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other·ophthalmology, dry eye, ocular immunology, nutrition, animal science·PMC10001612

Lacrimal Gland Epithelial Cells Shape Immune Responses through the Modulation of Inflammasomes and Lipid Metabolism

International Journal of Molecular Sciences · 9 authors, 4 centres

AI SUMMARY

FIDELITY 76%

POPULATIONMouse models: R26^ASC-citrine^ reporter mice, C57BL/6J wild-type, TSP-1^-/-, NOD.H2^b^ (Sjögren's syndrome model), and BALBc control mice

INTERVENTIONIntraglandular injection of LPS/nigericin (bacterial mimic) or IL-1α (acute injury); chronic inflammation studied in NOD.H2^b^ and TSP-1^-/- mice

COMPARISONVehicle-injected or uninjected controls; healthy BALBc or TSP-1^WT^ mice

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TL;DR

This study demonstrates that lacrimal gland epithelial cells form inflammasomes in response to both acute and chronic inflammation, and that sustained inflammasome activation combined with altered lipid metabolism (reduced fatty acid synthesis and increased cholesterol metabolism) drives Sjögren's syndrome-like pathogenesis in mouse models. These findings suggest that targeting inflammasome pathways and restoring lipid homeostasis could represent new therapeutic strategies for dry eye disease.

Full summary

4,165 CHARS

**Background:** Dry eye disease, particularly aqueous-deficient dry eye, is often caused by chronic inflammation of the lacrimal gland (LG) in conditions like Sjögren's syndrome (SS). Inflammasomes are intracellular complexes that activate inflammatory cytokines IL-1β and IL-18 and can induce pyroptosis. While aberrant inflammasome activation is implicated in autoimmune diseases, its role in LG inflammation and the potential involvement of lipid metabolism in resolving inflammation were not well understood. This study aimed to characterize inflammasome activation during acute and chronic LG inflammation and identify metabolic pathways that may regulate the resolution of inflammation.

**Methods:** The authors used multiple mouse models: (1) R26^ASC-citrine^ reporter mice to visualize inflammasome formation via fluorescent ASC specks; (2) acute injury induced by intraglandular injection of IL-1α; (3) bacterial mimicry using LPS/nigericin injection; (4) chronic inflammation models including NOD.H2^b^ mice (a primary SS model) and TSP-1^-/- mice. Inflammasome activation was assessed by immunostaining for ASC specks, Western blotting for CASP1, IL-1β, and GSDMD cleavage, and qRT-PCR. RNA-seq data from acute injury (GSE99093) and chronic inflammation (GSE210332) were analyzed for differentially expressed genes (DEGs) and pathway enrichment using Metascape and ROSALIND software. Lipid metabolism genes were specifically examined during the resolution phase (day 3 post-injury) and in chronically inflamed LGs.

**Key Results:** - In R26^ASC-citrine^ mice, LPS/nigericin injection induced numerous ASC specks in LG epithelial cells (acinar cells, myoepithelial cells, and ducts) within 6 hours. Similarly, IL-1α injection led to ASC speck formation at 6 hours, with increased speck number and diameter by 12 hours, accompanied by nuclear fragmentation indicative of cell death. - In chronically inflamed NOD.H2^b^ LGs (6-month-old males), Western blotting showed increased cleaved CASP1 (p33), mature IL-1β (17 kDa), and a ~5-fold increase in cleaved GSDMD compared to BALBc controls. TSP-1^-/-:R26^ASC-citrine^ mice also showed significantly more ASC specks at 2 and 6 months, with numbers increasing with disease progression. - RNA-seq analysis of acute injury revealed upregulation of inflammasome sensors Nlrp3, Aim2, Ifi204, Mefv, Naip5, and caspases Casp1 and Casp4 on days 1-2, returning to basal levels by day 3. In chronic NOD.H2^b^ LGs, these same sensors plus Nlrc4 were upregulated from 2 months of age, with further increases at 4 months. Il1b and Il18 were significantly elevated in both models. - Pathway analysis of day 3 post-injury (resolution phase) identified 66 genes specifically upregulated, with the most significant cluster being "Cholesterol metabolism with Bloch and Kandutsch-Russell pathways" (18 genes). Key lipogenic genes included Srebf1 (encoding SREBP-1), Acacb, Sqle, and Acss2, peaking on day 3. - In contrast, chronically inflamed NOD.H2^b^ LGs showed downregulation of PPARα (Ppara), Srebf1, and genes for fatty acid biosynthesis (Acacb) and mitochondrial β-oxidation, while cholesterol biosynthesis genes (Hmgcr, Sqle, Soat1/2) were upregulated. These changes were evident as early as 2 months of age.

**Clinical Implications:** This study provides direct evidence that LG epithelial cells act as sentinel cells by forming inflammasomes in response to microbial and sterile stimuli, and that sustained inflammasome activation contributes to chronic inflammation in SS models. The discovery that lipid metabolism shifts from fatty acid synthesis (promoting resolution) to cholesterol accumulation (promoting inflammation) during chronic disease suggests a metabolic mechanism driving LG dysfunction. The authors propose that PPARα activators like fenofibrate, which can restore fatty acid β-oxidation and reduce inflammation, combined with anti-inflammatory agents targeting IL-1 or GSDMD, may offer new therapeutic strategies for SS-associated dry eye disease. These findings highlight the importance of epithelial cell metabolism in regulating immune responses and tissue regeneration in the lacrimal gland.

PICO

PPOPULATION

Mouse models: R26^ASC-citrine^ reporter mice, C57BL/6J wild-type, TSP-1^-/-, NOD.H2^b^ (Sjögren's syndrome model), and BALBc control mice

IINTERVENTION

Intraglandular injection of LPS/nigericin (bacterial mimic) or IL-1α (acute injury); chronic inflammation studied in NOD.H2^b^ and TSP-1^-/- mice

OOUTCOME

Inflammasome formation (ASC specks), expression of inflammasome-related genes (Nlrp3, Aim2, Ifi204, Mefv, Naip5, Nlrc4, Casp1, Casp4, Il1b, Il18), IL-1β maturation, GSDMD cleavage, lipid metabolism gene expression (Srebf1, Ppara, Acacb, Hmgcr, Sqle), and cell death

STUDY TYPE

other

SPECIALTY

ophthalmology

SUMMARISED BY

AI pipeline

FIDELITY CHECK

76% · B

ORIGINAL PAPER

PMC10001612

Lacrimal Gland Epithelial Cells Shape Immune Responses through the Modulation of Inflammasomes and Lipid Metabolism

International Journal of Molecular Sciences

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ABSTRACT

Lacrimal gland inflammation triggers dry eye disease through impaired tear secretion by the epithelium. As aberrant inflammasome activation occurs in autoimmune disorders including Sjögren’s syndrome, we analyzed the inflammasome pathway during acute and chronic inflammation and investigated its potential regulators. Bacterial infection was mimicked by the intraglandular injection of lipopolysaccharide (LPS) and nigericin, known to activate the NLRP3 inflammasome. Acute injury of the lacrimal gland was induced by interleukin (IL)-1α injection. Chronic inflammation was studied using two Sjögren’s syndrome models: diseased NOD.H2 b compared to healthy BALBc mice and Thrombospondin-1-null (TSP-1 -/- ) compared to TSP-1 WT C57BL/6J mice. Inflammasome activation was investigated by immunostaining using the R26 ASC-citrine reporter mouse, by Western blotting, and by RNAseq. LPS/Nigericin, IL-1α and chronic inflammation induced inflammasomes in lacrimal gland epithelial cells. Acute and chronic inflammation of the lacrimal gland upregulated multiple inflammasome sensors, caspases 1/4, and interleukins Il1b and Il18 . We also found increased IL-1β maturation in Sjögren’s syndrome models compared with healthy control lacrimal glands. Using RNA-seq data of regenerating lacrimal glands, we found that lipogenic genes were upregulated during the resolution of inflammation following acute injury. In chronically inflamed NOD.H2 b lacrimal glands, an altered lipid metabolism was associated with disease progression: genes for cholesterol metabolism were upregulated, while genes involved in mitochondrial metabolism and fatty acid synthesis were downregulated, including peroxisome proliferator-activated receptor alpha (PPARα)/sterol regulatory element-binding 1 (SREBP-1)-dependent signaling. We conclude that epithelial cells can promote immune responses by forming inflammasomes, and that sustained inflammasome activation, together with an altered lipid metabolism, are key players of Sjögren’s syndrome-like pathogenesis in the NOD.H2 b mouse lacrimal gland by promoting epithelial dysfunction and inflammation.

CITATION EXPORT

Vanessa Delcroix, Olivier Mauduit, Menglu Yang et al.. Lacrimal Gland Epithelial Cells Shape Immune Responses through the Modulation of Inflammasomes and Lipid Metabolism. International Journal of Molecular Sciences. (2023). https://doi.org/10.3390/ijms24054309

AUTHORS

9 · 4 CENTRES

Vanessa Delcroix, Olivier Mauduit, Menglu Yang, Amrita Srivastava, Takeshi Umazume, Cintia S. de Paiva, Valery I. Shestopalov, Darlene A. Dartt, Helen P. Makarenkova

1Department of Molecular Medicine, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA
2Schepens Eye Research Institute/Massachusetts Eye and Ear, Department of Ophthalmology, Harvard Medical School, Boston, MA 02114, USA
3The Ocular Surface Center, Cullen Eye Institute, Department of Ophthalmology, Baylor College of Medicine, Houston, TX 77030, USA
4Bascom Palmer Eye Institute, University of Miami School of Medicine, Miami, FL 33136, USA

EDITOR / REVIEWER

Mariarosaria Boccellino