Early life incision induces an epigenetic modification in macrophages to drive neonatal nociceptive priming

Abstract

Muscle injury evokes multiple biological responses including immune cell activation. Among these, macrophages are key players that regulate inflammation and nociception. They are also "trained" through pattern recognition and epigenetic modifications to facilitate future activity. Neonatal mechanisms of nociception are unique from that of adults but are less well studied. Basic and clinical studies agree that neonatal injury "primes" the individual, predisposing them to worse outcomes following subsequent injury. We, therefore, hypothesized that developing macrophages may uniquely modulate neonatal nociceptive priming through sustained epigenetic modifications. We performed surgical hindpaw incision in mice of both sexes at postnatal day 7 (P7) and/or P35 and observed spontaneous paw guarding and evoked muscle mechanical hypersensitivity. Macrophage fas-induced apoptosis (MaFIA) animals were used to ablate macrophages and adoptive transfer (AT) of neonatal macrophages were used to assess necessity and sufficiency. Isolated macrophages from reporter mice (LysM;tdTomato) were used for ATAC- and RNA-sequencing. Bone marrow stem cells were differentiated into macrophages (BMDMs) from our groups and assessed for pro-inflammatory responses. Neonatal incision induced both acute pain-like behaviors (guarding and mechanical hypersensitivity) and "primed" animals to a later in life injury at P35. Depletion of macrophages during neonatal incision partially blocked acute pain-like behaviors but prevented prolonged pain-related behaviors to re-injury. AT of "primed" macrophages resulted in more severe mechanical hypersensitivity compared to transfer of "naïve" macrophages. Sequencing experiments found that P7 injury drives differential chromatin accessibility and gene expression in P35 macrophages compared to naïve cells. BMDMs of "primed" macrophages demonstrated an enhanced pro-inflammatory response to LPS+IFNy stimulation compared to "naïve" cells. Together, these data indicate that early life injury alters macrophages which are necessary and partially sufficient to drive neonatal nociceptive priming possibly through epigenetic modifications. Results may provide insight into persistent pain development that may occur after surgical injury.