Tire Rubber Pollutant 6PPD-Quinone Linked to Erectile Dysfunction Through HIF-1α/eNOS/NO Axis

A chemical born from the wear and tear of everyday driving may be quietly undermining male sexual health. 6PPD-quinone, or 6PPD-Q, forms when the tire antioxidant 6PPD reacts with ozone in the atmosphere, and it has become one of the most closely watched emerging contaminants of the decade. Now, a research team from Beijing Anzhen Nanchong Hospital of Capital Medical University, Nanjing Drum Tower Hospital, and the Affiliated Hospital of Southwest Medical University has published evidence in Frontiers in Endocrinology that 6PPD-Q exposure can impair erectile function in rats by dysregulating the HIF-1α/eNOS/nitric oxide axis, a signaling pathway central to penile vascular physiology. The study, led by Chunyang Meng and colleagues, integrates an unusually broad methodological toolkit, spanning network toxicology, bulk and single-cell transcriptomics, machine learning, Mendelian randomization, molecular docking, molecular dynamics simulation, and in vivo animal experiments, to build a mechanistic case linking a ubiquitous traffic-derived pollutant to a condition that affects hundreds of millions of men worldwide.

The story of 6PPD-Q begins on the road. Every time a vehicle brakes, accelerates, or corners, its tires shed microscopic particles and a complex cocktail of chemicals into the surrounding environment. Among these chemicals, 6PPD-Q is generated by atmospheric oxidation of the tire antioxidant 6PPD and has attracted particular concern because of its extraordinary toxicity in sensitive aquatic species, most famously the acute mortality it induces in coho salmon. Environmental monitoring has since documented 6PPD-Q in air, dust, soil, road runoff, sediment, surface water, and even edible plants, creating multiple potential routes of human contact. Detection of the compound in human urine and cerebrospinal fluid indicates that environmental exposure translates into internal exposure, raising urgent questions about what such internal doses might be doing to human organs and tissues.

Previous research had already flagged the reproductive system as a potential target of 6PPD-Q. In ovarian granulosa cells, the compound has been reported to suppress PI3K/AKT/FOXO1 signaling and promote apoptosis. Chronic exposure in male mice disrupted endocrine balance, impaired sperm quality, increased germ-cell death, and altered sphingolipid signaling. An independent multi-omics study of non-obstructive azoospermia implicated oxidative injury, abnormal cell-cycle regulation, apoptosis, and loss of germ-cell homeostasis in 6PPD-Q-associated testicular damage. Growing evidence also links environmental pollution more broadly to male sexual dysfunction through oxidative stress, endocrine disruption, vascular endothelial injury, and impaired nitric oxide signaling, with occupational bisphenol A exposure, lead, arsenic, mixed heavy metals, and organophosphate metabolites all showing associations with erectile dysfunction. Yet whether 6PPD-Q directly impairs erectile function, and through what molecular mechanism, remained unclear until now.

To answer that question, the team first assembled the molecular landscape. They retrieved potential 6PPD-Q targets from SwissTargetPrediction, SEA, TargetNet, and the Comparative Toxicogenomics Database, and erectile dysfunction-associated genes from GeneCards. The intersection yielded 134 genes present in both collections, designated 6PPD-Q-ED-related genes. A protein-protein interaction network built in STRING and visualized in Cytoscape showed that these genes were mainly involved in responses to peptides and peptide hormones, responses to xenobiotic stimuli, and negative regulation of transport, while Gene Ontology and KEGG enrichment identified metabolism-, signaling-, and cancer-related pathways. The enrichment profile suggested that 6PPD-Q exposure might disrupt cellular homeostasis through mechanisms related to xenobiotic stress responses and altered transport regulation, providing the first hint of a biologically coherent link between the pollutant and cavernosal dysfunction.

The next step placed these candidate signals within the cellular architecture of the corpus cavernosum itself. Using single-cell RNA sequencing data from five human erectile tissue samples, the researchers resolved nineteen clusters and annotated six major cell types: fibroblasts, endothelial cells, smooth muscle cells, macrophages, Schwann cells, and T cells. Fibroblasts were the predominant population, accounting for 48.98 percent of all cells, followed by endothelial cells at 22.91 percent and smooth muscle cells at 22.49 percent. Gene set scoring with AUCell showed that the 6PPD-Q-ED-related gene set was relatively enriched in macrophages and moderately enriched in fibroblasts and endothelial cells, suggesting that 6PPD-Q exposure may disrupt the cavernosal microenvironment through inflammatory responses, extracellular matrix remodeling, and endothelial dysfunction, the very processes that govern penile vascular health.

To prioritize the leading candidates, the team integrated three rat corpus cavernosum transcriptomic datasets covering diabetes-associated, hypertension-associated, and neurogenic erectile dysfunction, batch-correcting the combined matrices with ComBat. Differential expression analysis identified 1,003 genes, seventeen of which overlapped with the 6PPD-Q-ED-related gene set. Two independent machine learning procedures, LASSO regression and support vector machine-recursive feature elimination, converged on four genes: ACO1, HIF1A, PDK4, and PTPN11. All four showed area under the receiver operating characteristic curve values greater than 0.70, with HIF1A reaching an AUC of 0.926. Critically, two-sample Mendelian randomization using cis-eQTL instruments from eQTLGen and erectile dysfunction summary statistics from FinnGen, comprising 1,154 cases and 94,024 controls, showed that genetically predicted HIF1A expression was significantly associated with erectile dysfunction risk, with an inverse-variance weighted odds ratio of 1.384 and a 95 percent confidence interval of 1.022 to 1.874. No significant associations were found for the other three genes, and sensitivity analyses including MR-Egger, weighted median, MR-PRESSO, and leave-one-out procedures supported the stability of the HIF1A result.

With HIF1A prioritized, the team turned to structural biology to ask whether 6PPD-Q could physically interact with HIF-1α, the protein it encodes. Molecular docking with AutoDock Vina predicted that 6PPD-Q could bind to a pocket in HIF-1α with a binding energy of −6.8 kcal/mol, below the −5 kcal/mol threshold conventionally interpreted as suggestive of potentially favorable binding. The docked complex also showed hydrogen-bond interactions between 6PPD-Q and nearby amino acid residues. A 100-nanosecond molecular dynamics simulation in GROMACS confirmed that the complex reached a relatively stable trajectory, with root-mean-square deviation rising during early structural accommodation and subsequently fluctuating around a plateau. Radius of gyration and solvent-accessible surface area decreased modestly before stabilizing, consistent with a more compact ensemble, and intermittent hydrogen bonds persisted throughout the trajectory. The free-energy landscape contained a dominant low-energy basin, providing a computational structural basis for the proposed interaction.

The decisive evidence came from the animal experiments. Twenty-four male Sprague-Dawley rats were allocated to four groups: control, 6PPD-Q, 6PPD-Q plus PX-478, and PX-478 alone. 6PPD-Q was suspended in corn oil and given by oral gavage at 4 mg/kg once daily for six weeks, while PX-478, a HIF-1α inhibitor, was administered orally at 10 mg/kg three times per week. After the exposure period, the researchers measured erectile hemodynamics by recording intracavernosal pressure in response to electrical stimulation of the pelvic ganglion, normalizing maximum intracavernosal pressure to mean arterial pressure as the functional endpoint. Rats exposed to 6PPD-Q showed a significantly lower ICPmax/MAP ratio than controls, indicating impaired erectile function. Co-treatment with PX-478 partially restored this ratio, providing pharmacological evidence that HIF-1α activation contributes to the erectile impairment.

The molecular changes in the corpus cavernosum told a coherent story. Immunohistochemistry showed increased HIF-1α staining in endothelial and smooth muscle cells of 6PPD-Q-exposed rats, which was reduced by PX-478 treatment. Western blot analysis revealed increased HIF-1α expression alongside decreased phosphorylated eNOS, decreased total eNOS, and a reduced p-eNOS/eNOS ratio in the 6PPD-Q group. Cavernosal nitric oxide levels, measured colorimetrically, were decreased following 6PPD-Q exposure and increased after PX-478 treatment. Notably, serum testosterone levels did not differ significantly among the four groups, suggesting that the erectile impairment in this model was attributable to local vascular dysfunction rather than altered androgen levels. The eNOS/NO pathway is the core mechanism responsible for maintaining normal erectile responses, as endothelial cell-derived nitric oxide promotes relaxation of corpus cavernosum smooth muscle and sinusoidal filling, and its disruption by 6PPD-Q points directly to endothelial injury as the site of toxic action.

The findings suggest that 6PPD-Q may contribute to erectile dysfunction by activating HIF1A-related stress responses and impairing eNOS/NO-mediated endothelial relaxation. HIF-1α is a central transcriptional regulator of cellular responses to hypoxia and stress, and excessive activation may enhance the release of inflammatory mediators and activate tissue remodeling signals, contributing to endothelial injury and smooth muscle dysfunction. Previous erectile dysfunction studies have shown that cavernosal nerve injury or hypoxic conditions are accompanied by increased HIF-1α expression, elevated fibrosis-related factors, and reduced NOS expression, while interventions that improve erectile function are often associated with HIF-1α downregulation and restoration of the NOS system. The authors acknowledge important limitations: only a single dose and exposure period were evaluated, precluding dose-response assessment; the transcriptomic datasets were derived from rat models; the Mendelian randomization analysis was constrained by available public data and tissue mismatch; downstream cGMP signaling was not evaluated; and the proposed 6PPD-Q-HIF-1α interaction lacked direct biochemical validation by surface plasmon resonance or co-immunoprecipitation. Nevertheless, this study extends the reproductive toxicological effects of 6PPD-Q to penile erectile function for the first time, adding a new dimension to concerns about traffic-derived contaminants and male sexual health, and suggesting that reducing environmental 6PPD-Q exposure could become a modifiable factor in preventing vascular erectile dysfunction.

Subject of Research: Mechanistic evidence linking the tire-derived environmental contaminant 6PPD-quinone to erectile dysfunction through dysregulation of the HIF-1α/eNOS/nitric oxide axis

Article Title: 6PPD-quinone exposure impairs erectile function through dysregulation of the HIF-1α/eNOS/NO axis: multi-omics and in vivo evidence

Article References: Meng, C., Li, Y., Zhu, X., Li, Y., Du, Y., & Xia, Z. (2026). 6PPD-quinone exposure impairs erectile function through dysregulation of the HIF-1α/eNOS/NO axis: multi-omics and in vivo evidence. Frontiers in Endocrinology, 17, Article 1912092. https://doi.org/10.3389/fendo.2026.1912092

Image Credits: AI Generated

DOI: 10.3389/fendo.2026.1912092

Keywords: 6PPD-quinone, erectile dysfunction, HIF-1α, eNOS, nitric oxide, environmental contaminant, tire rubber pollutant, corpus cavernosum, network toxicology, single-cell RNA sequencing, Mendelian randomization, molecular docking