Small-molecule CBP/p300 histone acetyltransferase inhibition mobilizes leukocytes from the bone marrow via the endocrine stress response.

Mutations of the CBP/p300 histone acetyltransferase (HAT) domain can be linked to leukemic transformation in humans, suggestive of a checkpoint of leukocyte compartment sizes. Here, we examined the impact of reversible inhibition of this domain by the small-molecule A485. We found that A485 triggered acute and transient mobilization of leukocytes from the bone marrow into the blood. Leukocyte mobilization by A485 was equally potent as, but mechanistically distinct from, granulocyte colony-stimulating factor (G-CSF), which allowed for additive neutrophil mobilization when both compounds were combined. These effects were maintained in models of leukopenia and conferred augmented host defenses. Mechanistically, activation of the hypothalamus-pituitary-adrenal gland (HPA) axis by A485 relayed shifts in leukocyte distribution through corticotropin-releasing hormone receptor 1 (CRHR1) and adrenocorticotropic hormone (ACTH), but independently of glucocorticoids. Our findings identify a strategy for rapid expansion of the blood leukocyte compartment via a neuroendocrine loop, with implications for the treatment of human pathologies.

Lysophospholipid Acyltransferase 9 Promotes Emphysema Formation via Platelet-activating Factor.

Cigarette smoking is known to be the leading cause of chronic obstructive pulmonary disease (COPD). However, the detailed mechanisms have not been elucidated. PAF (platelet-activating factor), a potent inflammatory mediator, is involved in the pathogenesis of various respiratory diseases such as bronchial asthma and COPD. We focused on LPLAT9 (lysophospholipid acyltransferase 9), a biosynthetic enzyme of PAF, in the pathogenesis of COPD. LPLAT9 gene expression was observed in excised COPD lungs and single-cell RNA sequencing data of alveolar macrophages (AMs). LPLAT9 was predominant and upregulated in AMs, particularly monocyte-derived AMs, in patients with COPD. To identify the function of LPLAT9/PAF in AMs in the pathogenesis of COPD, we exposed systemic LPLAT9-knockout (LPALT9-/-) mice to cigarette smoke (CS). CS increased the number of AMs, especially the monocyte-derived fraction, which secreted MMP12 (matrix metalloprotease 12). Also, CS augmented LPLAT9 phosphorylation/activation on macrophages and, subsequently, PAF synthesis in the lung. The LPLAT9-/- mouse lung showed reduced PAF production after CS exposure. Intratracheal PAF administration accumulated AMs by increasing MCP1 (monocyte chemoattractant protein-1). After CS exposure, AM accumulation and subsequent pulmonary emphysema, a primary pathologic change of COPD, were reduced in LPALT9-/- mice compared with LPLAT9+/+ mice. Notably, these phenotypes were again worsened by LPLAT9+/+ bone marrow transplantation in LPALT9-/- mice. Thus, CS-induced LPLAT9 activation in monocyte-derived AMs aggravated pulmonary emphysema via PAF-induced further accumulation of AMs. These results suggest that PAF synthesized by LPLAT9 has an important role in the pathogenesis of COPD.

Multiomics identifies the link between intratumor steatosis and the exhausted tumor immune microenvironment in hepatocellular carcinoma.

BACKGROUND AND AIMS: Immunotherapy has become the standard-of-care treatment for hepatocellular carcinoma (HCC), but its efficacy remains limited. To identify immunotherapy-susceptible HCC, we profiled the molecular abnormalities and tumor immune microenvironment (TIME) of rapidly increasing nonviral HCC. APPROACHES AND RESULTS: We performed RNA-seq of tumor tissues in 113 patients with nonviral HCC and cancer genome sequencing of 69 genes with recurrent genetic alterations reported in HCC. Unsupervised hierarchical clustering classified nonviral HCCs into three molecular classes (Class I, II, III), which stratified patient prognosis. Class I, with the poorest prognosis, was associated with TP53 mutations, whereas class III, with the best prognosis, was associated with cadherin-associated protein beta 1 (CTNNB1) mutations. Thirty-eight percent of nonviral HCC was defined as an immune class characterized by a high frequency of intratumoral steatosis and a low frequency of CTNNB1 mutations. Steatotic HCC, which accounts for 23% of nonviral HCC cases, presented an immune-enriched but immune-exhausted TIME characterized by T cell exhaustion, M2 macrophage and cancer-associated fibroblast (CAF) infiltration, high PD-L1 expression, and TGF-ß signaling activation. Spatial transcriptome analysis suggested that M2 macrophages and CAFs may be in close proximity to exhausted CD8+ T cells in steatotic HCC. An in vitro study showed that palmitic acid-induced lipid accumulation in HCC cells upregulated PD-L1 expression and promoted immunosuppressive phenotypes of cocultured macrophages and fibroblasts. Patients with steatotic HCC, confirmed by chemical-shift MR imaging, had significantly longer PFS with combined immunotherapy using anti-PD-L1 and anti-VEGF antibodies. CONCLUSIONS: Multiomics stratified nonviral HCCs according to prognosis or TIME. We identified the link between intratumoral steatosis and immune-exhausted immunotherapy-susceptible TIME.

Ubiquitin Specific Protease 1 Expression and Function in T Cell Immunity.

T cells are essential mediators of immune responses against infectious diseases and provide long-lived protection from reinfection. The differentiation of naive to effector T cells and the subsequent differentiation and persistence of memory T cell populations in response to infection is a highly regulated process. E protein transcription factors and their inhibitors, Id proteins, are important regulators of both CD4+ and CD8+ T cell responses; however, their regulation at the protein level has not been explored. Recently, the deubiquitinase USP1 was shown to stabilize Id2 and modulate cellular differentiation in osteosarcomas. In this study, we investigated a role for Usp1 in posttranslational control of Id2 and Id3 in murine T cells. We show that Usp1 was upregulated in T cells following activation in vitro or following infection in vivo, and the extent of Usp1 expression correlated with the degree of T cell expansion. Usp1 directly interacted with Id2 and Id3 following T cell activation. However, Usp1 deficiency did not impact Id protein abundance in effector T cells or alter effector T cell expansion or differentiation following a primary infection. Usp1 deficiency resulted in a gradual loss of memory CD8+ T cells over time and reduced Id2 protein levels and proliferation of effector CD8+ T cell following reinfection. Together, these results identify Usp1 as a player in modulating recall responses at the protein level and highlight differences in regulation of T cell responses between primary and subsequent infection encounters. Finally, our observations reveal differential regulation of Id2/3 proteins between immune versus nonimmune cell types.

Mutagenesis and homology modeling reveal a predicted pocket of lysophosphatidylcholine acyltransferase 2 to catch Acyl-CoA.

Platelet-activating factor (PAF) is a potent proinflammatory phospholipid mediator that elicits various cellular functions and promotes several pathological events, including anaphylaxis and neuropathic pain. PAF is biosynthesized by two types of lyso-PAF acetyltransferases: lysophosphatidylcholine acyltransferase 1 (LPCAT1) and LPCAT2, which are constitutive and inducible forms of lyso-PAF acetyltransferase, respectively. Because LPCAT2 mainly produces PAF under inflammatory stimuli, understanding the structure of LPCAT2 is important for developing specific drugs against PAF-related inflammatory diseases. Although the structure of LPCAT2 has not been determined, the crystal structure was reported for Thermotoga maritima PlsC, an enzyme in the same gene family as LPCAT2. Here, we identified residues in mouse LPCAT2 essential for its enzymatic activity and a potential acyl-coenzyme A (CoA)-binding pocket, based on homology modeling of mouse LPCAT2 with PlsC. We also found that Ala115 of mouse LPCAT2 was important for acyl-CoA selectivity. In conclusion, these results predict the three-dimensional (3D) structure of mouse LPCAT2. Our findings have implications for the future development of new drugs against PAF-related diseases.

Influence of right versus left unilateral spatial neglect on the functional recovery after rehabilitation in sub-acute stroke patients.

Unilateral spatial neglect (USN) may lead to poor functional rehabilitation outcomes. However, studies investigating the rehabilitation outcomes of right-sided USN are lacking. We aimed to investigate (1) the clinical impacts of USN, including right-sided USN, for stroke patients in sub-acute rehabilitation, and (2) evaluate the differences in clinical characteristics and rehabilitation outcomes between right- and left-sided USN patients. We retrospectively screened the medical records of 297 inpatients at the Tokyo-Bay Rehabilitation Hospital who experienced a cerebrovascular accident with supratentorial lesions between January 1st, 2014 and December 31st, 2016. We performed independent multiple regression analysis in patients with left and right hemisphere damage. The Behavioral Inattention Test was a significant independent variable for predicting the motor, cognitive, and total functional independence measure (FIM), compared to the Stroke Impairment Assessment Set and Mini-Mental State Examination. USN affects motor FIM recovery more than cognitive FIM recovery regardless of the damaged hemisphere. Our study results confirm that both right- and left-sided USN influence the functional recovery of stroke patients. USN occurs, slightly less frequently, following a left hemisphere stroke. However, USN negatively affected rehabilitation outcomes, regardless of the neglected side. Therefore, USN treatment is necessary for patients with left and right hemisphere damage.

Biosynthetic Enzymes of Membrane Glycerophospholipid Diversity as Therapeutic Targets for Drug Development.

Biophysical properties of membranes are dependent on their glycerophospholipid compositions. Lysophospholipid acyltransferases (LPLATs) selectively incorporate fatty chains into lysophospholipids to affect the fatty acid composition of membrane glycerophospholipids. Lysophosphatidic acid acyltransferases (LPAATs) of the 1-acylglycerol-3-phosphate O-acyltransferase (AGPAT) family incorporate fatty chains into phosphatidic acid during the de novo glycerophospholipid synthesis in the Kennedy pathway. Other LPLATs of both the AGPAT and the membrane bound O-acyltransferase (MBOAT) families further modify the fatty chain compositions of membrane glycerophospholipids in the remodeling pathway known as the Lands' cycle. The LPLATs functioning in these pathways possess unique characteristics in terms of their biochemical activities, regulation of expressions, and functions in various biological contexts. Essential physiological functions for LPLATs have been revealed in studies using gene-deficient mice, and important roles for several enzymes are also indicated in human diseases where their mutation or dysregulation causes or contributes to the pathological condition. Now several LPLATs are emerging as attractive therapeutic targets, and further understanding of the mechanisms underlying their physiological and pathological roles will aid in the development of novel therapies to treat several diseases that involve altered glycerophospholipid metabolism.

Lysophosphatidylethanolamine acyltransferase 1/membrane-bound O-acyltransferase 1 regulates morphology and function of P19C6 cell-derived neurons.

Glycerophospholipids, which are components of biomembranes, are formed de novo by the Kennedy pathway and subsequently mature through the Lands cycle. Lysophospholipid acyltransferases (LPLATs) are key enzymes in both pathways and influence the fatty acid composition of biomembranes. Neuronal differentiation is characterized by neurite outgrowth, which requires biomembrane biosynthesis. However, the role of LPLATs in neuronal differentiation remains unknown. In this study, we examined whether LPLATs are involved in neuronal differentiation using all-trans-retinoic acid (ATRA)-treated P19C6 cells. In these cells, mRNA levels of lysophosphatidylethanolamine acyltransferase (LPEAT)-1/membrane-bound O-acyltransferase (MBOAT)-1 were higher than those in undifferentiated cells. LPEAT enzymatic activity increased with 16:0- and 18:1-CoA as acyl donors. When LPEAT1/MBOAT1 was knocked down with small interfering RNA (siRNA), outgrowth of neurites and expression of neuronal markers decreased in ATRA-treated P19C6 cells. Voltage-dependent calcium channel activity was also suppressed in these cells transfected with LPEAT1/MBOAT1 siRNA. These results suggest that LPEAT1/MBOAT1 plays an important role in neurite outgrowth and function.-Tabe, S., Hikiji, H., Ariyoshi, W., Hashidate-Yoshida, T., Shindou, H., Okinaga, T., Shimizu, T., Tominaga, K., Nishihara, T. Lysophosphatidylethanolamine acyltransferase 1/membrane-bound O-acyltransferase 1 regulates morphology and function of P19C6 cell-derived neurons.

Obese Japanese Patients with Stroke Have Higher Functional Recovery in Convalescent Rehabilitation Wards: A Retrospective Cohort Study.

BACKGROUND: A protective effect of excessive body mass index (BMI) on mortality or functional outcome in patients with stroke is not well established in the Asian population. This study aimed to explore whether obese patients with stroke have advantages for functional improvement in Japanese rehabilitation wards. METHOD: This retrospective cohort study included consecutive patients with stroke admitted and discharged from convalescent rehabilitation wards between 2011 and 2015. Demographic data, BMI, Functional Independence Measure (FIM) score, and nutritional status were analyzed. Participants were classified into 4 groups according to BMI (underweight <18.5 kg/m(2), standard 18.5-<23 kg/m(2), overweight 23-<27.5 kg/m(2), obese ≥27.5 kg/m(2)). The primary outcome was the FIM gain, and the secondary outcome was the FIM score at discharge. Multiple regression analysis was performed to analyze the relationship between BMI and functional recovery. RESULTS: In total, 897 participants (males 484, females 413; mean age 71.6 years) were analyzed and classified as underweight (134), standard (432), overweight (277), and obese (54). The median FIM gain and the FIM score at discharge were 30 and 114, respectively. The FIM gain in the obese group was significantly higher than those in the other groups. Multiple regression analysis revealed that obesity was independently correlated with the FIM gain, and those at discharge after adjusting for confounders such as age, gender, and FIM score on admission. CONCLUSIONS: Obese Japanese convalescent patients with stroke may have some advantages for functional recovery in rehabilitation wards.

Essential Role of Lysophosphatidylcholine Acyltransferase 3 in the Induction of Macrophage Polarization in PMA-Treated U937 Cells.

Lysophospholipid acyltransferases (LPLATs) regulate the diversification of fatty acid composition in biological membranes. Lysophosphatidylcholine acyltransferases (LPCATs) are members of the LPLATs that play a role in inflammatory responses. M1 macrophages differentiate in response to lipopolysaccharide (LPS) and are pro-inflammatory, whereas M2 macrophages, which differentiate in response to interleukin-4 (IL-4), are anti-inflammatory and involved in homeostasis and wound healing. In the present study, we showed that LPCATs play an important role in M1/M2-macrophage polarization. LPS changed the shape of PMA-treated U937 cells from rounded to spindle shaped and upregulated the mRNA and protein expression of the M1 macrophage markers CXCL10, TNF-α, and IL-1ß. IL-4 had no effect on the shape of PMA-treated U937 cells and upregulated the M2 macrophage markers CD206, IL-1ra, and TGF-ß in PMA-treated U937 cells. These results suggest that LPS and IL-4 promote the differentiation of PMA-treated U937 cells into M1- and M2-polarized macrophages, respectively. LPS significantly downregulated the mRNA expression of LPCAT3, one of four LPCAT isoforms, and suppressed its enzymatic activity toward linoleoyl-CoA and arachidonoyl-CoA in PMA-treated U937 cells. LPCAT3 knockdown induced a spindle-shaped morphology typical of M1-polarized macrophages, and increased the secretion of CXCL10 and decreased the levels of CD206 in IL-4-activated U937 cells. This indicates that knockdown of LPCAT3 shifts the differentiation of PMA-treated U937 cells to M1-polarized macrophages. Our findings suggest that LPCAT3 plays an important role in M1/M2-macrophage polarization, providing novel potential therapeutic targets for the regulation of immune and inflammatory disorders.

Macrophage/microglia-producing transient increase of platelet-activating factor is involved in neuropathic pain.

Peripheral nerve injury (PNI) induces debilitating neuropathic pain symptoms, such as tactile allodynia. Accumulating evidence suggests that the expression levels of various transcripts and proteins are drastically changed after PNI. Recent lipidome analysis demonstrates increased levels of diverse lipids in chronic pain conditions. We show that PNI transiently increases platelet-activating factor (PAF) levels, a potent inflammatory phospholipid mediator, in the dorsal root ganglia (DRG) and spinal cord. We revealed that macrophage and microglia-specific PAF-producing enzyme LPLAT9/LPCAT2 knockout mice (Cx3cr1CreERT2;Lpcat2flox/flox) failed to develop mechanical allodynia and to increase PAF levels in the DRG and spinal cord after PNI. Moreover, we observed the suppression of PNI-induced PAF increase in the spinal cord of PAF receptor knockout mice, indicating a self-amplification loop of PAF production. In conclusion, macrophages and microglia enhance PAF production, contributing to PNI-induced neuropathic pain. Additionally, PAF-PAF receptor signaling is a potential target of neuropathic pain control.

Japanese Patient Engagement Promotion Training (J-PEPT): Learning course on the implementation strategy of patient engagement.

Patient engagement for patient safety is emphasized in recent years. Therefore, the Committee on Quality and Patient Safety of the Japan Primary Care Association developed a Japanese Patient Engagement Promotion Training (J-PEPT) course. J-PEPT promotes to facilitate the implementation of PE strategies and contributes to nationwide dissemination for patient safety.

Sphingosine-1-phosphate promotes liver fibrosis in metabolic dysfunction-associated steatohepatitis.

AIM: Metabolic dysfunction-associated steatohepatitis (MASH) is one of the most prevalent liver diseases and is characterized by steatosis and the accumulation of bioactive lipids. This study aims to understand the specific lipid species responsible for the progression of liver fibrosis in MASH. METHODS: Changes in bioactive lipid levels were examined in the livers of MASH mice fed a choline-deficient diet (CDD). Additionally, sphingosine kinase (SphK)1 mRNA, which generates sphingosine 1 phosphate (S1P), was examined in the livers of patients with MASH. RESULTS: CDD induced MASH and liver fibrosis were accompanied by elevated levels of S1P and increased expression of SphK1 in capillarized liver sinusoidal endothelial cells (LSECs) in mice. SphK1 mRNA also increased in the livers of patients with MASH. Treatment of primary cultured mouse hepatic stellate cells (HSCs) with S1P stimulated their activation, which was mitigated by the S1P receptor (S1PR)2 inhibitor, JTE013. The inhibition of S1PR2 or its knockout in mice suppressed liver fibrosis without reducing steatosis or hepatocellular damage. CONCLUSION: S1P level is increased in MASH livers and contributes to liver fibrosis via S1PR2.

An immune-adrenergic pathway induces lethal levels of platelet-activating factor in mice.

Acute immune responses with excess production of cytokines, lipid/chemical mediators, or coagulation factors, often result in lethal damage. In addition, the innate immune system utilizes multiple types of receptors that recognize neurotransmitters as well as pathogen-associated molecular patterns, making immune responses complex and clinically unpredictable. We here report an innate immune and adrenergic link inducing lethal levels of platelet-activating factor. Injecting mice with toll-like receptor (TLR) 4 ligand lipopolysaccharide (LPS), cell wall N-glycans of Candida albicans, and the α2-adrenergic receptor (α2-AR) agonist medetomidine induces lethal damage. Knocking out the C-type lectin Dectin-2 prevents the lethal damage. In spleen, large amounts of platelet-activating factor (PAF) are detected, and knocking out lysophospholipid acyltransferase 9 (LPLAT9/LPCAT2), which encodes an enzyme that converts inactive lyso-PAF to active PAF, protects mice from the lethal damage. These results reveal a linkage/crosstalk between the nervous and the immune system, possibly inducing lethal levels of PAF.

hMLH1 promoter methylation status causes different expression patterns of estrogen receptor protein with endometrial lesion progression.

Mismatch repair (MMR) deficiency is reported to be an important factor in the early process of endometrial carcinogenesis. Although estrogen exposure is a crucial risk factor for endometrial carcinoma (EMCa), estrogen function is mediated by the estrogen receptor (ER). However, the relationship between ER and MMR, such as hMLH1 (human mutL homolog 1) activity, remains undetermined. In this study, we investigated the relationship between ER expression and hMLH1 promoter methylation status in atypical hyperplasia (AEH) and EMCa. ER expression was examined by immunohistochemical staining and the hMLH1 methylation status was evaluated using a methylation-specific PCR method. ER expression was significantly high in AEH, and extremely decreased with progression to EMCa. The hMLH1 methylation status allowed classification into methylated and unmethylated groups. Regarding the relationship between ER expression and hMLH1 methylation status, ER expression differed significantly between AEH and EMCa, and decreased with progression of the lesion in the unmethylated group, while it did not decrease with progression in the methylated group. These findings suggest that for a precursor lesion with hMLH1 unmethylated status, a decrease in ER expression is important for the development of carcinogenesis, while progression of a lesion with hMLH1 methylated status is not affected by ER expression.

Aromatic D-amino acids act as chemoattractant factors for human leukocytes through a G protein-coupled receptor, GPR109B.

GPR109B (HM74) is a putative G protein-coupled receptor (GPCR) whose cognate ligands have yet to be characterized. GPR109B shows a high degree of sequence similarity to GPR109A, another GPCR that was identified as a high-affinity nicotinic acid (niacin) receptor. However, the affinity of nicotinic acid to GPR109B is very low. In this study, we found that certain aromatic D-amino acids, including D-phenylalanine, D-tryptophan, and the metabolite of the latter, D-kynurenine, decreased the activity of adenylate cyclase in cells transfected with GPR109B cDNA through activation of pertussis toxin (PTX)-sensitive G proteins. These D-amino acids also elicited a transient rise of intracellular Ca(2+) level in cells expressing GPR109B in a PTX-sensitive manner. In contrast, these D-amino acids did not show any effects on cells expressing GPR109A. We found that the GPR109B mRNA is abundantly expressed in human neutrophils. D-phenylalanine and D-tryptophan induced a transient increase of intracellular Ca(2+) level and a reduction of cAMP levels in human neutrophils. Furthermore, knockdown of GPR109B by RNA interference inhibited the D-amino acids-induced decrease of cellular cAMP levels in human neutrophils. These D-amino acids induced chemotactic activity of freshly prepared human neutrophils. We also found that D-phenylalanine and D-tryptophan induced chemotactic responses in Jurkat cells transfected with the GPR109B cDNA but not in mock-transfected Jurkat cells. These results suggest that these aromatic D-amino acids elicit a chemotactic response in human neutrophils via activation of GPR109B.

Basic principles of neuroimmunology.

The brain is an immune-privileged organ such that immune cell infiltration is highly regulated and better tolerating the introduction of antigen to reduce risk of harmful inflammation. Thus, the composition and the nature of the immune response is fundamentally different in the brain where avoiding immunopathology is prioritized compared to other peripheral organs. While the principle of immune privilege in the central nervous system (CNS) still holds true, the role of the immune system in the CNS has been revisited over the recent years. This redefining of immune privilege in the brain is a result of the recent re-discovery of the extensive CNS meningeal lymphatic system and the identification of resident T cells in the brain, meningeal layers, and its surrounding cerebrospinal fluid (CSF) in both humans and rodents. While neuro-immune interactions have been classically studied in the context of neuroinflammatory disease, recent works have also elucidated unconventional roles of immune-derived cytokines in neurological function, highlighting the many implications and potential of neuro-immune interactions. As a result, the study of neuro-immune interactions is becoming increasingly important in understanding both CNS homeostasis and disease. Here, we review the anatomically distinct immune compartments within the brain, the known mechanisms of leukocyte trafficking and infiltration into the CNS and unique transcriptional and functional characteristics of CNS-resident immune cells.

Profiling of fatty acid metabolism in the dorsal root ganglion after peripheral nerve injury.

Peripheral nerve injury (PNI) induces neuronal hyperexcitability, which underlies neuropathic pain. The emergence of RNA sequencing technologies has enabled profiling of transcriptional changes in pathological conditions. However, these approaches do not provide information regarding metabolites such as lipids that are not directly encoded by genes. Fatty acids (FAs) are some of the essential lipids in mammalian organisms and are mainly stored as membrane phospholipids. In response to various biological stimuli, FAs are rapidly released and converted into several mediators, such as eicosanoids and docosanoids. FAs themselves or their metabolites play important roles in physiology and pathology. In this study, using a comprehensive lipidomic analysis of FA metabolites, 152 species were measured in the dorsal root ganglia of mice at multiple time points after PNI. We found that PNI increased the ω-6 FA metabolites produced by cyclooxygenases but not those produced by lipoxygenases or cytochrome P450 enzymes in the dorsal root ganglia. In contrast, ω-3 FA metabolites biosynthesized by any enzyme transiently increased after nerve injury. Overall, these findings provide a new resource and valuable insights into PNI pathologies, including pain and nerve regeneration.

A multiple sclerosis-protective coding variant reveals an essential role for HDAC7 in regulatory T cells.

Genome-wide association studies identifying hundreds of susceptibility loci for autoimmune diseases indicate that genes active in immune cells predominantly mediate risk. However, identification and functional characterization of causal variants remain challenging. Here, we focused on the immunomodulatory role of a protective variant of histone deacetylase 7 (HDAC7). This variant (rs148755202, HDAC7.p.R166H) was identified in a study of low-frequency coding variation in multiple sclerosis (MS). Through transcriptomic analyses, we demonstrate that wild-type HDAC7 regulates genes essential for the function of Foxp3+ regulatory T cells (Tregs), an immunosuppressive subset of CD4 T cells that is generally dysfunctional in patients with MS. Moreover, Treg-specific conditional hemizygous deletion of HDAC7 increased the severity of experimental autoimmune encephalitis (EAE), a mouse model of neuroinflammation. In contrast, Tregs transduced with the protective HDAC7 R166H variant exhibited higher suppressive capacity in an in vitro functional assay, mirroring phenotypes previously observed in patient samples. In vivo modeling of the human HDAC7 R166H variant by generation of a knock-in mouse model bearing an orthologous R150H substitution demonstrated decreased EAE severity linked to transcriptomic alterations of brain-infiltrating Tregs, as assessed by single-cell RNA sequencing. Our data suggest that dysregulation of epigenetic modifiers, a distinct molecular class associated with disease risk, may influence disease onset. Last, our approach provides a template for the translation of genetic susceptibility loci to detailed functional characterization, using in vitro and in vivo modeling.

Lysophosphatidylcholine Acyltransferase 1 Deficiency Promotes Pulmonary Emphysema via Apoptosis of Alveolar Epithelial Cells.

Chronic obstructive pulmonary disease (COPD) is primarily caused by inhalation of cigarette smoke and is the third leading cause of death worldwide. Pulmonary surfactant, a complex of phospholipids and proteins, plays an essential role in respiration by reducing the surface tension in the alveoli. Lysophosphatidylcholine acyltransferase 1 (LPCAT1) is an enzyme that catalyzes the biosynthesis of surfactant lipids and is expressed in type 2 alveolar epithelial cells. Its dysfunction is suggested to be involved in various lung diseases; however, the relationship between LPCAT1 and COPD remains unclear. To investigate the role of LPCAT1 in the pathology of COPD, we analyzed an elastase-induced emphysema model using Lpcat1 knockout (KO) mice. In Lpcat1 KO mice, elastase-induced emphysema was significantly exacerbated with increased apoptotic cells, which was not ameliorated by supplementation with dipalmitoylphosphatidylcholine, which is a major component of the surfactant synthesized by LPCAT1. We subsequently evaluated the effects of cigarette smoking on primary human type 2 alveolar epithelial cells (hAEC2s) and found that cigarette smoke extract (CSE) downregulated the expression of Lpcat1. Furthermore, RNA sequencing analysis revealed that the apoptosis pathway was significantly enriched in CSE-treated primary hAEC2s. Finally, we downregulated the expression of Lpcat1 using small interfering RNA, which resulted in enhanced CSE-induced apoptosis in A549 cells. Taken together, cigarette smoke-induced downregulation of LPCAT1 can promote the exacerbation of pulmonary emphysema by increasing the susceptibility of alveolar epithelial cells to apoptosis, thereby suggesting that Lpcat1 is a novel therapeutic target for irreversible emphysema.