BACKGROUND: Irregular bleeding affects up to 60% of hormone replacement therapy (HRT) users. The mechanism of this bleeding is not understood. Reduced endometrial microvascular integrity appears to underlie breakthrough bleeding in pre‐menopausal women and the aim of this study was to establish whether similar changes are seen in HRT users and hence to elucidate a possible mechanism of irregular bleeding. METHODS: Endometrium from 34 HRT users with amenorrhoea, irregular bleeding or regular bleeding was assessed for endometrial endothelial cell density (anti‐CD34), number of blood vessels per mm², vascular basal lamina components (laminin, collagen IV and heparan sulphate proteoglycan) and in 32 subjects and 23 controls for perivascular smooth muscle α (SMA). Findings were compared with a control population of 29 post‐menopausal women not using HRT, other sex steroids or tamoxifen and with no vaginal bleeding. Staining intensity was assessed in a blinded fashion in all immunohistochemical studies. RESULTS: Four significant differences in endometrial blood vessels were observed between HRT users and controls: (i) a significantly lower density of endometrial endothelial cells (EC staining for CD34) per mm² was present in HRT users compared with controls (P < 0.001); (ii) endothelial cells (EC) were predominantly organized within blood vessels (83%) in controls but in HRT users EC were dispersed in the tissues with only 29% in organized vessels (P <0.001); (iii) supportive perivascular cell SMA was significantly reduced in 23 post‐menopausal HRT users compared with 23 post‐menopausal controls (n = 29, P = 0.013) and (iv) an atrophic or inactive histological pattern of endometrium was more frequently seen in the controls (P < 0.001). CONCLUSIONS: These findings support the hypothesis that exposure to HRT profoundly alters endometrial blood vessels, reducing structural integrity thereby predisposing to irregular bleeding in HRT users.