RESUMEN
Fibroblasts acquire a proinflammatory phenotype in inflammatory bowel disease, but the factors driving this process and how fibroblasts contribute to mucosal immune responses are incompletely understood. TNF superfamily member 12 (TNFSF12, or TNF-like weak inducer of apoptosis [TWEAK]) has gained interest as a mediator of chronic inflammation. In this study, we explore its role as a driver of inflammatory responses in fibroblasts and its contribution to fibroblast-monocyte interaction using human primary colonic fibroblasts, THP-1 and primary monocytes. Recombinant human TWEAK induced the expression of cytokines, chemokines, and immune receptors in primary colonic fibroblasts. The TWEAK upregulated transcriptome shared 29% homology with a previously published transcriptional profile of inflammatory fibroblasts from ulcerative colitis. TWEAK elevated surface expression of activated fibroblast markers and adhesion molecules (podoplanin [PDPN], ICAM-1, and VCAM-1) and secretion of IL-6, CCL2, and CXCL10. In coculture, fibroblasts induced monocyte adhesion and secretion of CXCL1 and IL-8, and they promoted a CD14high/ICAM-1high phenotype in THP-1 cells, which was enhanced when fibroblasts were prestimulated with TWEAK. Primary monocytes in coculture with TWEAK-treated fibroblasts had altered surface expression of CD16 and triggering receptor expressed on myeloid cells-1 (TREM-1) as well as increased CXCL1 and CXCL10 secretion. Conversely, inhibition of the noncanonical NF-κB pathway on colonic fibroblasts with a NF-κB-inducing kinase small molecule inhibitor impaired their ability to induce a CD14high phenotype on monocytes. Our results indicate that TWEAK promotes an inflammatory fibroblast-monocyte crosstalk that may be amenable for therapeutic intervention.
Asunto(s)
Diferenciación Celular , Colon , Citocina TWEAK , Fibroblastos , Monocitos , Humanos , Citocina TWEAK/metabolismo , Monocitos/inmunología , Monocitos/metabolismo , Fibroblastos/metabolismo , Fibroblastos/inmunología , Colon/inmunología , Colon/patología , Colon/metabolismo , Diferenciación Celular/inmunología , Comunicación Celular/inmunología , Inflamación/inmunología , Células THP-1 , Técnicas de Cocultivo , Citocinas/metabolismo , Adhesión CelularRESUMEN
BACKGROUND AND AIMS: Activation of vascular smooth muscle cell inflammation is recognised as an important early driver of vascular disease. We have previously identified the let-7 miRNA family as important regulators of inflammation in in vitro and in vivo models of atherosclerosis. Here we investigated a dual statin/let-7d-5p miRNA combination therapy approach to target human aortic SMC (HAoSMC) activation and inflammation. METHODS: In vitro studies using primary HAoSMCs were performed to investigate the effects of let-7d-5p miRNA overexpression and inhibition. HAoSMCs were treated with combinations of the inflammatory cytokine tumor necrosis factor-α (TNF-α), and atorvastatin or lovastatin. HAoSMC Bulk RNA-seq transcriptomics of HAoSMCs revealed downstream regulatory networks modulated by let-7d-5p miRNA overexpression and statins. Proteome profiler cytokine array, Western blotting and quantitative PCR analyses were performed on HAoSMCs to validate key findings. RESULTS: Let-7d-5p overexpression significantly attenuated TNF-α-induced upregulation of IL-6, ICAM1, VCAM1, CCL2, CD68, MYOCD gene expression in HAoSMCs (p<0.05). Statins (atorvastatin, lovastatin) significantly attenuated inflammatory gene expression and upregulated Let-7d levels in HAoSMCs (p<0.05). Bulk RNA-seq analysis of a dual Let-7d-5p overexpression/statin therapy in HAoSMCs revealed that let-7d-5p activation and statins converge on key inflammatory pathways (IL-6, IL-1ß, TNF-α, IFN-γ). Let-7d-5p overexpression led to reduced expression of the ox-LDL receptor OLR1, and this was associated with lower ox-LDL uptake in HAoSMCs. In silico analysis of smooth muscle cell phenotypic switching shows that overexpression of let-7d-5p in HAoSMCs maintains a contractile phenotype. CONCLUSIONS: Targeting the Let-7 network alongside statins can modulate HAoSMC activation and attenuate key inflammatory pathway signals.
Asunto(s)
Aorta , Atorvastatina , Inhibidores de Hidroximetilglutaril-CoA Reductasas , MicroARNs , Músculo Liso Vascular , Miocitos del Músculo Liso , Fenotipo , Transducción de Señal , MicroARNs/metabolismo , MicroARNs/genética , Humanos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/metabolismo , Miocitos del Músculo Liso/efectos de los fármacos , Aorta/metabolismo , Aorta/patología , Atorvastatina/farmacología , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Células Cultivadas , Factor de Necrosis Tumoral alfa/metabolismo , Inflamación/metabolismo , Inflamación/genética , Lovastatina/farmacología , Mediadores de Inflamación/metabolismo , Regulación de la Expresión Génica , Citocinas/metabolismoRESUMEN
OBJECTIVES: To understand commonalities and differences in injured patient experiences of accessing and receiving quality injury care across three lower-income and middle-income countries. DESIGN: A qualitative interview study. The interviews were audiorecorded, transcribed and thematically analysed. SETTING: Urban and rural settings in Ghana, South Africa and Rwanda. PARTICIPANTS: 59 patients with musculoskeletal injuries. RESULTS: We found five common barriers and six common facilitators to injured patient experiences of accessing and receiving high-quality injury care. The barriers encompassed issues such as service and treatment availability, transportation challenges, apathetic care, individual financial scarcity and inadequate health insurance coverage, alongside low health literacy and information provision. Facilitators included effective information giving and informed consent practices, access to health insurance, improved health literacy, empathetic and responsive care, comprehensive multidisciplinary management and discharge planning, as well as both informal and formal transportation options including ambulance services. These barriers and facilitators were prevalent and shared across at least two countries but demonstrated intercountry and intracountry (between urbanity and rurality) variation in thematic frequency. CONCLUSION: There are universal factors influencing patient experiences of accessing and receiving care, independent of the context or healthcare system. It is important to recognise and understand these barriers and facilitators to inform policy decisions and develop transferable interventions aimed at enhancing the quality of injury care in sub-Saharan African nations.