Abstract
Background Hand, foot, and mouth disease (HFMD) is a common infectious disease among children under five years old, with large-scale outbreaks reported worldwide, leading to numerous severe cases and fatalities. Since 2008, HFMD has ranked first in both incidence and mortality among Class C notifiable infectious diseases in China. Enterovirus 71 (EV71) is the primary causative agent of severe HFMD, associated with critical cardiopulmonary injuries and life-threatening neurological complications. Clinical studies have shown that hyperglycemia is frequently observed in severe HFMD cases, and elevated fasting blood glucose is an independent risk factor for encephalitis complications. Macrophages, as key players in antiviral immunity, can become hyperactivated during infection, releasing excessive pro-inflammatory cytokines such as IL-1β and TNF-α, which may trigger a cytokine storm, exacerbating tissue damage and dysregulating glucose metabolism. This study aims to investigate the mechanisms by which inflammatory mediators contribute to glucose dysregulation, providing potential therapeutic strategies to reduce the risk of encephalitis in severe HFMD.
Methods We first performed intraperitoneal injection of EV71 virus into BALB/c mice with severe HFMD as mice injected with saline serving as the control group. We subsequently used immunohistochemistry (IHC) and ELISA to verify the pathological damage and functional changes in the pancreatic tissues of severe HFMD mice, and employed transcriptome analysis and quantitative real-time PCR (RT-PCR) to detect changes in genes related to insulin secretion in severe HFMD mice. Then we induced an inflammation model by treating mouse macrophage RAW cells with EV71, with untreated RAW cells serving as the control group. Finally, RT-PCR was performed to detect the secretion of inflammatory factors, and mouse pancreatic β-cells were treated with 0, 5, 10, and 20 ng/ml IL-1β and 0, 5, 10, and 20 ng/ml TNF-α, respectively, as well as flow cytometry and CCK8 were used to detect the proliferation and apoptosis of mouse pancreatic β-cells. Results Theelevated blood glucose levels, pancreatic tissue damage, and impaired islet secretion function (P< 0.05), in addition the pancreatic tissues infiltrated by a large number of macrophages secreting inflammatory factors (P<0.01) were observed and examined in severe HFMD mice. We explored and found the insulin secretion-related genes Atp1b4, Irs1, and Irs4 that downregulated in severe HFMD mice (P<0.001) , and pancreatic islet cells inflammation through macrophages in EV71 infected mouse (P < 0.0001).
Conclusion In this study, we used an EV71-infected severe HFMD mouse model to investigate hyperglycemia-related mechanisms. We observed significantly elevated fasting blood glucose levels accompanied by pancreatic tissue damage and impaired insulin secretion, despite minimal EV71 VP1 replication in the pancreas, suggesting indirect injury rather than direct viral infection. Transcriptomic analysis revealed downregulation of insulin secretion-related genes, supported by KEGG pathway and RT-PCR validation. Notably, pancreatic tissues exhibited macrophage infiltration and upregulated pro-inflammatory cytokines (IL-1β, IL-6, TNF-α, IFN-γ). In vitro, EV71 triggered inflammatory responses in macrophages, while IL-1β and TNF-α suppressed MIN6 β-cell proliferation and promoted apoptosis. Our findings demonstrate that EV71-induced macrophage activation drives pancreatic injury via cytokine storm, disrupting β-cell function and insulin secretion, ultimately leading to hyperglycemia. Further studies are needed to clarify the causal link between insulin deficiency and sustained hyperglycemia in severe HFMD. (Acknowledgements:This study was supported by Grant from School of Public Health of Southern Medical University, China , Grant No.GW202329; Corresponding author: Hong Cao, gzhcao@smu.edu.cn).
