Effects of IGF-1R gene silencing on the expression of Caspase12 in L02 cells with peroxidation damage
10.3969/j.issn.1006-5725.2014.12.006
- VernacularTitle:IGF-1R基因沉默对L02细胞过氧化损伤时Caspase12表达的影响
- Author:
Jianhua XU
;
Qiping LU
- Publication Type:Journal Article
- Keywords:
Plasmid silence;
IGF-1R;
shRNA;
Caspase12
- From:
The Journal of Practical Medicine
2014;(12):1871-1874
- CountryChina
- Language:Chinese
-
Abstract:
Objective To observe the effects of silencing IGF-1 receptor on the expression of Caspase12 , a endoplasmic reticulum specificity apoptosis protein of the peroxidation damage in L02 cells. Methods Four the recombined plasmids expressing short hairpin RNA (shRNA) specific for sileincing IGF-1R gene were constructed and these shRNAs were potentially targeted on different mRNA sequences of the IGF-1 receptor gene (Sh-H-IG1R- 1108, Sh-H-IGF1R-2797, Sh-H-IGF1R-3215 and Sh-H-IGF1R-3787). These constructs were classed as the experimental group. Another two constructs containing an irrespective sequence and no sequence were defined as a positive control group a negative control group , respectively. All these constructs were transfected into the L02 cells by Lipefectamine 2000. The interference effect was detected by RT-PCR after 24 h and the plasmid stably silencing the expression of IGF-1 receptor was selected for further analysis which was divided into four groups: the blank control group(Group A), the peroxidation damage group(Group B), the negative plasmid transfection group (Group C) and the positive plasmid transfection group (Group D). The expression of Caspase12 was detected by Westen Blot after 12 h. Results The amplification folds of experimental groups were statistically significantly lower than those of the Sh-H-IGF1R-V1 group and Sh-H-IGF1R-V2 group (P < 0.05). The silence effect of the Sh-H-IGF1R-3215 group (0.02 ± 0.01) and the Sh-H-IGF1R-3787 group (0.05 ± 0.05) were most stable and the repeatability of the later was better. The expressions of Caspase12 in Group B (2.41 ± 0.03), Group C (2.45 ± 0.02) and Group D (3.60 ± 0.06) were higher than Group A (0.34 ± 0.02) (P < 0.01). The Group Cexpression was higher than that of the Group B (P > 0.05). The Caspase12 expression in Group D was higher than that in Group B and C (P < 0.01). Conclusion (1) Of all the four constructs in this study, Sh-H-IGF1R-3787 is the most suitable for silencing IGF-1 receptor. (2) Silencing IGF 1R expression using shRNA plasmids can aggravate the L02 cells apoptosis, suggesting that the IGF-1/IGF 1R signal pathway might be involved in regulating the activation of apoptosis in L02 cells with peroxidation damag via endoplasmic reticulum stress.
