Cells, Cultured ; Curcumin ; pharmacology ; Endothelium, Vascular ; drug effects ; pathology ; radiation effects ; Human Umbilical Vein Endothelial Cells ; drug effects ; radiation effects ; Humans ; Radiation Injuries ; Signal Transduction ; drug effects ; radiation effects

The present paper is aimed to investigate the effects of pulse electrical stimulation on mutual adhesion of vascular endothelial cell and endothelial progenitor cell (EPC). EPC was induced from periphery blood, labeled with fluorescence dye and then co-cultured with vascular endothelial cell. With a fixed electric voltage and frequency of 5V and 5Hz, respectively, the co-culture system was continually stimulated for 24h under different pulse width, 1, 3, 6 and 9ms. After pulse stimulation, fluorescence intensity of adherent labeled EPC was measured and converted to fluorescence ratio. Compared to that in the control group, fluorescence ratio of 3 ms and 6 ms group were significantly larger, while that in the 9 ms group was lower. The peak fluorescence ratio value was appeared at 6 ms group. It is indicated that suitable pulse electrical stimulation could benefit the adhesion of endothelial cell and EPC. All these results provide a new theoretical basis about why electrical stimulation could contribute to neovascularization.
Adult ; Cell Adhesion ; radiation effects ; Cells, Cultured ; Coculture Techniques ; Electric Stimulation ; Female ; Hematopoietic Stem Cells ; cytology ; radiation effects ; Human Umbilical Vein Endothelial Cells ; cytology ; radiation effects ; Humans ; Male ; Young Adult

OBJECTIVETo study the effects of curcumin on vascular endothelial injuries induced by radiation and the mechanism.

METHODSHuman umbilical vein endothelial cells (HUVEC) were isolated, cultured and divided into the control group and 4 groups exposed to 3-ray at the doses of 2, 4, 6 and 8 Gy. Cellular morphological and ultrastructural changes were examined under light microscopy and electron microscopy respectively. Flow cytometry was used to detect the cellular apoptosis, necrosis and intracellular reactive oxygen species(ROS) generation. The contents of Lactate dehydrogenase (LDH) and malondialdehyde (MDA) in the cultures were measured before and after irradiation.

RESULTSThe results of cellular morphological and ultrastructural ex-aminations shown that the typical apoptotic changes appeared after irradiation. The rates of apoptosis and necrosis in groups pretreated with curcumin were significantly lower than those in other groups (P<0.05). LDH and MDA in the irradiation groups were significantly higher than those in curcumin pretreatment groups (P<0.05). The ROS generation in radiation groups significantly increased with the radiation doses, as compared with the groups pre-treated with curcumin (P<0.05).

CONCLUSIONIrradiation induced the apoptosis and necrosis of HUVEC, and increased significantly the intracellular LDH and MDA levels in a dose-dependent. Curcumin had the protective effects on HUVEC from the apoptosis and necrosis induced by radiation.

Apoptosis ; drug effects ; radiation effects ; Curcumin ; pharmacology ; Human Umbilical Vein Endothelial Cells ; drug effects ; radiation effects ; Humans ; L-Lactate Dehydrogenase ; metabolism ; Malondialdehyde ; metabolism ; Reactive Oxygen Species ; analysis ; Vascular System Injuries

OBJECTIVE: Pyroptosis is an inflammatory form of programmed cell death. This phenomenon has been recently reported to play an important role in radiation-induced normal tissue injury. Connexin43 (Cx43) is a gap junction protein that regulates cell growth and apoptosis. In this study, we investigated the effect of Cx43 on X-ray-induced pyroptosis in the human umbilical vein endothelial cells (HUVECs). METHODS: HUVECs, Cx43 overexpression, and Cx43 knockdown strains were irradiated with 10 Gy. Proteins were detected using western blot analysis. Cell pyroptosis was evaluated using the fluorescence-labeled inhibitor of caspase assay (FLICA) and propidium iodide staining through flow cytometry and confocal microscopy. Cell morphology and cytotoxicity were detected by scanning electron microscopy and lactate dehydrogenase release assay, respectively. RESULTS: Irradiation with 10 Gy X-ray induced pyroptosis in the HUVECs and reduced Cx43 expression. The pyroptosis in the HUVECs was significantly attenuated by overexpression of Cx43 as it decreased the level of active caspase-1. However, interference of Cx43 expression with siRNA significantly promoted pyroptosis by increasing the active caspase-1 level. Pannexin1 (Panx1), a gap junction protein regulates pyroptosis, and its cleaved form is used to evaluate channel opening and active state. The level of cleaved Panx1 in the HUVECs and Cx43 knockdown strains increased in the presence of X-ray, but decreased in the Cx43 overexpression strains. Furthermore, interference of Panx1 with siRNA alleviated the upregulation of pyroptosis caused by Cx43 knockdown. CONCLUSION Results suggest that single high-dose X-ray irradiation induces pyroptosis in the HUVECs. In addition, Cx43 regulates pyroptosis directly by activating caspase-1 or indirectly by cleaving Panx1.
Caspase 1 ; genetics ; metabolism ; Connexin 43 ; genetics ; metabolism ; Connexins ; genetics ; metabolism ; Gene Expression Regulation ; radiation effects ; Human Umbilical Vein Endothelial Cells ; physiology ; radiation effects ; Humans ; Nerve Tissue Proteins ; genetics ; metabolism ; Pyroptosis ; X-Rays ; adverse effects

Combined radiation-wound injury (CRWI) is characterized by blood vessel damage and pro-inflammatory cytokine deficiency. Studies have identified that the direct application of leptin plays a significant role in angiogenesis and inflammation. We established a sustained and stable leptin expression system to study the mechanism. A lentivirus method was employed to explore the angiogenic potential and peripheral inflammation of irradiated human umbilical vein endothelial cells (HUVECs). Leptin was transfected into human placenta-derived mesenchymal stem cells (HPMSCs) with lentiviral vectors. HUVECs were irradiated by X-ray at a single dose of 20 Gy. Transwell migration assay was performed to assess the migration of irradiated HUVECs. Based on the Transwell systems, co-culture systems of HPMSCs and irradiated HUVECs were established. Cell proliferation was measured by cell counting kit-8 (CCK-8) assay. The secretion of pro-inflammatory cytokines (human granulocyte macrophage-colony stimulating factor (GM-CSF), interleukin (IL)-1α, IL-6, and IL-8) was detected by enzyme-linked immunosorbent assay (ELISA). The expression of pro-angiogenic factors (vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF)) mRNA was detected by real-time quantitative polymerase chain reaction (RT-qPCR) assay. Relevant molecules of the nuclear factor-κB (NF-κB) and Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathways were detected by western blot assay. Results showed that leptin-modified HPMSCs (HPMSCs/ leptin) exhibited better cell proliferation, migration, and angiogenic potential (expressed more VEGF and bFGF). In both the single HPMSCs/leptin and the co-culture systems of HPMSCs/leptin and irradiated HUVECs, the increased secretion of pro-inflammatory cytokines (human GM-CSF, IL-1α, and IL-6) was associated with the interaction of the NF-κB and JAK/STAT signaling pathways. We conclude that HPMSCs/leptin could promote angiogenic potential and peripheral inflammation of HUVECs after X-ray radiation.
Cell Proliferation ; Cells, Cultured ; Cytokines/biosynthesis* ; Female ; Human Umbilical Vein Endothelial Cells/radiation effects* ; Humans ; Inflammation/etiology* ; Leptin/pharmacology* ; Mesenchymal Stem Cells/physiology* ; Neovascularization, Physiologic/physiology* ; Placenta/cytology* ; Pregnancy ; STAT3 Transcription Factor/genetics* ; Transcription Factor RelA/genetics* ; X-Rays