A novel approach for transforming breast cancer stem cells into endothelial cells.

Tumor vascular endothelial cells play a pivotal in the tumor microenvironment, influencing the proliferation, invasion, and metastasis of tumor progression. The present study investigated a novel method for inducing the transformation of breast cancer stem cells into endothelial cells, providing a cellular model investigating anti-angiogenic mechanisms in vitro. The breast cancer cell line MCF-7 was used, and the expression of CD133 was initially detected using flow cytometry. CD133+ breast cancer cells were purified using immunomagnetic bead sorting technology, yielding an MCF-7CD133+ subpopulation. The proliferation ability of these cells was assessed using an MTT assay, while their microsphere formation ability was evaluated using a microsphere formation assay. Post-transformation in an optimized endothelial cell culture medium, expression of endothelial cell markers CD31 and CD105 were detected using flow cytometry. Endothelial cell tube formation assays and DiI-labeled acetylated low-density lipoprotein (DiI-Ac-LDL) assays were employed to analyze the endothelial cell function of the MCF-7CD133+ cells. MDM2/CEN12 gene amplification was detected through fluorescence in situ hybridization (FISH). The MCF-7 breast cancer cell line exhibited 1.7±0.3% trace cells expressing the stem cell surface marker CD133. After anti-CD133 immunomagnetic bead sorting, MCF-7CD133+ and MCF-7CD133- subpopulation cells were obtained, with CD133 expression rates of 85.6±2.8 and 0.18±0.08%, respectively. MTT assay results demonstrated that, after 7 days, the proliferation rate of MCF-7CD133+ cells was significantly higher compared with MCF-7CD133- cells. MCF-7CD133+ subpopulation cells displayed strong stem cell characteristics, growing in suspension in serum-free media and forming tumor cell spheres. In contrast, MCF-7CD133- cells failed to form microspheres. After culturing cells in endothelial cell differentiation and maintenance media, the percentage of MCF-7CD133+ cells before and after endothelial cell culture was 0.3±0.16 and 81.4±8.37% for CD31+ cells and 0.2±0.08 and 83.8±7.24% for CD105+ cells, respectively. Vascular-like structure formation and Ac-LDL phagocytosis with red fluorescence in the tube formation assays confirmed endothelial cell function in the MCF-7CD133+ cells. FISH was used to verify MDM2/CEN12 gene amplification in the induced MCF-7CD133+ cells, indicating tumor cell characteristics. The modified endothelial cell transformation medium effectively induced differentiated tumor stem cells to express vascular endothelial cell markers and exhibit endothelial functions, ideal for in vitro anti-angiogenesis research.

METTL3/IGF2BP3-regulated m6A modification of HYOU1 confers doxorubicin resistance in breast cancer.

Chemoresistance is a main reason for therapeutic failure and poor prognosis for breast cancer (BC) patients, especially for triple-negative BC patients. How the molecular mechanisms underlying the chemoresistance to doxorubicin (Dox) in BC is not well understood. Here, we revealed that METTL3/IGF2BP3-regulated m6A modification of HYOU1 increased Dox resistance in BC cells. CCK-8 and Annexin V-FITC/PI staining assays were employed to measure viability and cell death. Western blotting and qRT-PCR assays were applied to assay the expression of genes. Knockdown and rescue experiments were used to assay the role of METTL3, IGF2BP3 and HYOU1 in regulating BC cell responses to Dox. RIP, MeRIP and dual-luciferase activity assays were applied to examine the function of METTL3/IGF2BP3 in the m6A modification of HYOU1 mRNA. It was found that global mRNA m6A methylation levels were upregulated in Dox-resistant BC cell lines. The methyltransferase METTL3 was upregulated in Dox-resistant BC cell lines, and downregulation of METTL3 could overcome this resistance. Furthermore, HYOU1 was identified as a downstream target of METTL3-mediated m6A modification. Downregulation of HYOU1 could overcome Dox resistance, while forced expression of HYOU1 resulted in Dox resistance in BC cells. METTL3 cooperated with IGF2BP3 to modulate the m6A modification of HYOU1 mRNA and increase its stability. Collectively, our findings unveiled the key roles of the METTL3/IGF2BP3/HYOU1 axis in modulating Dox sensitivity in BC cells; thus, targeting this axis might be a potential strategy to increase Dox efficacy in the treatment of BC.

"Elastic stretch cavity building" system in endoscopic thyroidectomy via the axillary approach: a case series.

Background: Patients undergoing conventional endoscopic thyroidectomy via the axillary approach, which is commonly used clinically, suffered from a range of postoperative complications. This study aimed to prevent postoperative complications and evaluate patients' satisfaction with cosmetic outcomes in endoscopic thyroidectomy via the axillary with the use of "Elastic Stretch Cavity Building" System. Methods: In this retrospective case series study, the clinical data of patients who were admitted to the Thyroid Surgery Department of Ningbo Medical Centre Lihuili Hospital between December 2020 and December 2021 for endoscopic thyroidectomy via the axillary approach under the "Elastic Stretch Cavity Building" System. Results: A total of 67 patients were included, all surgeries were successfully completed. The operation time was 75.61 ± 13.67 minutes; the postoperative drainage volume was 109.97 ± 37.54 ml; the average postoperative hospital stay was 4 (2-6) days. There was no skin ecchymosis, effusion or infection, hypocalcemia, convulsions, upper extremity dyskinesia, and temporary hoarseness after the surgery. The patients were satisfied with the cosmetic effects, and the cosmetic score was 4 (3-4). Conclusion: The "Elastic Stretch Cavity Building" System in endoscopic thyroid surgery via the axillary approach might reduce the risks of complications and achieve satisfactory results with the cosmetic outcomes.

Corrigendum: Purified PTEN-Long induces liver cancer cells to undergo autophagy and apoptosis.

[This corrects the article DOI: 10.3389/fsurg.2022.767611.].

Purified PTEN-Long Induces Liver Cancer Cells to Undergo Autophagy and Apoptosis.

Background: PTEN-Long is a translational variant of phosphatase and tensin homolog deleted on chromosome 10 (PTEN). This tumor suppressor is frequently lost or mutated and even it has been shown as the determinant in several human tumors. Therefore, we will determine the significant roles of PTEN-Long in the development of liver cancer. Methods: In the present study, we characterized the antitumor effects of PTEN-Long and PTEN in proliferation, migration of HepG2 cells, apoptosis and autophagy in liver cancer cells. To extends, we have also measured the effects of purified PTEN and PTEN-Long in the above index of HepG2 cells. Results: PTEN and PTEN-Long were ectopic-expressed in HepG2 cells, and their phenotypic effects were recorded. As expected, there was less expression of PTEN-Long and PTEN in liver cancer samples than in paired normal tissues. Ectopic expression of PTEN-Long or PTEN significantly decreased the proliferation and migration of HepG2 cells and increased apoptosis. PTEN ectopic-expression increased the number of GFP-/RFP+-LC3 puncta and levels of beclin-1 and LC3BII/LC3BI, suggesting autophagy induction. Purified PTEN-Long freely entered cells, decreased proliferation, and increased autophagy and apoptosis, while purified PTEN did not. Conclusions: Our results identify an antitumor function of purified PTEN-Long and suggest its potential utility for liver cancer treatment.

Hypermethylated KLF9 Is An Independent Prognostic Factor For Favorable Outcome In Breast Cancer.

BACKGROUND AND OBJECTIVE: Breast cancer (BC) is the most lethal human malignancy and is the leading cause of cancer-associated death in women worldwide. Krüppel-like factor 9 (KLF9) belongs to a family of transcriptional regulators and its role in BC has not been fully investigated. METHOD: Data mining was used to analyze BC data from The Cancer Genome Atlas (TCGA) database, which was downloaded using the UCSC Xena browser. The differential expression and methylation level of KLF9 was analyzed in patients with BC and corresponding normal controls enrolled from our hospital. Besides, the correlation of KLF9 methylation and prognosis was explored, and gene set enrichment analysis (GSEA) was conducted to identify the potential signaling pathway of KLF9 involved. RESULTS: Both TCGA and BC tissues indicated hypermethylation of the KLF9 promoter region in patients with BC compared with normal controls, which might account for the dysregulation of KLF9 in patients with BC. Besides, hypermethylation of KLF9 was detected in patients with estrogen or progesterone receptor-positive and non-triple-negative disease. Further, hypermethylation of KLF9 was demonstrated to be a potential independent biomarker in obtaining favorable outcomes in BC. By GSEA, tumor-associated biological processes and signaling pathway were identified, which indicated that KLF9 might play a vital role in the carcinogenesis of BC. CONCLUSION: KFL9 plays an important role in the carcinogenesis of BC through the multiple tumor-associated signaling pathway. The hypermethylation of KLF9 resulted in its reduced expression in BC, while the hypermethylation of KLF9 has potential in the prediction of favorable outcomes in BC.