Fig 1: PRKCB Was Upregulated in GC and Positively Related to hsa_circ_0092306(A) Quantitative real-time PCR showed PRKCB was significantly upregulated in 45 paired GC and adjacent tissues. (B) The PRKCB expression was positively related to the hsa_circ_0092306 expression in 45 GC tissues. (C) The expressions of PKCß1 protein and PRKCB mRNA in the normal gastric mucosal epithelial cells and GC cells were detected by western blot and quantitative real-time PCR. (D and E) The expressions of PKCß1 protein and PRKCB mRNA after regulating the hsa_circ_0092306 expression were detected in MKN-45 cells (D) and tumor tissues (E). (F) The expression of PKCß1 protein after regulating the hsa_circ_0092306 expression was determined by immunohistochemistry. **p < 0.01 and ***p < 0.001 meant statistical significance.
Fig 2: PRKCB Was the Downstream Functional Gene of hsa_circ_0092306/miR-197-3p in MKN-45 Cells(A and B) The PKCβ1 protein (A) and PRKCB mRNA (B) levels after regulating miR-197-3p or hsa_circ_0092306 in MKN-45 cells were detected by western blot and quantitative real-time PCR, respectively. (C) Flow cytometry showed that PRCKB inhibited cell apoptosis in MKN-45 cells. (D) Cell viability measured by MTT showed a higher cell viability of MKN-45 cells in the PRCKB-overexpressed group. (E) Wound healing and Transwell assays showed higher cell migration and invasion in the PRCKB overexpression group. *p < 0.05, **p < 0.01, and ***p < 0.001 meant statistical significance.
Fig 3: Validation of the expression of the mTOR pathway markers in diabetic nephropathy. (A,B) Box plots of the expression of mTOR pathway markers EIF4B, RICTOR, and PRKCB in control and diabetic nephropathy samples in the (A) GSE111154 and (B) GSE142025 datasets. DN: diabetic nephropathy. (C) H&E staining of the morphology of kidney tissues in the control group and the diabetic nephropathy rat model group. Scale bar, 100 μm; magnification, ×400. (D) The mRNA expression of EIF4B, RICTOR, and PRKCB was determined in kidney tissues from the control and diabetic nephropathy groups through RT-qPCR. (E,F) The expression of EIF4B, RICTOR, and PRKCB proteins was determined in kidney tissues from the control and diabetic nephropathy groups through Western blot. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.
Fig 4: miR-197-3p Targeted hsa_circ_0092306 and PRKCB(A) Quantitative real-time PCR was performed and showed a lower level of miR-197-3p in 45 paired GC and adjacent tissues. (B) There was a negative relationship between hsa_circ_0092306 and miR-197-3p (R2 = 0.3678). (C) hsa_circ_0092306 suppressed the miR-197-3p expression in MKN-45 cells. (D) miR-197-3p was upregulated in the hsa_circ_0092306 knockdown tumors. (E) There was a negative relationship between PRKCB and miR-197-3p (R2 = 0.6766). (F) The targeting sequence between hsa_circ_0092306 and miR-197-3p was revealed by the Circular RNA Interactome, and the luciferase reporter experiment indicated the target relationship of hsa_circ_0092306 and miR-197-3p. (G) miR-197-3p enriched in the pull-down production with the hsa_circ_0092306 probe. (H) The targeting sequence between PRKCB and miR-197-3p was revealed by the TargetScan, and the luciferase reporter experiment indicated the target relationship of PRKCB and miR-197-3p. **p < 0.01 and ***p < 0.001 meant statistical significance.
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