Disruption of IL-10 receptor signaling promotes hematopoietic stem cell exhaustion and clonal expansion of JAK2V617F mutant cells during inflammatory stress Free

The ability of hematopoietic stem cells (HSCs) to return to quiescence after inflammatory activation is critical for long-term stem cell maintenance. While the regulation of HSC quiescence in steady-state conditions is well studied, much less is known about the signals that govern resolution of inflammatory activation. IL-10 is a key anti-inflammatory cytokine that limits excessive immune activation following Toll-like receptor (TLR) signaling. In monocytes, IL-10 acts through a negative feedback loop to suppress pro-inflammatory cytokine production. We asked whether IL-10 signaling plays a similar role in regulating inflammatory responses in HSCs.

We exposed mice to acute or chronic TLR4 stimulation with lipopolysaccharide (LPS), with or without IL-10 receptor (IL-10R) blockade, and evaluated HSC proliferation (BrdU, H2B-GFP), transcriptional changes (RNA-seq), polarity (image flow cytometry), and regenerative function (competitive transplantation). After a single dose of LPS, HSCs returned to quiescence by 48 hours. In contrast, IL-10R blockade prolonged HSC cycling, with a significant fraction continuing to proliferate 72 hours post-exposure. RNA-seq of HSCs from mice treated with LPS plus IL-10R blockade showed sustained activation of inflammatory and metabolic pathways (e.g., E2F, mTOR, type I IFN) and downregulation of genes involved in stem cell maintenance. In models of chronic inflammation, H2B-GFP label dilution revealed greater proliferative history in HSCs from mice treated with LPS + IL-10R blockade compared to LPS alone. Combination treated HSCs also exhibited reduced polarity, a feature associated with functional decline. In transplantation assays, bone marrow from LPS + IL-10R-treated mice showed impaired competitive transplantation ability.

Given that resistance to inflammatory stress is a hallmark of mutant clones in clonal hematopoiesis and myeloid malignancies, we asked whether IL-10R blockade might create selective pressure favoring inflammation-resistant HSCs. While common mutations such as TET2 and DNMT3A are selected under common inflammatory conditions encountered during aging, we hypothesized that more specific inflammatory contexts, such as defective IL-10R signaling, might favor clones like JAK2V617F. We previously observed defective IL-10R signaling in monocytes from JAK2V617F-positive MPN patients, and notably, this defect was also present in an unaffected identical twin, suggesting a possible inherited predisposition.

In Ba/F3 cells, Jak2V617F enhanced IL-10R signaling, supporting cell growth under limiting concentrations of IL-10 and resulted in cytokine independence with long latency after withdrawal of cytokines. In vivo, Jak2V617F knock-in HSCs exhibited elevated baseline proliferation but were resistant to the prolonged cycling triggered by IL-10R blockade. Unlike wild-type cells, Jak2V617F HSCs retained features of quiescence despite inflammatory challenge. In competitive repopulation, IL-10R blockade drove preferential expansion of Jak2V617F-mutant cells, which over time gave rise to a more aggressive MPN phenotype, including accentuated thrombocytosis, splenomegaly, and megakaryocytic hyperplasia.

These findings identify IL-10R signaling as a critical regulator of inflammatory resolution in HSCs. Disruption of this pathway prolongs inflammatory activation, accelerates HSC exhaustion, and creates a selective environment that promotes expansion of inflammation-resistant clones such as JAK2V617F. This work suggests that defects in anti-inflammatory signaling pathways may contribute to clonal evolution in early MPN and raises the possibility that enhancing IL-10R signaling could help preserve stem cell function in the setting of chronic inflammation.