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Galectin-3-induced oxidized low-density lipoprotein promotes the phenotypic transformation of vascular smooth muscle cells.

Tian L, Chen K, Cao J, Han Z, Gao L, Wang Y, Fan Y, Wang C - Mol Med Rep (2015)

Bottom Line: The oxLDL‑induced cells exhibited increased expression levels of osteopontin, a smooth muscle synthetic protein, and calponin and α‑actin, smooth muscle contractile proteins.The oxLDL‑induced changes in cellular phenotype were associated with increased migration, proliferation, and phagocytosis.Concordant with these results, oxLDL‑treated smooth muscle cells exhibited activation of canonical Wnt signaling, as determined by an increase in the protein expression levels of β‑catenin.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology, Shanghai Ninth People's Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200011, P.R. China.

ABSTRACT
Oxidized low-density lipoprotein (oxLDL) is involved in the pathological phenotypic transformation of vascular smooth muscle cells in atherosclerosis. Galectin‑3 also has an important role in atherosclerosis. However, little is currently known regarding the effects of galectin‑3 on the oxLDL‑induced phenotypic transformation of vascular smooth muscle cells. In the present study, primary culture human umbilical vascular smooth muscle cells were treated with various oxLDL concentrations (0‑50 µg/ml) for 72 h, and phenotypic changes were subsequently recorded. The results of the present study suggested that oxLDL increases the expression levels of galectin‑3, and induces the phenotypic transformation of vascular smooth muscle cells. The oxLDL‑induced cells exhibited increased expression levels of osteopontin, a smooth muscle synthetic protein, and calponin and α‑actin, smooth muscle contractile proteins. The oxLDL‑induced changes in cellular phenotype were associated with increased migration, proliferation, and phagocytosis. Concordant with these results, oxLDL‑treated smooth muscle cells exhibited activation of canonical Wnt signaling, as determined by an increase in the protein expression levels of β‑catenin. Silencing of galectin‑3 by small interfering RNA reversed the phenotypic transformation and functional changes observed in the oxLDL‑treated cells, suggesting these changes were dependent on the activation of galectin‑3. In addition, galectin‑3 knockdown decreased the protein expression levels of β‑catenin in both the cytoplasm and nucleus; however, the mRNA expression levels of β‑catenin remained unchanged. These results suggest that galectin‑3 is responsible for the phenotypic transformation of human umbilical vascular smooth muscle cells, and the canonical Wnt/β-catenin signaling pathway may be involved in this process.

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Galectin-3 (gal-3) knockdown reduces the oxidized low-density lipoprotein (oxLDL)-induced activation of human umbilical smooth muscle cells (HUSMCs). (A) Following transfection with either control or gal-3-specific small interfering (si)RNA for 48 h, cellular migration was analyzed using a transwell assay. The cells that migrated from the upper to the lower chamber were counted in five non-overlapping fields under a microscope (magnification, ×100). The cells that migrated to the lower surface of each chamber are shown. (B) The cells transfected with scramble siRNA were defined as control and set to 100. Following transfection with either control or gal-3-siRNA for 24 h, HUSMCs were incubated for 48 h in the absence or presence of 50 μg/ml oxidized low-density lipopolyprotein (oxLDL). *P<0.05, vs. control; #P<0.05, vs. oxLDL. (C) Following transfection, the HUSMCs were incubated for 24 or 48 h in the absence or presence of 50 μg/ml oxLDL. A Cell Counting kit-8 assay was used to detect cell proliferation. The cells transfected with scramble siRNA were defined as control. (D) The HUSMCs were stained with Oil Red O in order to detect neutral lipids. Representative images (magnification, ×40) of the cells stained with Oil Red O are shown. Data are presented as the mean ± standard deviation. *P<0.05, vs. the control. #P<0.05, vs. oxLDL.
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f3-mmr-12-04-4995: Galectin-3 (gal-3) knockdown reduces the oxidized low-density lipoprotein (oxLDL)-induced activation of human umbilical smooth muscle cells (HUSMCs). (A) Following transfection with either control or gal-3-specific small interfering (si)RNA for 48 h, cellular migration was analyzed using a transwell assay. The cells that migrated from the upper to the lower chamber were counted in five non-overlapping fields under a microscope (magnification, ×100). The cells that migrated to the lower surface of each chamber are shown. (B) The cells transfected with scramble siRNA were defined as control and set to 100. Following transfection with either control or gal-3-siRNA for 24 h, HUSMCs were incubated for 48 h in the absence or presence of 50 μg/ml oxidized low-density lipopolyprotein (oxLDL). *P<0.05, vs. control; #P<0.05, vs. oxLDL. (C) Following transfection, the HUSMCs were incubated for 24 or 48 h in the absence or presence of 50 μg/ml oxLDL. A Cell Counting kit-8 assay was used to detect cell proliferation. The cells transfected with scramble siRNA were defined as control. (D) The HUSMCs were stained with Oil Red O in order to detect neutral lipids. Representative images (magnification, ×40) of the cells stained with Oil Red O are shown. Data are presented as the mean ± standard deviation. *P<0.05, vs. the control. #P<0.05, vs. oxLDL.

Mentions: Previous studies (18–20) have reported that oxLDL may promote the activation of VSMCs, including migration, proliferation, and phagocytosis. The present study investigated whether gal-3 was involved in the oxLDL-induced activation of HUSMCs. Migration assays were performed in order to determine the effects of gal-3 silencing on cell motility, and cell numbers, which were determined using a microscope. Knockdown of endogenous gal-3 markedly reduced oxLDL-mediated cell migration in vitro (Fig. 3A and B). A previous study reported that 50 μg/ml oxLDL induced the proliferation of HUSMCs at 24 h (21). The present study demonstrated that silencing gal-3 reduced cell proliferation, and 50 μg/ml oxLDL induced proliferation at 24 h and 48 h (Fig. 3C). These results suggest that gal-3 may affect the proliferation of HUSMCs. The effects of gal-3 knockdown on cell phagocytosis were also investigated. The cells were treated with oxLDL for 48 h, following which lipid accumulation was stained with Oil Red O. A marked increase in Oil Red O staining in the control cells incubated with oxLDL was observed, as compared with the cells transfected with gal-3 siRNA (Fig. 3D). These results suggest that gal-3 has an important role in the oxLDL-induced activation of HUSMC.


Galectin-3-induced oxidized low-density lipoprotein promotes the phenotypic transformation of vascular smooth muscle cells.

Tian L, Chen K, Cao J, Han Z, Gao L, Wang Y, Fan Y, Wang C - Mol Med Rep (2015)

Galectin-3 (gal-3) knockdown reduces the oxidized low-density lipoprotein (oxLDL)-induced activation of human umbilical smooth muscle cells (HUSMCs). (A) Following transfection with either control or gal-3-specific small interfering (si)RNA for 48 h, cellular migration was analyzed using a transwell assay. The cells that migrated from the upper to the lower chamber were counted in five non-overlapping fields under a microscope (magnification, ×100). The cells that migrated to the lower surface of each chamber are shown. (B) The cells transfected with scramble siRNA were defined as control and set to 100. Following transfection with either control or gal-3-siRNA for 24 h, HUSMCs were incubated for 48 h in the absence or presence of 50 μg/ml oxidized low-density lipopolyprotein (oxLDL). *P<0.05, vs. control; #P<0.05, vs. oxLDL. (C) Following transfection, the HUSMCs were incubated for 24 or 48 h in the absence or presence of 50 μg/ml oxLDL. A Cell Counting kit-8 assay was used to detect cell proliferation. The cells transfected with scramble siRNA were defined as control. (D) The HUSMCs were stained with Oil Red O in order to detect neutral lipids. Representative images (magnification, ×40) of the cells stained with Oil Red O are shown. Data are presented as the mean ± standard deviation. *P<0.05, vs. the control. #P<0.05, vs. oxLDL.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4581830&req=5

f3-mmr-12-04-4995: Galectin-3 (gal-3) knockdown reduces the oxidized low-density lipoprotein (oxLDL)-induced activation of human umbilical smooth muscle cells (HUSMCs). (A) Following transfection with either control or gal-3-specific small interfering (si)RNA for 48 h, cellular migration was analyzed using a transwell assay. The cells that migrated from the upper to the lower chamber were counted in five non-overlapping fields under a microscope (magnification, ×100). The cells that migrated to the lower surface of each chamber are shown. (B) The cells transfected with scramble siRNA were defined as control and set to 100. Following transfection with either control or gal-3-siRNA for 24 h, HUSMCs were incubated for 48 h in the absence or presence of 50 μg/ml oxidized low-density lipopolyprotein (oxLDL). *P<0.05, vs. control; #P<0.05, vs. oxLDL. (C) Following transfection, the HUSMCs were incubated for 24 or 48 h in the absence or presence of 50 μg/ml oxLDL. A Cell Counting kit-8 assay was used to detect cell proliferation. The cells transfected with scramble siRNA were defined as control. (D) The HUSMCs were stained with Oil Red O in order to detect neutral lipids. Representative images (magnification, ×40) of the cells stained with Oil Red O are shown. Data are presented as the mean ± standard deviation. *P<0.05, vs. the control. #P<0.05, vs. oxLDL.
Mentions: Previous studies (18–20) have reported that oxLDL may promote the activation of VSMCs, including migration, proliferation, and phagocytosis. The present study investigated whether gal-3 was involved in the oxLDL-induced activation of HUSMCs. Migration assays were performed in order to determine the effects of gal-3 silencing on cell motility, and cell numbers, which were determined using a microscope. Knockdown of endogenous gal-3 markedly reduced oxLDL-mediated cell migration in vitro (Fig. 3A and B). A previous study reported that 50 μg/ml oxLDL induced the proliferation of HUSMCs at 24 h (21). The present study demonstrated that silencing gal-3 reduced cell proliferation, and 50 μg/ml oxLDL induced proliferation at 24 h and 48 h (Fig. 3C). These results suggest that gal-3 may affect the proliferation of HUSMCs. The effects of gal-3 knockdown on cell phagocytosis were also investigated. The cells were treated with oxLDL for 48 h, following which lipid accumulation was stained with Oil Red O. A marked increase in Oil Red O staining in the control cells incubated with oxLDL was observed, as compared with the cells transfected with gal-3 siRNA (Fig. 3D). These results suggest that gal-3 has an important role in the oxLDL-induced activation of HUSMC.

Bottom Line: The oxLDL‑induced cells exhibited increased expression levels of osteopontin, a smooth muscle synthetic protein, and calponin and α‑actin, smooth muscle contractile proteins.The oxLDL‑induced changes in cellular phenotype were associated with increased migration, proliferation, and phagocytosis.Concordant with these results, oxLDL‑treated smooth muscle cells exhibited activation of canonical Wnt signaling, as determined by an increase in the protein expression levels of β‑catenin.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiology, Shanghai Ninth People's Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200011, P.R. China.

ABSTRACT
Oxidized low-density lipoprotein (oxLDL) is involved in the pathological phenotypic transformation of vascular smooth muscle cells in atherosclerosis. Galectin‑3 also has an important role in atherosclerosis. However, little is currently known regarding the effects of galectin‑3 on the oxLDL‑induced phenotypic transformation of vascular smooth muscle cells. In the present study, primary culture human umbilical vascular smooth muscle cells were treated with various oxLDL concentrations (0‑50 µg/ml) for 72 h, and phenotypic changes were subsequently recorded. The results of the present study suggested that oxLDL increases the expression levels of galectin‑3, and induces the phenotypic transformation of vascular smooth muscle cells. The oxLDL‑induced cells exhibited increased expression levels of osteopontin, a smooth muscle synthetic protein, and calponin and α‑actin, smooth muscle contractile proteins. The oxLDL‑induced changes in cellular phenotype were associated with increased migration, proliferation, and phagocytosis. Concordant with these results, oxLDL‑treated smooth muscle cells exhibited activation of canonical Wnt signaling, as determined by an increase in the protein expression levels of β‑catenin. Silencing of galectin‑3 by small interfering RNA reversed the phenotypic transformation and functional changes observed in the oxLDL‑treated cells, suggesting these changes were dependent on the activation of galectin‑3. In addition, galectin‑3 knockdown decreased the protein expression levels of β‑catenin in both the cytoplasm and nucleus; however, the mRNA expression levels of β‑catenin remained unchanged. These results suggest that galectin‑3 is responsible for the phenotypic transformation of human umbilical vascular smooth muscle cells, and the canonical Wnt/β-catenin signaling pathway may be involved in this process.

Show MeSH
Related in: MedlinePlus