ObjectiveNon-suicidal self-injury (NSSI) is a common but maladaptive behavior among adolescents. Previous studies demonstrated a significant association of anxiety and depression with NSSI. However, the psychological and neuroendocrine mechanisms through which negative emotions influence NSSI remain unclear. The present study aimed to investigate whether the personality trait of neuroticism mediates the relationship between negative emotions and NSSI, and whether thyroid hormones moderate this pathway.MethodsA total of 104 Han Chinese adolescents and young adults (aged 12–22 years) who exhibited NSSI behaviors were recruited. The participants completed questionnaires to assess NSSI severity (Questionnaire for Middle School Students’ Behavior, QMSSB), depressive symptoms (Hamilton Depression Rating Scale, HAMD), anxiety symptoms (Hamilton Anxiety Rating Scale, HAMA) and personality traits (Eysenck Personality Questionnaire, EPQ). Venous blood was collected for thyroid function tests (TT3, TT4, FT3, FT4 and TSH). Mediation and moderation analyses were conducted using the PROCESS plug-in for SPSS software.ResultsNeuroticism mediated the relationship between emotional symptoms and NSSI. For HAMA in particular, neuroticism fully mediated the association with NSSI (effect = 0.0798, 95% bootstrap confidence interval [0.0387, 0.1367]). TT3 (b = 0.3112, p = 0.0147) and FT3 (b = 0.1118, p = 0.0371) positively moderated the HAMD-NSSI relationship, while TSH negatively moderated this relationship in the full sample (b = -0.0976, p = 0.0011) and remained significant in the high-neuroticism subgroup (β = -1.1116, p = 0.0026). Following FDR correction, TT3 (q = 0.0245), FT3 (q = 0.0412) and TSH (q = 0.0055) were found to significantly moderate the HAMD-NSSI relationship in the entire sample. The negative moderating effect of TSH remained significant in the high-neuroticism subgroup (q = 0.0078). However, no moderating effects in the low-neuroticism subgroup survived FDR correction (all q > 0.05). No significant moderating effects of thyroid hormones were found on the HAMA-NSSI pathway (all p > 0.05).ConclusionsNeuroticism mediates the mood-NSSI link. TT3 and FT3 positively moderate the depression-NSSI link, while TSH negatively moderates it. This negative moderating effect of TSH remains significant only in the high-neuroticism subgroup after FDR correction. These findings integrate psychological and neuroendocrine mechanisms for the identification and intervention of NSSI risk in youths.