Obesity is a recognized risk factor for hypertension (1). Besides, hypertension constitutes one of the diagnostic criteria for metabolic syndrome, which is identified in approximately 40% of individuals who are obese (2). The incidence of hypertension among children with obesity can reach as high as 35% (3). The rise of childhood obesity has become a prominent public health concern, with a significant surge in its prevalence over recent decades. This condition transcends mere aesthetic issues, being linked to a multitude of serious health threats, including hypertension, which can have enduring implications for cardiovascular well-being.

Children who are overweight or obese are at an increased risk of developing hypertension in their adult years. The focus on preventative measures is crucial, especially for hypertension, where effective early diagnostics and biomarkers are sought. For pediatric hypertension, 24-hour ABPM is an effective diagnostic tool (4–6), and the innovative genetic methods like microarrays may help identify biomarkers.

The escalating prevalence of childhood obesity highlights the pressing necessity to unravel the intricate mechanisms that associate obesity with hypertension, as this insight could guide both preventive and therapeutic interventions. Although an array of genetic, environmental, and the lifestyle factors has been identified as contributors to the emergence of obesity in children, there remains a significant void in our comprehension of the precise molecular pathways that link obesity to the heightened susceptibility to hypertension in this at-risk demographic (7, 8).

Current investigations have revealed several genes linked to obesity; however, the specific contributions of the genes in facilitating the connection between obesity and hypertension remain inadequately examined. Among these, the S100A9 gene has attracted interest due to its role in inflammatory mechanisms and vascular impairment, both of which are important elements in the pathophysiology of hypertension. Prior research has underscored the importance of inflammatory indicators within the realm of obesity-related hypertension, yet the potential involvement of S100A9 in this dynamic is not comprehensively defined (9, 10). Consequently, there exists an urgent necessity for studies that explore the particular functions and regulatory pathways of S100A9 in relations to childhood obesity and its associated hypertension.

To address this research gap, our study employs a multifaceted approach that integrates gene expression analysis, WGCNA and functional assays. This combination of methods allows for a comprehensive exploration of gene expression profiles, co-expression networks, and identification of hub genes that may play pivotal roles in obesity-related hypertension. Such methodologies have proven advantageous in elucidating the complex biological processes and may facilitate the identification of novel therapeutic targets (11). The primary objective of this work is to elucidate the molecular mechanisms linking S100A9 to hypertension in children with obesity, thereby identifying key genes and pathways that could serve as potential intervention points.

Childhood obesity is a growing public health issue linked to serious health outcomes like hypertension in youth. Its origins are complex, involving genetic, environmental, and lifestyle factors, leading to increased prevalence. Children with obesity face immediate and long-term health challenges, including cardiovascular diseases, emphasizing the need for effective prevention and intervention.

This study explores how the S100A9 gene connects obesity and hypertension in children, identifying S100A9 as potential biomarker and mechanistic mediator. Findings show S100A9 affects the inflammation and vascular dysfunction, crucial for understanding obesity-related hypertension, and highlights its role in activating NADPH oxidase, increasing ROS, and decreasing NO, suggesting it as a target for treatment.

S100A9 is a calcium-binding protein, belonging to the damage-associated molecular pattern (DAMP) protein category, and is widely involved in inflammatory responses and fibrotic processes. In the context of childhood hypertension, the potential of S100A9 as a biomarker is mainly reflected in its association with inflammation and immune activation. Our study suggested that S100A9 is positively correlated with hypertension in obese children, which is consistent with the reports in the literature that S100A9 serves as an inflammatory marker in various diseases (20, 21). For instance, in cardiovascular diseases, S100A9 is identified as a key biomarker, involved in pathological processes such as inflammation, oxidative stress and fibrosis, which are also common in the pathogenesis of hypertension (21).

Compared with the existing biomarkers of childhood hypertension [C-reactive protein (CRP), renin, aldosterone], S100A9 may have unique advantages. Traditional biomarkers such as CRP are mainly used to assess systemic inflammation, but S100A9 is more specifically associated with the activation of neutrophils and macrophages (20, 22). For example, in obesity-related diseases, the level of S100A9 is associated with metabolic disorders and inflammatory states, while existing markers may lack sensitivity to early immune dysregulation. Specifically, S100A9 has shown association with hyperglycemia and insulin resistance in the diabetes and obesity models, while in children with hypertension, neutrophil activation as a major feature may make S100A9 more specific in identifying obesity-related hypertension subgroups (23, 24). In terms of clinical diagnostic utility, the S100A9 may achieve minimally invasive screening by testing serum or blood samples, and at the same time, it can be combined with other markers (such as BMI) to improve accuracy (21, 24).

The investigation reveals how childhood obesity affects immune functionality through neutrophil degranulation pathways, suggesting persistent inflammation leads to comorbidities like hypertension. Understanding S100A9 interactions may clarify obesity's immunological impact, aiding in developing immune modulators for better obesity management. Limitations include small sample size, lack of clinical validation, and potential batch effects, affecting the robustness of findings.

Paquinimod is an S100A9 inhibitor that has shown therapeutic potential in multiple studies, but its safety data in the pediatric population is limited. According to the literature, the safety and pharmacokinetics of Paquinimod have been confirmed in adult clinical trials (IPF and diabetes) (22, 23). For instance, in idiopathic pulmonary fibrosis model, Paquinimod improved the pathological changes of fibrosis and no serious adverse events were reported (22). Similarly, in retinopathy studies, Paquinimod inhibited S100A9-driven NLRP3 inflammasome activation in vitro (25). However, children's responses to therapeutic interventions may differ from those of adults, mainly because their developing immune systems and organs are more susceptible to interference. Potential developmental effects may include unknown effects on neural development or bone growth, as S100A9 plays a role in neuro-inflammation (26), but there is a lack of long-term follow-up data for pediatric populations. Subsequent clinical applications require a large amount of follow-up data from specific populations as a basis before conducting in vivo studies on children.

In terms of long-term consequences, S100A9 inhibition may regulate TLR4 pathway (20). Age-appropriate dosing strategies should be based on pediatric pharmacological principles, such as adjusting the dosage according to body weight or body surface area, similar to the application of other immunomodulators in children (22). Therefore, targeted clinical trials need to be conducted in children to evaluate safety of low-dose initiation and progressive administration (27). Overall, the safety of Paquinimod in adults provides a basis for pediatric application, but the individual differences and developmental stage-specific risks need to be carefully evaluated (21, 23).

Our research has found that neutrophil activation is the main immune feature in obese children with hypertension, which is consistent with the S100A9-mediated inflammatory pathway in the literature. S100A9 drives the inflammatory response by activating Toll-like receptor 4 (TLR4) and NLRP3 inflammosomes, which has been confirmed in various diseases (20, 22, 25). Among the existing mechanisms of pediatric hypertension, the traditional pathways mainly involve the activation of the renin-angiotensin system (RAS), sympathetic hyperactivity or endothelial dysfunction (21, 28). In contrast, the neutrophil activation pathway may represent a novel mechanism that more directly links obesity-related metabolic disorders and the development of hypertension. Specifically, in obesity models, upregulation of S100A9 leads to energy metabolism disorders and increased inflammation (24). Our research shows that this pathway is prominent in childhood hypertension, suggesting it may be particularly targeted at obesity-related hypertension, as S100A9 is highly expressed in both obese and diabetic states (23, 24). However, S100A9 also plays a role in non-obesity-related diseases (such as myocardial infarction), indicating that it may be involved in a broader range of inflammatory responses, not limited to obesity (11, 27). For instance, in cardiovascular diseases, S100A9 promotes fibrosis and inflammation, processes that are common in various subtypes of hypertension (21, 28). Therefore, the patient population for targeted S100A9 inhibition therapy may include children with obesity-related hypertension, but it may also extend to other cases of inflammation-driven hypertension (20, 27). This distinction is crucial for determining the treatment strategy: if neutrophil activation is only targeted at obesity, the treatment may focus on metabolic intervention; if the response is widespread, the S100A9 inhibitor (Paquinimod) may become a supplement to the current pediatric hypertension treatment, especially for patients who do not respond to traditional therapies. At present, the field of pediatric hypertension treatment lacks options targeting immune regulation. Therefore, approach discovered in this study may fill this gap by inhibiting S100A9 to alleviate the progression of inflammatory hypertension.

The sample size of the GSE87493 dataset used in this study is relatively limited, which is a key limitation of this research. The stability of module detection in WGCNA, the estimation of network topological parameters, and the identification of hub genes all largely depend on the sample size. A smaller sample size may increase statistical variation, resulting in less clear boundaries for certain co-expression modules or less stable rankings and connections for some hub genes. Although S100A9 showed relatively high module affiliation and connectivity in the analysis, its absolute strength of “hub” status in small sample setting may need to be interpreted with caution. It may represent the most prominent association signal in the current dataset, but its network centrality may slightly change in a larger sample. The smaller sample size may limit our ability to detect weak but biologically significant co-expression patterns, meaning that some genes or modules related to obesity-induced hypertension may not be captured by the current analysis.

To compensate for this deficiency, we do not merely rely on the results of WGCNA. The selected core regulatory genes all come from the intersection of DEGs and key modules, which enhances their biological relevance. More importantly, we conducted the independent functional experimental verification of S100A9 (in the endothelial cell function experiment), providing functional-level support for the key role of S100A9 in endothelial dysfunction. At the same time, we referred to existing literature in the discussion, pointing out the role of S100A9 in adult inflammation and cardiovascular diseases, which provides indirect biological rationality support for our findings in the pediatric population.

In this study, the HUVECs used as an in vitro primary endothelial cell model, although it is a classic tool for studying endothelial function, has inherent simplification. It cannot fully simulate the complexity of the vascular environment in children's bodies and lacks the interactions among various cell types (such as smooth muscle cells, immune cells) that exist in the body. Therefore, the effects of S100A9 on endothelial function observed in the HUVECs experiments and the protective effect of Paquinimod provide strong “principle verification” for mechanism hypothesis, but they are not directly equivalent to the therapeutic effects in the children with hypertension. Therefore, our findings require further experiments, including animal models and a large number of clinical studies, for validation.

Furthermore, one of the core early events related to childhood obesity-induced hypertension is vascular endothelial dysfunction. HUVECs are the most commonly used and mature models for studying the proliferation, migration, apoptosis, permeability, and nitric oxide (NO) signaling pathways of endothelial cells (29). Therefore, using it to investigate whether S100A9 directly impairs endothelial cell function is relatively appropriate. Meanwhile, although HUVECs are derived from umbilical cords, the basic endothelial cell biological characteristics they exhibit (forming tubular structures and responding to inflammatory factors) are highly conserved in vascular endothelial cells in children.

Therefore, based on the above research, we clearly state that the main value of this study lies in: (1) By combining bioinformatics with experiments, S100A9 has been identified as a potential key molecular node connecting childhood obesity and endothelial dysfunction; (2) The feasibility of targeting and inhibiting S100A9 (using Paquinimod) to improve endothelial function has been verified at the cellular level. This provides important preclinical evidence for “targeting S100A9 may become a therapeutic strategy”, but its ultimate clinical applicability depends on future evaluations of its efficacy, safety, and pediatric-specific pharmacokinetics in animal models and well-designed clinical trials.

In conclusion, this study reveals key molecular mechanisms and pathways linked to hypertension in children with obesity, emphasizing S100A9 as a critical mediator of vascular dysfunction and inflammation, and a potential biomarker and mechanistic mediator. It identifies important genes and interactions, suggesting future targeted interventions to reduce hypertension risk in this group, with further research (including a broader collection of clinical data and animal model experiments) needed to validate these findings and their implications.

In summary, this research aims to bridge the existing knowledge gap regarding the molecular mechanisms underlying childhood obesity and hypertension, with a specific focus on the role of S100A9. We seek to uncover the intricate relationships between gene expression, inflammatory responses, and vascular health in children with obesity via leveraging advanced analytical techniques, ultimately, the findings from this study may contribute to the development of targeted therapeutic strategies aimed at mitigating the risks of hypertension in this population.