Overcoming chemoresistance in non-angiogenic colorectal cancer by metformin via inhibiting endothelial apoptosis and vascular immaturity

The development of chemoresistance which results in a poor prognosis often renders current treatments for colorectal cancer(CRC).In this study,we identified reduced microvessel density(MVD)and vascular immaturity resulting from endothelial apoptosis as therapeutic targets for overcoming chemoresistance.We focused on the effect of metformin on MVD,vascular maturity,and endothelial apoptosis of CRCs with a non-angiogenic phenotype,and further investigated its effect in overcoming chemoresistance.In situ transplanted cancer models were established to compare MVD,endothelial apoptosis and vascular maturity,and function in tumors from metformin-and vehicle-treated mice.An in vitro co-culture system was used to observe the effects of metformin on tumor cell-induced endothelial apoptosis.Transcriptome sequencing was performed for genetic screening.Non-angiogenic CRC developed independently of angiogenesis and was characterized by vascular leakage,immaturity,reduced MVD,and non-hypoxia.This phenomenon had also been observed in human CRC.Furthermore,non-angiogenic CRCs showed a worse response to chemotherapeutic drugs in vivo than in vitro.By suppressing endothelial apoptosis,metformin sensitized non-angiogenic CRCs to chemo-drugs via elevation of MVD and improvement of vascular maturity.Further results showed that endothelial apoptosis was induced by tumor cells via activation of caspase signaling,which was abrogated by metformin administration.These findings provide pre-clinical evidence for the involvement of endothelial apoptosis and subsequent vascular immaturity in the chemoresistance of non-angiogenic CRC.By suppressing endothelial apoptosis,metformin restores vascular maturity and function and sensitizes CRC to chemotherapeutic drugs via a vascular mechanism.

Integrin β5 subunit regulates hyperglycemia-induced vascular endothelial cell apoptosis through FoxO1-mediated macroautophagy

Background:Hyperglycemia frequently induces apoptosis in endothelial cells and ultimately contributes to microvascular dysfunction in patients with diabetes mellitus(DM).Previous research reported that the expression of integrins as well as their ligands was elevated in the diseased vessels of DM patients.However,the association between integrins and hyperglycemia-induced cell death is still unclear.This research was designed to investigate the role played by integrin subunit β5(ITGB5)in hyperglycemia-induced endothelial cell apoptosis.Methods:We used leptin receptor knockout(Lepr-KO)(db/db)mice as spontaneous diabetes animal model.Selective deletion of ITGB5 in endothelial cell was achieved by injecting vascular targeted adeno-associated virus via tail vein.Besides,we also applied small interfering RNA in vitro to study the mechanism of ITGB5 in regulating high glucose-induced cell apoptosis.Results:ITGB5 and its ligand,fibronectin,were both upregulated after exposure to high glucose in vivo and in vitro.ITGB5 knockdown alleviated hyperglycemia-induced vascular endothelial cell apoptosis and microvascular rarefaction in vivo.In vitro analysis revealed that knockdown of either ITGB5 or fibronectin ameliorated high glucose-induced apoptosis in human umbilical vascular endothelial cells(HUVECs).In addition,knockdown of ITGB5 inhibited fibronectin-induced HUVEC apoptosis,which indicated that the fibronectin-ITGB5 interaction participated in high glucose-induced endothelial cell apoptosis.By using RNA-sequencing technology and bioinformatic analysis,we identified Forkhead Box Protein O1(FoxO1)as an important downstream target regulated by ITGB5.Moreover,we demonstrated that the excessive macroautophagy induced by high glucose can contribute to HUVEC apoptosis,which was regulated by the ITGB5-FoxO1 axis.Conclusion:The study revealed that high glucose-induced endothelial cell apoptosis was positively regulated by ITGB5,which suggested that ITGB5 could potentially be used to predict and treat DM-related vascular complications.

Sulindac derivative-induced apoptosis in a human umbilical vein endothelial cell line ECV304

Objective To investigate the effects of sulindac metabolites on proliferation and apoptosis in the human umbilical vein endothelial cell line ECV304 in vitro Methods The proliferation profile of ECV304 was determined by methyl thiazolyl tetrazolium (MTT) method Cell cycle distribution, apoptosis and the ultrastructure of ECV304 were detected by flow cytometry (FCM) and electron microscopy, respectively Results MTT assay showed that the sulfide inhibited the proliferation of ECV304 and its effect was dose dependent; the IC 50 was 200 μmol/L FCM showed that the sulfide changed cell cycle distribution The cell cycle distribution was as follows: G 1 phase (control group 77 74%±1 58%; sulfone group 75 63%±2 12%; sulfide group 46 12%±1 60%); S phase (control group 13 64%±1 22%; sulfone group 16 40±2 30%; sulfide group 27 26%±2 08%); G 2 M phase (control group 8 61%±0 67%; sulfone group 7 98%±0 49%; sulfide group 26 62%±3 54%) The apoptosis rates in the control group, sulfone group and sulfide group were 6 08%±3 39%, 4 81%±2 14% and 51 90%±5 67%, respectively Sulfide reduced the proportion of G 1 phase, increased the proportion of S phase, G 2 M phase and the apoptosis rate significantly ( P <0 01, vs control) In the sulfide treated cells, there were nuclear fragmentation and chromosomal condensation, shrinkage of the cell and loss of contact with neighboring cells Apoptotic bodies were observed Sulfone showed no effect on cell proliferation, cell cycle distribution or cell morphology Conclusions Sulfide can significantly reduce the proliferation of ECV304, change the cell cycle distribution and arrest cells in G 2 M phase where apoptosis may be induced Sulfone has no such effects on this cell line

Effects of salvianolic acids on endothelial cells against damage induced by cholestane-3β-5α-6β-triol

To investigate the effects of salvianolic acids on human umbilical vein endothelial cells (HUVEC) against damage induced by cholestane-3β-5α-6β-triol (chol-triol) Methods The viability of HUVEC was measured by MTT method The apoptosis of HUVEC induced by chol-triol was detected by flow cytometry and TUNEL assay The production of malondialdehyd (MDA) in HUVEC was tested by thiobarbaturic acid (TBA) assay Results The viability of HUVEC treated with chol-triol 100 μmol/L decreased by 39 8% while salvianolic acids 100 μg/ml increased by 27 9% The apoptotic rate of HUVEC measured by PI staining increased from 6%-8% to 17%-20% after chol-triol treatment for 12 h Salvianolic acids 100 μg/ml reduced the apoptotic rate to 10%-14% after treatment HUVEC for 1 h prior to chol-triol treatment In another experiment, chol-triol increased the number of TUNEL-positive cells 5 times, but salvianolic acids 10 μg/ml and 100 μg/ml reduced the number of TUNEL-positive cells by 36 9% and 61 2%, respectively The production of MDA in HUVEC increased by 120 7% after chol-triol treatment for 12 h Salvianolic acids 10 μg/ml and 100 μg/ml also decreased the concentration of MDA by 28 7% and 39 8%, respectively Conclusion Salvianolic acids has protective effect on endothelial cells against damage induced by chol-triol