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Involvement of calreticulin in cell proliferation, invasion and differentiation in diallyl disulfide-treated HL-60 cells

View Article: PubMed Central - PubMed

ABSTRACT

Diallyl disulfide (DADS) has shown potential as a therapeutic agent in various cancers. Previously, calreticulin (CRT) was found to be downregulated in differentiated HL-60 cells treated with DADS. The present study investigated the role of CRT proteins in DADS-induced proliferation, invasion and differentiation in HL-60 cells. The present study demonstrated that DADS treatment significantly changed the morphology of HL-60 cells and caused the significant time-dependent downregulation of CRT. Small interfering RNA (siRNA)-mediated knockdown of CRT expression significantly inhibited proliferation, decreased invasion ability, increased the expression of cluster of differentiation (CD)11b and reduced the expression of CD33 in DADS-treated HL-60 cells. DADS also significantly affected cell proliferation, invasion and differentiation in CRT-overexpressed HL-60 cells. Nitroblue tetrazolium (NBT) reduction assays showed decreased NBT reduction activity in the CRT overexpression group and increased NBT reduction in the CRT siRNA group. Following treatment with DADS, the NBT reduction abilities in all groups were increased. In conclusion, the present study clearly demonstrates the downregulation of CRT during DADS-induced differentiation in HL-60 cells and indicates that CRT is involved in cell proliferation, invasion and differentiation in DADS-treated HL-60 cells.

No MeSH data available.


Role of CRT in cell differentiation of HL-60 cells induced by DADS. (A) Representative flow cytometry results showing CD33 expression. (B) Representative flow cytometry results showing CD11b expression. (C) Nitroblue tetrazolium reduction assays: 1, HL-60; 2, DADS+ HL-60; 3, HL-60/CRT; 4, DADS+HL-60/CRT; 5, HL-60/CRT siRNA; 6, DADS+HL-60/CRT siRNA. Data are the mean ± standard deviation of three experiments. *P<0.05 vs. control; Student's t-test. CRT, calreticulin; DADS, diallyl disulfide; CD33, cluster of differentiation 33; CD11b, cluster of differentiation 11b; OD, optical density.
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f4-ol-0-0-4850: Role of CRT in cell differentiation of HL-60 cells induced by DADS. (A) Representative flow cytometry results showing CD33 expression. (B) Representative flow cytometry results showing CD11b expression. (C) Nitroblue tetrazolium reduction assays: 1, HL-60; 2, DADS+ HL-60; 3, HL-60/CRT; 4, DADS+HL-60/CRT; 5, HL-60/CRT siRNA; 6, DADS+HL-60/CRT siRNA. Data are the mean ± standard deviation of three experiments. *P<0.05 vs. control; Student's t-test. CRT, calreticulin; DADS, diallyl disulfide; CD33, cluster of differentiation 33; CD11b, cluster of differentiation 11b; OD, optical density.

Mentions: The present study finally investigated the role of CRT in cell differentiation induced by DADS treatment. As shown in Fig. 4A, FCM analysis showed that, compared with control cells, the expression of CD33 in the CRT siRNA group was markedly reduced (93.53 vs. 51.22%, respectively), while the CRT-overexpressed groups showed slightly increased CD33 expression (95.76%). Following treatment with DADS for 48 h, the expression of CD33 was significantly decreased in non-transfected HL-60 cells (55.46%), CRT-overexpressing HL-60 cells (63.09%), and CRT siRNA HL-60 cells (31.46%) (P=0.037). As shown in Fig. 4B, compared with the control cells (1.00%), the CRT-overexpressed group showed decreased CD11b expression (0.81%) and the CRT siRNA group showed significantly increased CD11b expression (7.37%). After DADS treatment for 48 h, the control group, CRT-overexpressed group and the CRT siRNA group all showed increased CD11b expression (8.24, 6.55 and 12.42%, respectively; P=0.033). NBT reduction assays showed decreased NBT reduction activity in the CRT overexpression group and increased NBT reduction in the CRT siRNA group (Fig. 4C). Following treatment with DADS, the NBT reduction abilities in all groups were increased (P=0.024).


Involvement of calreticulin in cell proliferation, invasion and differentiation in diallyl disulfide-treated HL-60 cells
Role of CRT in cell differentiation of HL-60 cells induced by DADS. (A) Representative flow cytometry results showing CD33 expression. (B) Representative flow cytometry results showing CD11b expression. (C) Nitroblue tetrazolium reduction assays: 1, HL-60; 2, DADS+ HL-60; 3, HL-60/CRT; 4, DADS+HL-60/CRT; 5, HL-60/CRT siRNA; 6, DADS+HL-60/CRT siRNA. Data are the mean ± standard deviation of three experiments. *P<0.05 vs. control; Student's t-test. CRT, calreticulin; DADS, diallyl disulfide; CD33, cluster of differentiation 33; CD11b, cluster of differentiation 11b; OD, optical density.
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Related In: Results  -  Collection

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f4-ol-0-0-4850: Role of CRT in cell differentiation of HL-60 cells induced by DADS. (A) Representative flow cytometry results showing CD33 expression. (B) Representative flow cytometry results showing CD11b expression. (C) Nitroblue tetrazolium reduction assays: 1, HL-60; 2, DADS+ HL-60; 3, HL-60/CRT; 4, DADS+HL-60/CRT; 5, HL-60/CRT siRNA; 6, DADS+HL-60/CRT siRNA. Data are the mean ± standard deviation of three experiments. *P<0.05 vs. control; Student's t-test. CRT, calreticulin; DADS, diallyl disulfide; CD33, cluster of differentiation 33; CD11b, cluster of differentiation 11b; OD, optical density.
Mentions: The present study finally investigated the role of CRT in cell differentiation induced by DADS treatment. As shown in Fig. 4A, FCM analysis showed that, compared with control cells, the expression of CD33 in the CRT siRNA group was markedly reduced (93.53 vs. 51.22%, respectively), while the CRT-overexpressed groups showed slightly increased CD33 expression (95.76%). Following treatment with DADS for 48 h, the expression of CD33 was significantly decreased in non-transfected HL-60 cells (55.46%), CRT-overexpressing HL-60 cells (63.09%), and CRT siRNA HL-60 cells (31.46%) (P=0.037). As shown in Fig. 4B, compared with the control cells (1.00%), the CRT-overexpressed group showed decreased CD11b expression (0.81%) and the CRT siRNA group showed significantly increased CD11b expression (7.37%). After DADS treatment for 48 h, the control group, CRT-overexpressed group and the CRT siRNA group all showed increased CD11b expression (8.24, 6.55 and 12.42%, respectively; P=0.033). NBT reduction assays showed decreased NBT reduction activity in the CRT overexpression group and increased NBT reduction in the CRT siRNA group (Fig. 4C). Following treatment with DADS, the NBT reduction abilities in all groups were increased (P=0.024).

View Article: PubMed Central - PubMed

ABSTRACT

Diallyl disulfide (DADS) has shown potential as a therapeutic agent in various cancers. Previously, calreticulin (CRT) was found to be downregulated in differentiated HL-60 cells treated with DADS. The present study investigated the role of CRT proteins in DADS-induced proliferation, invasion and differentiation in HL-60 cells. The present study demonstrated that DADS treatment significantly changed the morphology of HL-60 cells and caused the significant time-dependent downregulation of CRT. Small interfering RNA (siRNA)-mediated knockdown of CRT expression significantly inhibited proliferation, decreased invasion ability, increased the expression of cluster of differentiation (CD)11b and reduced the expression of CD33 in DADS-treated HL-60 cells. DADS also significantly affected cell proliferation, invasion and differentiation in CRT-overexpressed HL-60 cells. Nitroblue tetrazolium (NBT) reduction assays showed decreased NBT reduction activity in the CRT overexpression group and increased NBT reduction in the CRT siRNA group. Following treatment with DADS, the NBT reduction abilities in all groups were increased. In conclusion, the present study clearly demonstrates the downregulation of CRT during DADS-induced differentiation in HL-60 cells and indicates that CRT is involved in cell proliferation, invasion and differentiation in DADS-treated HL-60 cells.

No MeSH data available.