Fig 1: ANGPTL3 inhibited the activation of the JAK/STAT3 pathway. SKOV3 ovarian cancer cells were transfected with the recombinant ANGPTL3 plasmids or vector. Then, the protein expression levels of p-JAK2, JAK2, p-STAT3, STAT3, MMP-2 and PD-L1 were analyzed by western blotting (A). The corresponding quantification of the above protein bands was evaluated by ImageJ software (B–D). *P < 0.05 vs. control groups. n = 3. The differences among three groups were analyzed using ANOVA with SNK post-hoc test. JAK2, Janus Kinase2; p-JAK2, phospho-Janus Kinase2; STAT3, Signal transducer and activator of transcription 3; p-STAT3, phospho-Signal transducer and activator of transcription 3; MMP-2, matrix metallopeptidase 2; PD-L1, programmed cell death 1; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.
Fig 2: ANGPTL3 enhancements elevated NK cell cytotoxicity to ovarian cancer cells. (A, B) NK cells activated by IL-2, or not, were co-cultured with ANGPTL3-overexpressed ovarian cancer cells in a Transwell system, including SKOV3 (A) and ES-2 (B) cells. Then, the mRNA expression of CD69 was detected by qRT-PCR. n = 4. (C, D) The contents of TNF-a (C) and IFN-? (D) were evaluated in supernatants from the co-culture of IL-2-treated NK cells and ovarian cancer cells. n = 3. (E, F) NK cell-mediated cytotoxicity was analyzed using an LDH cytotoxic assay kit. (G) Ovarian cancer cell apoptosis was analyzed by flow cytometry. The percentage of apoptotic cells was analyzed by calculating the total percentage of early apoptotic (Annexin V+/PI-) and late necrotic (Annexin V+/PI+) cells. n = 3. The differences among multiple groups were analyzed using ANOVA with SNK post-hoc test. *P < 0.05 vs. control groups. #P < 0.05 vs. IL-2 groups. IL-2, interleukin 2; TNF-a, tumor necrosis factor-alpha; IFN-?, interferon gamma; ANGPTL3, Angiopoietin-like protein 3; E:T, effector-to-target.
Fig 3: Up-regulation of ANGPTL3 inhibited the metastatic potential of ovarian cancer cells. Ovarian cancer cells (SKOV3 and ES-2) were transfected with empty vector or recombinant ANGPTL3 plasmids. Then, cell invasion (A and B) was evaluated by Transwell assay. n = 3. (C–F) SKOV3 and ES-2 cells were treated with TGF-ß and ANGPTL3 recombinant plasmids. Then, the mRNA and protein levels of E-cadherin and N-cadherin were analyzed by qRT-PCR (C, E) and western blotting (D, F). *P < 0.05 vs. control groups. The differences among three or four groups were analyzed using ANOVA with SNK post-hoc test. n = 4. #P < 0.05 vs. TGF-ß groups. TGF-ß, transforming growth factor-ß; ANGPTL3, Angiopoietin-like protein 3; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.
Fig 4: ANGPTL3 overexpression antagonized the proliferation of ovarian cancer cells in vitro. SKOV3 (A) and ES-2 ovarian cancer cells (B) were transfected with the recombinant ANGPTL3 plasmids (ANGPTL3 group) or empty vectors (Vector group, the negative control group). Cells without any transfection were defined as the control group. Then, the mRNA levels of ANGPTL3 were determined by qRT-PCR. (C) Then, the protein expression of ANGPTL3 was analyzed in ovarian cancer cells by western blotting. (D) The corresponding bands were quantified using the Image J software. (E, F) After the transfection with ANGPTL3 vectors, the viability of SKOV3 (E) and ES-2 (F) was evaluated by CCK-8 assay. Control: without any transfection; Vector: pcDNA3.1 (+) negative control transfection; ANGPTL3: recombinant pcDNA3.1-ANGPTL3 plasmid transfection. n = 4. The differences among three groups were analyzed using ANOVA with SNK post-hoc test. *P < 0.05 vs. control groups. ANGPTL3, Angiopoietin-like protein 3; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.
Fig 5: Decreased expression of ANGPTL3 indicated a poor prognosis for patients with ovarian cancer. (A) GEPIA2 database analyzed the expression of ANGPTL3 in ovarian cancer tissues and normal specimens. (B) Expression of ANGPTL3 in various stages of ovarian cancer patients was analyzed using the UALCAN database. (C) The qRT-PCR assay was performed to determine the mRNA levels of ANGPTL3 in collected ovarian cancer and adjacent tissues. n = 4. (D) Protein expression of ANGPTL3 in specimens from 21 ovarian cancer patients was detected by Western blotting. n = 3. (E–G) The correlation between ANGPTL3 and survival of ovarian cancer patients was analyzed by the GEPIA2 (E, F) and Kaplan-Meier Plotter (G). (H, I) The mRNA (H) and protein (I) levels of ANGPTL3 were detected in ovarian cancer cells by qRT-PCR and western blotting. The differences among two or multiple groups were analyzed using Student t-test or ANOVA with SNK post-hoc test. n = 3. OV, Ovarian serous cystadenocarcinoma; TCGA, The Cancer Genome Atlas; ANGPTL3, Angiopoietin-like protein 3; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.
Supplier Page from Abcam for Anti-ANGPTL3 antibody