HMEC-1

Moderate Levels of Haem can Relieve the Inhibition of Hyperoxia on HMEC-1 Proliferation

ROP is prevalent in premature infants due to enhanced perinatal care and neonatology, mainly caused by retinal angiogenesis stunted by relative hyperoxia, suppressing VEGF expression. While BACH1 inhibits VEGF, haem degrades BACH1, but its effect on endothelial cells in ROP is unknown.

Lan's team model phase I ROP's hyperoxia using HMEC-1 cells exposed to 40% hyperoxia. Various haem doses are tested on cell proliferation, migration, and angiogenesis. Under hyperoxic conditions, HMEC-1 cell proliferation is inhibited, but 5–40 μM haem enhances it, while 80 μM or more inhibits it. Under normoxic conditions, 10–40 μM haem boosts proliferation, whereas 160 μM or more inhibits it. The optimal dose for HMEC-1 proliferation under both conditions is 20 μM haem (Fig. 1a). Over time, proliferation of HMEC-1 peaks at 72 hours. The proliferation rate with 20 μM haem is higher than the control under both conditions, although no significant difference exists between the 20 μM haem hyperoxia and normoxia control groups, indicating that 20 μM haem alleviates hyperoxia-induced inhibition (Fig. 1b). The EdU assay shows green fluorescence in proliferating cells (Fig. 1c). Analysis reveals that under hyperoxic conditions, total cell number, proliferative cell number, and percentage decrease. Treatment with 20 μM haem increases these metrics under both hyperoxic and normoxic conditions. No significant difference is found between the 20 μM haem hyperoxia group and the normoxic control, confirming that 20 μM haem alleviates hyperoxia-induced proliferation inhibition (Fig. 1d-f).

Fig. 1. Effect of haem on HMEC-1 cell proliferation (Jian L, Mei Y, et al., 2021).

Doxorubicin Induces Senescence in HMEC-1 Cells

Aging impacts chronic disease development, affecting endothelial and platelet functions and involving cellular senescence. Senescent cells secrete bioactive molecules through SASP, potentially influencing platelet behavior. Venturini et al. investigated how SASP factors from senescent endothelial cells, induced by doxorubicin, affect platelet activity.

To generate senescent endothelial cells, HMEC-1 cells were treated with doxorubicin at concentrations of 0.05, 0.1, and 0.5 µM for 48 or 96 hours. Proliferation decreased at all three concentrations and further declined with longer exposure (Fig. 2A). SA-β-Gal assays confirmed increased senescence with higher doxorubicin doses (Fig. 2B-D). Even the lowest dose (0.05 µM) induced senescence, with higher doses compromising cell viability, especially at 96 hours (Fig. 2B). Thus, 0.05 µM doxorubicin was used for senescence induction in subsequent experiments. Senescent markers increased with time, peaking after 72 hours (Fig. 2D). CDKN1A mRNA levels also rose after 72 hours, supporting senescence induction (Fig. 2E).

Fig. 2. Analysis of proliferation and senescence in doxorubicin-treated HMEC-1 cells (Venturini W, Olate-Briones A, et al., 2020).