Fig 1: The 3′ LTR of the provirus drives the overexpression of STAT3.There is a normal allele of STAT3 in tumor 1A. A diagram showing the two STAT3 alleles, and their expression, is on the left. A ddPCR assay (see main text and Materials and Methods) was developed that distinguishes the transcripts from each other and from genomic DNA (which is unspliced). As shown on the right, the LTR-driven STAT3 is expressed at about a 30-fold higher level than the normal transcript.
Fig 2: EP inhibited the abnormal activation of the JAK2/STAT3 signaling pathway. (a) Western blot was employed to determine the protein expression of JAK2, p‐JAK2, STAT3, and p‐STAT3. (b, c) Graphical representation of p‐JAK2 and p‐STAT3 normalized to JAK2 and STAT3, respectively (x¯ ± s, n = 5/group). *p < .05; **p < .01; ns: not significant.
Fig 3: Effect of an miR-1279 mimic on circ_0007059-induced cell viability, apoptosis, and IFN-associated inflammation of RMCs and HEK293 cells. RMC and HEK293 cells were cotransfected with a circ_0007059-overexpressing vector and/or NC/miR-1278 mimic for 24 h, followed by treatment with 1000 units/mL IFN for 24 h. Expression of A, B miR-1278 or C, D SHP-1 was evaluated by RT-qPCR in cells with different transfected constructs (biological replicates, 1; technical replicates, 3; repeat time, 3). E, F WB was performed to detect SHP-1, STAT3, and phosphorylated STAT3 levels in RMCs and HEK293 cells (biological replicates, 1; technical replicates, 3; repeat time, 3). G, H cell viability of RMCs and HEK293 cells was examined using the MTT assay (biological replicates, 1; technical replicates, 3; repeat time, 3). I, J the number of apoptotic cells was assessed by flow cytometry (biological replicates, 1; technical replicates, 3; repeat time, 3). K, L expression of CXCL10, IFIT1, ISG15, and MX1 mRNA in RMCs and HEK293 cells was determined by RT-qPCR (biological replicates, 1; technical replicates, 3; repeat time, 3). Results are expressed as the mean ± SEM for technical replicates. *P < 0.05, **P < 0.01 vs. the indicated group
Fig 4: Influence of SHP-1 overexpression on viability, apoptosis, and IFN-associated inflammation of RMCs and HEK293 cells. RMC and HEK293 cells were transfected with an SHP-1-overexpressing vector or NC vector for 24 h, followed by treatment with 1,000 units/mL IFN for 24 h. Expression of (A, B) SHP-1 was measured in cells with different transfection constructs by RT-qPCR (biological replicates, 2; technical replicates, 3; repeat time, 3). C, D WB was performed to detect SHP-1, STAT3, and phosphorylated STAT3 levels in RMCs and HEK293 cells (biological replicates, 1; technical replicates, 3; repeat time, 3). E, F cell viability of RMCs and HEK293 cells were determined by MTT assay (biological replicates, 1; technical replicates, 3; repeat time, 3). G, H the number of apoptotic cells was evaluated by flow cytometry (biological replicates, 1; technical replicates, 3; repeat time, 3). I, J expression of mRNA for CXCL10, IFIT1, ISG15, and MX1 gene in RMCs and HEK293 cells as determined by RT-qPCR (biological replicates, 2; technical replicates, 3; repeat time, 3). Results are expressed as the mean ± SEM for technical replicates. *P < 0.05, **P < 0.01 vs. the indicated group
Fig 5: circ_0007059 targeted miR-1278 and miR-1278, in turn, targeted SHP-1. A Graphical illustration of the conserved circ_0007059 binding motifs in the 3′-UTR of SHP-1 for miR-1278 and miR-1278-binding motifs. B, C a luciferase reporter assay was used to measure luciferase activity, including wild-type or mutated (MU) copies of human circ_0007059 and SHP-1 after transfection an miR-1278 mimic in HEK293 cells. Luciferase activity was normalized to that of Renilla luciferase (biological replicates, 1; technical replicates, 3; repeat time, 3). D, E the expression of miR-1278 and SHP-1 mRNA in renal biopsy samples from patients with LN (n = 30) and adjacent normal tissue samples (n = 10) as measured by RT-qPCR are shown. RMC and HEK293 cells were transfected with a circ_0007059-overexpressing vector or NC vector for 24 h, followed by treatment with 1000 units/mL IFN for 24 h (biological replicates, as indicated; technical replicates, 3; repeat time, 3). F, G miR-1278 levels in RMCs and HEK293 cells were measured by RT-qPCR (biological replicates, 1; technical replicates, 3; repeat time, 3). H, I RT-qPCR and J, K WB was performed to detect SHP-1, STAT3, and phosphorylated STAT3 levels in RMCs and HEK293 cells (biological replicates, 1; technical replicates, 3; repeat time, 3). Results are expressed as the mean ± SEM for technical replicates. *P < 0.05, **P < 0.01 vs. the indicated group
Supplier Page from Abcam for Anti-STAT3 antibody