lrp6 shrna  (Santa Cruz Biotechnology)

Journal: Endocrine Connections

Article Title: IUGR with catch-up growth programs impaired insulin sensitivity through LRP6/IRS-1 in male rats

doi: 10.1530/ec-21-0203

Figure Lengend Snippet: Figure 3 The expression of LRP6 was decreased in the skeletal muscle of male CG-IUGR. The protein (A) and mRNA (B) levels of LRP6 and β-catenin in skeletal muscle were reduced in the male CG-IUGR rats. Data are presented as mean ± s.e.m. (n = 5). *P < 0.05, **P < 0.01 compared with same-age controls.

Article Snippet: LRP6 shRNA, control shRNA, polybrene and puromycin dihydrochloride were purchased from Santa Cruz Biotechnology Inc (TX).

Techniques: Expressing

Journal: Endocrine Connections

Article Title: IUGR with catch-up growth programs impaired insulin sensitivity through LRP6/IRS-1 in male rats

doi: 10.1530/ec-21-0203

Figure Lengend Snippet: Figure 4 LRP6 is requisite for normal expression of the IR-β/IRS-1. LRP6 specific shRNA decreased the protein (A) and mRNA (B) expression of LRP6, IR-β and IRS-1 in C2C12 cells. Data are presented as mean ±s.e.m. (n = 3). **P < 0.01 compared with same-age controls.

Article Snippet: LRP6 shRNA, control shRNA, polybrene and puromycin dihydrochloride were purchased from Santa Cruz Biotechnology Inc (TX).

Techniques: Expressing, shRNA

Journal: Endocrine Connections

Article Title: IUGR with catch-up growth programs impaired insulin sensitivity through LRP6/IRS-1 in male rats

doi: 10.1530/ec-21-0203

Figure Lengend Snippet: Figure 5 Wnt3a/LRP6 enhanced the expression of IRS-1 and IGF-1R in muscle cells of male CG-IUGR. (A) The curve of LRP6 mRNA expression in primary muscle cells treated with Wnt3a (30 ng/mL) for different time spans in male CG-IUGR rats. (B and C) Upon Wnt3a stimulation for 12 h, the protein levels of LRP6 (B), β-catenin (B) and IRS-1 (C) were increased in the primary muscle cells of male CG-IUGR rats and the expression of IR-β (C) was not altered. (D) The expression of IGF-1R was reduced in the skeletal muscle of male CG-IUGR rats. (E) The expression of IGF-1R was increased upon Wnt-3a stimulation in the primary muscle cells of male CG-IUGR. Data are presented as mean ± s.e.m. (n = 3). **P < 0.01 compared with same age controls.

Article Snippet: LRP6 shRNA, control shRNA, polybrene and puromycin dihydrochloride were purchased from Santa Cruz Biotechnology Inc (TX).

Techniques: Expressing

Journal: Endocrine Connections

Article Title: IUGR with catch-up growth programs impaired insulin sensitivity through LRP6/IRS-1 in male rats

doi: 10.1530/ec-21-0203

Figure Lengend Snippet: Figure 6 Wnt3a/LRP6 increased the expression of mTOR/S6K in skeletal muscle cells of male CG-IUGR rats. (A) The protein levels of mTOR and P70S6K were decreased in skeletal muscle of male CG-IUGR rats (n = 5). (B) The expression of mTOR and P70S6K was increased upon stimulating with Wnt3a (30 ng/mL) for 12 h in skeletal muscle cells of male CG-IUGR rats.

Article Snippet: LRP6 shRNA, control shRNA, polybrene and puromycin dihydrochloride were purchased from Santa Cruz Biotechnology Inc (TX).

Techniques: Expressing

Journal: Journal of Cellular and Molecular Medicine

Article Title: Low‐density lipoprotein receptor‐related protein 6 regulates alternative pre‐ mRNA splicing

doi: 10.1111/jcmm.13682

Figure Lengend Snippet: LRP 6 regulates the muscle‐specific splicing of integrin‐β1D during the development of striated muscle. (A) Time‐lapse microscopy of C2C12 myoblasts throughout the proliferation and myodifferentiation phases. GM : growth medium; DM : differentiation medium. Scale bar: 10 μm. Quantitative PCR (B) and Western blotting (C) analysis of LRP 6 and myodifferentiation markers at indicated time points. C‐left: typical blots; C‐right: pooled data. * P < .05 compared with Ctrl, # P < .05 compared with groups other than Ctrl. (E) Measurement of integrin‐β1 transcripts and proteins in C2C12 cells. Upper: examination of integrin‐β1 transcripts during myodifferentiation using gel electrophoresis; Middle and Lower: Western blotting examination of integrin‐β1 ( ITGB 1) and ITGB 1D during myodifferentiation (middle) and under Lrp6 ‐deficient conditions (lower). E‐middle and ‐lower left: typical blots; E‐middle and ‐lower right: pooled data. * P < .05 compared with Ctrl. Representative images from five independent experiments with similar results are shown

Article Snippet: To generate vectors expressing different LRP6 domains including P1E1P2E2, P3E3P4E4 and ICD, the corresponding sequences were amplified from Lrp6 plasmid (Origene) and subcloned into the pcDNA3.1 expression vector (Invitrogen) using BamHI and XbaI restriction sites.

Techniques: Time-lapse Microscopy, Real-time Polymerase Chain Reaction, Western Blot, Nucleic Acid Electrophoresis

Journal: Journal of Cellular and Molecular Medicine

Article Title: Low‐density lipoprotein receptor‐related protein 6 regulates alternative pre‐ mRNA splicing

doi: 10.1111/jcmm.13682

Figure Lengend Snippet: Roles of splicing repressors in the splicing switching of Itgb1 isoforms. (A) Western blotting examination of hn RNP A1 and PTB proteins during myodifferentiation of C2C12 myoblasts. DM , differentiation medium. Bottom: pooled data. * P < .05 compared with day 0. (B) Schematic of hn RNP A1 and PTB ‐binding sites in the flanking introns of alternative exon D in the splicing reporter of Itgb1 . (C) Effects of hn RNP A1 on the protein expression of ITGB 1D in myocytes. (D) Effects of LRP 6 on the protein expression of hn RNP A1 and PTBP 1. The proteins were extracted from neonatal cardiomyocytes subject to hn RNP A1 ‐ or Lrp6 ‐si RNA s for 48 h. Right: pooled data. * P < .05 compared with si‐Ctrl. (E‐F) Effects of the splice repressor PTBP 1 on the splicing of Itgb1D in the presence and absence of LRP 6 in culture neonatal cardiomyocytes. Western blotting examination of LRP 6, PTBP 1 and ITGB 1D proteins (E). E‐right: pooled data. * P < .05 compared with si‐Ctrl. (F‐top) Gel electrophoresis analysis of Itgb1 RNA s; (F‐bottom) gel electrophoresis analysis of the specific activity of PTBP 1 on Itgb1 pre‐ mRNA using the minigene. (G) Model of the LRP 6‐mediated splicing of Itgb1D in striated muscle cells. Representative blots from five independent experiments with similar results are shown

Article Snippet: To generate vectors expressing different LRP6 domains including P1E1P2E2, P3E3P4E4 and ICD, the corresponding sequences were amplified from Lrp6 plasmid (Origene) and subcloned into the pcDNA3.1 expression vector (Invitrogen) using BamHI and XbaI restriction sites.

Techniques: Western Blot, Binding Assay, Expressing, Nucleic Acid Electrophoresis, Activity Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: Low‐density lipoprotein receptor‐related protein 6 regulates alternative pre‐ mRNA splicing

doi: 10.1111/jcmm.13682

Figure Lengend Snippet: LRP 6 guides the global RNA splicing regulation. (A‐C) Genome‐wide identification by RNA ‐seq of Lrp6 loss‐induced alternative splicing ( AS ) events in cardiomyocytes ( NRCM ), HeLa cells and human umbilical vein endothelial cells ( HUVEC ). (A) Venn diagram for the differential AS events in wild type (Ctrl) and Lrp6 knockdown ( KD ) cells; (B) The pie chart for the percentage of different types of the differential AS events; (C) Number of total LRP 6‐dependent AS events. (D) Functional categories of a conserved set of 94 genes with LRP 6‐dependent AS between humans and rats. (E) The AS of genes identified by RNA ‐seq was verified through RT ‐ PCR analysis in NRCM s. WT , wild type; KD , Lrp6 knockdown

Article Snippet: To generate vectors expressing different LRP6 domains including P1E1P2E2, P3E3P4E4 and ICD, the corresponding sequences were amplified from Lrp6 plasmid (Origene) and subcloned into the pcDNA3.1 expression vector (Invitrogen) using BamHI and XbaI restriction sites.

Techniques: Genome Wide, RNA Sequencing Assay, Functional Assay, Reverse Transcription Polymerase Chain Reaction