Enhancement of innate immunity in gingival epithelial cells by vitamin D and HDAC inhibitors.

Introduction: The human host defense peptide LL-37 is a component of the innate immune defense mechanisms of the oral cavity against colonization by microbes associated with periodontal disease. We have previously shown that the active form of vitamin D, 1,25(OH)2D3, can induce the expression of LL-37 in gingival epithelial cells (GEC), and prevent the invasion and growth of periopathogenic bacteria in these cells. Further, experimental vitamin D deficiency resulted in increased gingival inflammation and alveolar bone loss. Epidemiological studies have shown associations between vitamin D deficiency and periodontal disease in humans, suggesting application of vitamin D could be a useful therapeutic approach. Further, since we have shown the local activation of vitamin D by enzymes expressed in the GEC, we hypothesized that we could observe this enhancement with the stable, and inexpensive inactive form of vitamin D, which could be further increased with epigenetic regulators. Methods: We treated 3-dimensional primary cultures of GEC topically with the inactive form of vitamin D, in the presence and absence of selected histone deacetylase (HDAC) inhibitors. LL-37 mRNA levels were quantified by quantitative RT-PCR, and inhibition of invasion of bacteria was measured by fluorescence microscopy. Results: Vitamin D treatment led to an induction of LL-37 mRNA levels, as well as an inhibition of pro-inflammatory cytokine secretion. This effect was further enhanced by HDAC inhibitors, most strongly when the HDAC inhibitor, phenyl butyrate (PBA) was combined with Vitamin D3. This was observed both in solution and in a prototype gel formulation using sodium butyrate. Finally, this combination treatment led to an increase in the antimicrobial activity against infection by Porphyromonas gingivalis and Filifactor alocis, bacteria associated with periodontal lesions, as well as herpes simplex virus, which has also been shown to be associated with periodontal lesions. Conclusions: Our results demonstrate that a combination of inactive vitamin D and sodium butyrate could be developed as a safe treatment for periodontal disease.

Disease Course, Treatments, and Outcomes of Children With Systemic Juvenile Idiopathic Arthritis-Associated Lung Disease.

OBJECTIVE: Systemic juvenile idiopathic arthritis-associated lung disease (SJIA-LD) is a life-threatening disease complication. Key questions remain regarding clinical course and optimal treatment approaches. The objectives of the study were to detail management strategies after SJIA-LD detection, characterize overall disease courses, and measure long-term outcomes. METHODS: This was a prospective cohort study. Clinical data were abstracted from the electronic medical record, including current clinical status and changes since diagnosis. Serum biomarkers were determined and correlated with presence of LD. RESULTS: We enrolled 41 patients with SJIA-LD, 85% with at least one episode of macrophage activation syndrome and 41% with adverse reactions to a biologic. Although 93% of patients were alive at last follow-up (median 2.9 years), 37% progressed to requiring chronic oxygen or other ventilator support, and 65% of patients had abnormal overnight oximetry studies, which changed over time. Eighty-four percent of patients carried the HLA-DRB1*15 haplotype, significantly more than patients without LD. Patients with SJIA-LD also showed markedly elevated serum interleukin-18 (IL-18), variable C-X-C motif chemokine ligand 9 (CXCL9), and significantly elevated matrix metalloproteinase 7. Treatment strategies showed variable use of anti-IL-1/6 biologics and addition of other immunomodulatory treatments and lung-directed therapies. We found a broad range of current clinical status independent of time from diagnosis or continued biologic treatment. Multidomain measures of change showed imaging features were the least likely to improve with time. CONCLUSION: Patients with SJIA-LD had highly varied courses, with lower mortality than previously reported but frequent hypoxia and requirement for respiratory support. Treatment strategies were highly varied, highlighting an urgent need for focused clinical trials.

IFN-γ is essential for alveolar macrophage-driven pulmonary inflammation in macrophage activation syndrome.

Macrophage activation syndrome (MAS) is a life-threatening cytokine storm complicating systemic juvenile idiopathic arthritis (SJIA) driven by IFN-γ. SJIA and MAS are also associated with an unexplained emerging inflammatory lung disease (SJIA-LD), with our recent work supporting pulmonary activation of IFN-γ pathways pathologically linking SJIA-LD and MAS. Our objective was to mechanistically define the potentially novel observation of pulmonary inflammation in the TLR9 mouse model of MAS. In acute MAS, lungs exhibit mild but diffuse CD4-predominant, perivascular interstitial inflammation with elevated IFN-γ, IFN-induced chemokines, and alveolar macrophage (AMϕ) expression of IFN-γ-induced genes. Single-cell RNA sequencing confirmed IFN-driven transcriptional changes across lung cell types with myeloid expansion and detection of MAS-specific macrophage populations. Systemic MAS resolution was associated with increased AMϕ and interstitial lymphocytic infiltration. AMϕ transcriptomic analysis confirmed IFN-γ-induced proinflammatory polarization during acute MAS, which switches toward an antiinflammatory phenotype after systemic MAS resolution. Interestingly, recurrent MAS led to increased alveolar inflammation and lung injury, and it reset AMϕ polarization toward a proinflammatory state. Furthermore, in mice bearing macrophages insensitive to IFN-γ, both systemic features of MAS and pulmonary inflammation were attenuated. These findings demonstrate that experimental MAS induces IFN-γ-driven pulmonary inflammation replicating key features of SJIA-LD and provides a model system for testing potentially novel treatments directed toward SJIA-LD.