The Mechanism Of Galectin Family Members In Inducing Apoptosis Of HIV-infected Macrophages

Object AIDS is a disease that seriously threatens human health.Currently,it is usually treated with highly effective antiretroviral therapy.This therapy can significantly improve the quality of life of AIDS patients but it cannot completely cure AIDS due to the presence of virus reserviors.Mononuclear-macrophages are important target cells for HIV infection.How to induce apoptosis of mononuclear macrophages infected by HIV is one of the current research hotspots.Therefore,we studied the mechanism of Galectin family members inducing apoptosis of HIV infected macrophages in this paper.Methods In order to study the effect of Galectin family members on the apoptosis of HIV-infected macrophages,First,different concentrations of Galectins were used to induce apoptosis of monocyte(THP-1)cells,and the suitable Galectin concentration was found.Then PMA was used to stimulate the differentiation of mononuclear cells(THP-1)into macrophages(THP-1-M?),Using the prepared HIV-1 virus to infect macrophages.Finally,5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,3uM Galectin-8,luM Galectin-9 were used to treat monocyte(THP-1),macrophage(THP-1-M?)and HIV-infected macrophages(HIV-THP-1-M?),and its apoptosis was detected.In order to investigate whether the apoptosis of THP-1 induced by Galectin family members is related to the caspase pathway,first of all,different concentration of Caspase broad-spectrum inhibitor(Z-VAD),Caspase-3 inhibitor(Z-DEVD)and Caspase-9 inhibitor(Z-LEHD)were added to the induction of apoptosis in JE6-1 cells with FasL,and the suitable use concentration of Caspase broad-spectrum inhibitor(Z-VAD),Caspase-3 inhibitor(Z-DEVD)and inhibitor inhibitor was found.Then Caspase broad-spectrum inhibitors(Z-VAD),Caspase-3 inhibitor(Z-DEVD)and Caspase-9 inhibitor(Z-LEHD)was added when apoptosis of monocytic cells(THP-1)which were induced by 5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,3uM Galectin-8,luM Galectin-9,respectively.At the same time,Caspase-3 activation and PARP cleavage of apoptotic monocytes(THP-1)induced by 5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,3uM Galectin-8 and luM Galectin-9 were detected by Western blot.Results For macrophages(THP-1-M?),the apoptosis rate of control macrophages(THP-1-Mcp)without addition of Galectin family members was 4.39%±0.74%,the apoptosis rate of 5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,3uM Galectin-8,and 1 uM Galectin-9 induced macrophages(THP-1-M?)were 4.78%± 0.41%,7.21%± 1.46%,3.78%± 1.03%,5.88%± 2.08%and 8.10%± 4.13%,respectively.There was no significant difference between each Galectin treatment groups and the control group.The apoptosis rate of HIV-1-infected macrophage(HIV-1-THP-1-Mcp)without Galectin family member was 12.69%± 1.116%;the apoptosis rates of 5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,3uM Galectin-8 and luM Galectin-9 induced HIV-1-infected macrophages(HIV-1-THP-1-M?)were 11.69%±0.90%,17.45%±t1.30%,32.01%±1.30%,15.77%±11.21%,and 19.27%±2.13%,respectively.The Galectin-7 treatment group had statistically significant differences compared with the control group(p<0.001).The apoptosis rates of 5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,3uM Galectin-8,and luM Galectin-9 induced monocytic cells(THP-1)were 19.66%±0.74%,56.31%±10.60%,21.74%±2.39%,31.85%±5.56%and 88.75%±4.25%.After adding Caspase inhibitor(Z-VAD),the apoptosis rates were 7.33%± 0.47%,8.41%± 0.47%,6.50%± 1.90%,17.74%± 0.23%,8.34%± 0.90%.After adding Caspase-3 inhibitor(Z-DEVD),the apoptosis rates were 9.17%:± 0.42%,10.37%± 0.15%,7.54%±1.65%,20.98±%11.96%,9.03%±0.92%.After adding Caspase-9 inhibitor(Z-LEHD),the apoptosis rates were 25.41%±0.21%,24.28%±3.33%,15.27%±0.89%,25.42%±5.11%,21.58%± 4.74%.The statistical results show that when 5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,and luM Galectin-9 induced apoptosis of monocyte(THP-1),compared with the control group that did not add Caspase inhibitors,the rate of apoptosis induced by 5uM Galectin-2 and luM Galectin-9 was significantly decreased after the addition of Caspase broad-spectrum inhibitor(Z-VAD)and Caspase-3 inhibitor(Z-DEVD)(P<0.01),the rate of apoptosis induced by 5uM Galectin-4 and 7.5uM Galectin-7 was significantly decreased after the addition of Caspase broad-spectrum inhibitor(Z-VAD)and Caspase-3 inhibitor(Z-DEVD)(P<0.05).The apoptosis rate of monocytes(THP-1)induced by 5uM Galectin-2 increased after the addition of Caspase-9 inhibitor(Z-LEHD),there was a significant difference(P<0.05).5uM Galectin-4,7.5uM Galectin-7 induced apoptosis of monocyte(THP-1),when the addition of Caspase-9 inhibitor(Z-LEHD)had no obvious effect on the apoptosis rate,there was no significant difference between the results(P>0.05).The apoptosis rate of monocytes(THP-1)induced by luM Galectin-9 decreased after the addition of Caspase-9 inhibitor(Z-LEHD),there was a significant difference(P<0.01).Compared with the control group without Caspase inhibitor,when 3uM Galectin-8 induced apoptosis of mononuclear cells(THP-1),Caspase inhibitor(Z-VAD),Caspase-3 inhibitor(Z-DEVD)and Caspase-9 inhibitor(Z-LEHD)was added,the apoptosis rate showed no significant difference(P>0.05).At the same time,we detected the activation of Caspase-3 and the cleavage of PARP induced by 5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,3uM Galectin-8 and luM Galectin-9 of mononuclear cells(THP-1)with Western blot we found that 5uM Galectin-2,5uM Galectin-4,7.5uM Galectin-7,luM Galectin-9 induced monocyte(THP-1)apoptosis can detect the size of 32kD Caspase-3 and can detect the size of 17kD and 12kD cleaved Caspase-3.At the same time,the PARP band was weakened and the cleaved PARP band was enhanced.When 3uM Galectin-8 induced monocyte(THP-1)apoptosis,only the size of 32kD Caspase-3 was detected.The PARP and cleaved PARP bands had no significantly difference from the control group.Conclusion Compared with HIV-1-infected macrophages without added Galectin family members,7.5uM Galectin-7 specificity induced massive apoptosis of HIV-1-infected macrophages.Galectin-2,Galectin-4,Galectin-7,Galectin-9 induced monocyte(THP-1)apoptosis was related to Caspase pathway,Galectin-8 induced monocyte(THP-1)apoptosis was not related to Caspase pathway.