalling pathway?…”

IL-11 Is Elevated and Drives the Profibrotic Phenotype Transition of Orbital Fibroblasts in Thyroid-Associated Ophthalmopathy

“…STAT3 activity has been implicated in proteotoxicity due to overloading of the endoplasmic reticulum. It has been demonstrated that Stattic prevented IL-11-induced fibrogenesis by inhibiting both ERK and STAT3 (53).…”

“…STAT3 activity has been implicated in proteotoxicity due to overloading of the endoplasmic reticulum. It has been demonstrated that Stattic prevented IL-11-induced fibrogenesis by inhibiting both ERK and STAT3 ( 53 ). The extent to which STAT3 and ERK interact under IL-11 stimulation in OFs is currently being investigated.…”