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Figure 5. Expression of caspase-3 in HEI-OC1 cells. (a) 2% agarose gel electrophoresis of the RT-PCR products of Gapdh, Cs, and <t>Cdh23</t> in HEI-OC1 cells. Lane M, 50 bp DNA ladders; lanes 1–2, 3–4, and 5–6 represent RT-PCR products of Gapdh (100 bp), Cdh23 (228 bp), and Cs (144 bp), respectively. (b) Transcription levels of Cs and Cdh23 in HEI-OC1 cells transfected with shRNA. mRNA levels of Cs was significantly suppressed (p <.05) by Cs shRNA (Cs knockdown group) compared with that by control shRNA (control, limited to1 unit for the level of each gene). mRNA levels of Cs and Cdh23 in cotransfencted group (Cdh23 shRNA þCs shRNA) were also lower than those of the controls. (c) Caspase-3 levels determined by Western blotting. Caspase-3 expression levels were markedly elevated in Cs knockdown group or cotransfected group compared with those of controls, whereas were not elevated in Cdh23 knockdown group. (d) The average gray intensity of caspase-3 protein detected by Western blotting was standardized to that of b-actin. Caspase-3 expression levels in Cs knockdown group or cotransfected group were significantly higher than those of the controls. Data are shown as mean SD of triplicate experiments. *p <.05. shRNA ¼ short hairpin RNA; RT-PCR ¼ reverse transcription polymerase chain reaction; mRNA ¼ messenger RNA; Gapdh ¼ glyceraldehyde 3-phosphate dehydrogenase.
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Figure 5. Expression of caspase-3 in HEI-OC1 cells. (a) 2% agarose gel electrophoresis of the RT-PCR products of Gapdh, Cs, and <t>Cdh23</t> in HEI-OC1 cells. Lane M, 50 bp DNA ladders; lanes 1–2, 3–4, and 5–6 represent RT-PCR products of Gapdh (100 bp), Cdh23 (228 bp), and Cs (144 bp), respectively. (b) Transcription levels of Cs and Cdh23 in HEI-OC1 cells transfected with shRNA. mRNA levels of Cs was significantly suppressed (p <.05) by Cs shRNA (Cs knockdown group) compared with that by control shRNA (control, limited to1 unit for the level of each gene). mRNA levels of Cs and Cdh23 in cotransfencted group (Cdh23 shRNA þCs shRNA) were also lower than those of the controls. (c) Caspase-3 levels determined by Western blotting. Caspase-3 expression levels were markedly elevated in Cs knockdown group or cotransfected group compared with those of controls, whereas were not elevated in Cdh23 knockdown group. (d) The average gray intensity of caspase-3 protein detected by Western blotting was standardized to that of b-actin. Caspase-3 expression levels in Cs knockdown group or cotransfected group were significantly higher than those of the controls. Data are shown as mean SD of triplicate experiments. *p <.05. shRNA ¼ short hairpin RNA; RT-PCR ¼ reverse transcription polymerase chain reaction; mRNA ¼ messenger RNA; Gapdh ¼ glyceraldehyde 3-phosphate dehydrogenase.
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Figure 5. Expression of caspase-3 in HEI-OC1 cells. (a) 2% agarose gel electrophoresis of the RT-PCR products of Gapdh, Cs, and <t>Cdh23</t> in HEI-OC1 cells. Lane M, 50 bp DNA ladders; lanes 1–2, 3–4, and 5–6 represent RT-PCR products of Gapdh (100 bp), Cdh23 (228 bp), and Cs (144 bp), respectively. (b) Transcription levels of Cs and Cdh23 in HEI-OC1 cells transfected with shRNA. mRNA levels of Cs was significantly suppressed (p <.05) by Cs shRNA (Cs knockdown group) compared with that by control shRNA (control, limited to1 unit for the level of each gene). mRNA levels of Cs and Cdh23 in cotransfencted group (Cdh23 shRNA þCs shRNA) were also lower than those of the controls. (c) Caspase-3 levels determined by Western blotting. Caspase-3 expression levels were markedly elevated in Cs knockdown group or cotransfected group compared with those of controls, whereas were not elevated in Cdh23 knockdown group. (d) The average gray intensity of caspase-3 protein detected by Western blotting was standardized to that of b-actin. Caspase-3 expression levels in Cs knockdown group or cotransfected group were significantly higher than those of the controls. Data are shown as mean SD of triplicate experiments. *p <.05. shRNA ¼ short hairpin RNA; RT-PCR ¼ reverse transcription polymerase chain reaction; mRNA ¼ messenger RNA; Gapdh ¼ glyceraldehyde 3-phosphate dehydrogenase.
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Figure 5. Expression of caspase-3 in HEI-OC1 cells. (a) 2% agarose gel electrophoresis of the RT-PCR products of Gapdh, Cs, and Cdh23 in HEI-OC1 cells. Lane M, 50 bp DNA ladders; lanes 1–2, 3–4, and 5–6 represent RT-PCR products of Gapdh (100 bp), Cdh23 (228 bp), and Cs (144 bp), respectively. (b) Transcription levels of Cs and Cdh23 in HEI-OC1 cells transfected with shRNA. mRNA levels of Cs was significantly suppressed (p <.05) by Cs shRNA (Cs knockdown group) compared with that by control shRNA (control, limited to1 unit for the level of each gene). mRNA levels of Cs and Cdh23 in cotransfencted group (Cdh23 shRNA þCs shRNA) were also lower than those of the controls. (c) Caspase-3 levels determined by Western blotting. Caspase-3 expression levels were markedly elevated in Cs knockdown group or cotransfected group compared with those of controls, whereas were not elevated in Cdh23 knockdown group. (d) The average gray intensity of caspase-3 protein detected by Western blotting was standardized to that of b-actin. Caspase-3 expression levels in Cs knockdown group or cotransfected group were significantly higher than those of the controls. Data are shown as mean SD of triplicate experiments. *p <.05. shRNA ¼ short hairpin RNA; RT-PCR ¼ reverse transcription polymerase chain reaction; mRNA ¼ messenger RNA; Gapdh ¼ glyceraldehyde 3-phosphate dehydrogenase.

Journal: ASN neuro

Article Title: Caspase-mediated apoptosis in the cochleae contributes to the early onset of hearing loss in A/J mice.

doi: 10.1177/1759091415573985

Figure Lengend Snippet: Figure 5. Expression of caspase-3 in HEI-OC1 cells. (a) 2% agarose gel electrophoresis of the RT-PCR products of Gapdh, Cs, and Cdh23 in HEI-OC1 cells. Lane M, 50 bp DNA ladders; lanes 1–2, 3–4, and 5–6 represent RT-PCR products of Gapdh (100 bp), Cdh23 (228 bp), and Cs (144 bp), respectively. (b) Transcription levels of Cs and Cdh23 in HEI-OC1 cells transfected with shRNA. mRNA levels of Cs was significantly suppressed (p <.05) by Cs shRNA (Cs knockdown group) compared with that by control shRNA (control, limited to1 unit for the level of each gene). mRNA levels of Cs and Cdh23 in cotransfencted group (Cdh23 shRNA þCs shRNA) were also lower than those of the controls. (c) Caspase-3 levels determined by Western blotting. Caspase-3 expression levels were markedly elevated in Cs knockdown group or cotransfected group compared with those of controls, whereas were not elevated in Cdh23 knockdown group. (d) The average gray intensity of caspase-3 protein detected by Western blotting was standardized to that of b-actin. Caspase-3 expression levels in Cs knockdown group or cotransfected group were significantly higher than those of the controls. Data are shown as mean SD of triplicate experiments. *p <.05. shRNA ¼ short hairpin RNA; RT-PCR ¼ reverse transcription polymerase chain reaction; mRNA ¼ messenger RNA; Gapdh ¼ glyceraldehyde 3-phosphate dehydrogenase.

Article Snippet: When the cells reached 60% confluency in the culture dishes, the cells were transfected with Cs shRNA (Cs shRNA Plasmid, sc-96228-SH, Santa Cruz Biotechnology), Cdh23 shRNA (cadherin 23 shRNA Plasmid, sc-43009-SH, Santa Cruz Biotechnology) or Cs shRNA and Cdh23 shRNA to knockdown the transcription levels of Cs and Cdh23 genes.

Techniques: Expressing, Agarose Gel Electrophoresis, Reverse Transcription Polymerase Chain Reaction, Transfection, shRNA, Knockdown, Control, Western Blot, Reverse Transcription, Polymerase Chain Reaction