p38 MAP Kinase Signaling in Microglia Plays a Sex-Specific Protective Role in CNS Autoimmunity and Regulates Microglial Transcriptional States

Abstract

Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system, representing the leading cause of non-traumatic neurologic disease in young adults. This disease is three times more common in women, yet more severe in men, but the mechanisms underlying these sex differences remain largely unknown. MS is initiated by autoreactive T helper cells, but CNS-resident and CNS-infiltrating myeloid cells are the key proximal effector cells regulating disease pathology. We have previously shown that genetic ablation of p38α MAP kinase broadly in the myeloid lineage is protective in the autoimmune model of MS, experimental autoimmune encephalomyelitis (EAE), but only in females, and not males. To precisely define the mechanisms responsible, we used multiple genetic approaches and bone marrow chimeras to ablate p38α in microglial cells, peripheral myeloid cells, or both. Deletion of p38α in both cell types recapitulated the previous sex difference, with reduced EAE severity in females. Unexpectedly, deletion of p38α in the periphery was protective in both sexes. In contrast, deletion of p38α in microglia exacerbated EAE in males only, revealing opposing roles of p38α in microglia vs. periphery. Bulk transcriptional profiling revealed that p38α regulated the expression of distinct gene modules in male vs. female microglia. Single-cell transcriptional analysis of WT and p38α-deficient microglia isolated from the inflamed CNS revealed a diversity of complex microglial states, connected by distinct convergent transcriptional trajectories. In males, microglial p38α deficiency resulted in enhanced transition from homeostatic to disease-associated microglial states, with the downregulation of regulatory genes such as Atf3, Rgs1, Socs3, and Btg2, and increased expression of inflammatory genes such as Cd74, Trem2, and MHC class I and II genes. In females, the effect of p38α deficiency was divergent, exhibiting a unique transcriptional profile that included an upregulation of tissue protective genes, and a small subset of inflammatory genes that were also upregulated in males. Taken together, these results reveal a p38α-dependent sex-specific molecular pathway in microglia that is protective in CNS autoimmunity in males, suggesting that autoimmunity in males and females is driven by distinct cellular and molecular pathways, thus suggesting design of future sex-specific therapeutic approaches.

Glucose Transporter-2 Regulation of Male versus Female Hypothalamic Astrocyte MAPK Expression and Activation: Impact of Glucose

“…Incorporation of a plan to investigate potential sex differences in GLUT2 regulation of hypothalamic astrocyte MAPK family protein expression and phosphorylation aligns with the current U.S. National Institutes of Health policy emphasis on consideration of sex as a critical biological variable. This focus is supported by reports that the MAPK profile is sex-dimorphic in several organs [22][23][24][25][26] and is sensitive to estradiol [27], and by recent findings that GLUT2 imposes sex-specific control of astrocyte glucose and glycogen metabolism [15]. The work performed herein used siRNA gene silencing tools along with immunoblotting to determine if and how GLUT2 gene knockdown may affect total versus phosphorylated protein expression profiles for p44/21 MAPK (ERK1/2), p38 MAPK, and SAPK/JNK molecular-weight isoforms in glucose-supplied versus glucose-deprived hypothalamic astrocytes from each sex.…”

Early chronic suppression of microglial p38α in a model of Alzheimer’s disease does not significantly alter amyloid-associated neuropathology

“…This also left the final sex ratios skewed across groups and underpowered to assess sex differences. This is not only an important caveat to consider in the context of disease progression [ 68 ], but also with respect to microglial p38 signaling, as some evidence has reported significant differences between male and female animals on measures of p38-mediated myeloid responses to Aβ as well as on the microglial transcriptome [ 69 ]. Finally, as AD presents with multiple pathologies and not only amyloid deposition [ 70 ], investigations of p38α in models that also present with other relevant neurodegenerative changes, such as those modeling Tau-associated pathologies [ 71 ], should be considered.…”

The multifaceted roles of activating transcription factor 3 (ATF3) in inflammatory responses – Potential target to regulate neuroinflammation in acute brain injury

“…101,102 Subsequently, the activation of cerebral resident neural and peripheral immune cells amplifies neuroinflammatory responses due to the compromised permeability of the blood brain barrier (BBB) and the proinflammatory cytokine storm. 103,104 Evidence shows that ATF3 can be expressed in microglia, 105,106 brain ECs, 67,107 and neurons 9,108 in the CNS and can exert effects on neuroinflammatory modulators to affect brain physiologic function or development. 31,32 Similarly, the expression of ATF3 in peripheral immune cells, such as macrophages, is involved in peripheral inflammation.…”

“…Evidence shows that ATF3 can be expressed in microglia, 105,106 brain ECs, 67,107 and neurons 9,108 in the CNS and can exert effects on neuroinflammatory modulators to affect brain physiologic function or development. 31,32 Similarly, the expression of ATF3 in peripheral immune cells, such as macrophages, is involved in peripheral inflammation.…”