ABSTRACT

Background Free testosterone (FT) demonstrates significant associations with cardiovascular diseases (CVDs). However, the precise relationship and underlying mediators linking FT and abdominal aortic calcification (AAC) remain incompletely understood. This study combined data from NHANES cross-sectional studies with Mendelian randomization (MR) and mediation MR analyses to assess the causal relationship between FT and AAC, and further identified lipid metabolites as potential mediators.

Methods Cross-sectional analysis utilized National Health and Nutrition Examination Survey (NHANES) data collected during 2013–2014, encompassing 2,654 individuals. Associations between severe AAC (SAAC) and FT were investigated using weighted multivariable regression, subgroup analyses, and restricted cubic spline (RCS) regression models. Moreover, to verify causal inference, bidirectional two-sample MR analyses were implemented utilizing European ancestry genome-wide association study (GWAS) data. Subsequently, mediation MR analyses were conducted to quantify the contribution of lipid metabolites to the observed associations.

Results A significant inverse association was observed between FT and SAAC; compared to participants in the lowest FT quartile, those in the highest quartile had a 67.9% lower risk (odds ratio [OR] = 0.321, 95% confidence interval [CI]: 0.194– 0.529, P value [P] < 0.001). The nonlinear inverse relationship between FT and SAAC was further confirmed by RCS regression (P for nonlinearity = 0.010). MR and mediation analyses corroborated the causal impact of FT on AAC (inverse variance-weighted [IVW] OR = 0.969, 95% CI: 0.941–0.997, P = 0.033). Four lipid-associated metabolites mediated approximately 36.13% of this causal link, namely mean low-density lipoprotein (LDL) particle diameter, triglycerides to total lipid ratio in large high-density lipoprotein (HDL) particles, total cholesterol to total lipid ratio in chylomicrons and extremely large very low-density lipoprotein (VLDL), and cholesteryl ester to total lipid ratio in chylomicrons and extremely large VLDL.

Conclusions Elevated FT independently associates with a reduced AAC risk and exerts causal effects mediated by lipid metabolic pathways. These findings highlight FT regulation as a potential target for AAC prevention and provide new insights for therapeutic strategies against vascular calcification (VC).

Competing Interest Statement

The authors have declared no competing interest.

Funding Statement

This research was supported by the following organizations: 1) National Natural Science Foundation of China (No. 82170517) for HD. Hu; 2) National Natural Science Foundation of China (No. 82170821) for Y. Qiu; 3) Basic Research Program from Department of Education of Liaoning Province (JC2019004); and 4) 345 Talent Project of Shengjing Hospital, China Medical University.

Author Declarations

I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.

Yes

The details of the IRB/oversight body that provided approval or exemption for the research described are given below:

All protocols used in the research were approved by the Ethics Committee at NCHS, with each participant providing informed consent. Publicly available summary statistics from GWAS datasets were utilized in MR analyses. Ethical approval and informed consent procedures had already been fulfilled by the original GWAS studies through their respective institutional review boards; hence, no further ethical approvals were required for the present analysis.

I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals.

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LIST OF ABBREVIATIONS(FT)Free testosterone(NHANES)National Health and Nutrition Examination Survey(CVDs)Cardiovascular diseases(AAC)Abdominal aortic calcification(MR)Mendelian randomization(SAAC)Severe abdominal aortic calcification(RCS)Restricted cubic spline(GWAS)Genome-wide association study[OR]Odds ratio[95% CI]95% confidence interval[P]P value(LDL)Low-density lipoprotein(HDL)High-density lipoprotein(VLDL)Very low-density lipoprotein(VC)Vascular calcification(CAD)Coronary artery disease(SHBG)Sex hormone-binding globulin(HDL-C)High-density lipoprotein cholesterol(SD)Standard deviations(DXA)Dual-energy X-ray absorptiometry(MEC)Mobile Examination Center(alb)Albumin(IVs)Instrumental variables(SNP)Single nucleotide polymorphism(NCHS)National Center for Health Statistics(IVW)Inverse variance weighted(ASO)Arteriosclerosis obliterans(VCAM-1)Vascular cell adhesion molecule-1(LOX-1)Lectin-type oxidized LDL receptor 1(LDLR)LDL receptors(TRLs)Triglyceride-rich lipoproteins