MKRN1 promotes colorectal cancer metastasis by activating the TGF-ß signalling pathway through SNIP1 protein degradation.

BACKGROUND: The Makorin ring finger protein 1 (MKRN1) gene, also called RNF61, is located on the long arm of chromosome 7 and is a member of the RING finger protein family. The E3 ubiquitin ligase MKRN1 is closely linked to tumour development, but the exact mechanism needs to be elucidated. In this study, we aimed to investigate the specific mechanism and role of MKRN1 in colorectal cancer (CRC) development. METHODS: MKRN1 expression in CRC was analysed using the Cancer Cell Line Encyclopaedia and the Cancer Genome Atlas (TCGA) databases. Rectal tumour tissues were frozen to explore the MKRN1 expression in CRC and its clinical significance. The impact of MKRN1 on CRC cell proliferation and migration was observed using CCK8, colony formation, wound healing, and transwell assays. A combination of MKRN1 quantitative proteomics, ubiquitination modification omics analysis, and a string of in vitro and in vivo experiments revealed the potential mechanisms by which MKRN1 regulates CRC metastasis. RESULTS: MKRN1 expression was significantly elevated in CRC tissues compared to paracancerous tissues and was positively linked with prognosis (P < 0.01). MKRN1 downregulation inhibits CRC cell proliferation, migration, and invasion. Conversely, MKRN1 overexpression promotes the proliferation, migration, and invasion of CRC cells. Mechanistically, MKRN1 induces epithelial-mesenchymal transition (EMT) in CRC cells via ubiquitination and degradation of Smad nuclear-interacting protein 1 (SNIP1). Furthermore, SNIP1 inhibits transforming growth factor-ß (TGF-ß) signalling, and MKRN1 promotes TGF-ß signalling by degrading SNIP1 to induce EMT in CRC cells. Finally, using conditional knockout mice, intestinal lesions and metastatic liver microlesions were greatly reduced in the intestinal knockout MKRN1 group compared to that in the control group. CONCLUSIONS: High MKRN1 levels promote TGF-ß signalling through ubiquitination and degradation of SNIP1, thereby facilitating CRC metastasis, and supporting MKRN1 as a CRC pro-cancer factor. The MKRN1/SNIP1/TGF-ß axis may be a potential therapeutic target in CRC.

The Management of Portal Vein Thrombosis after Adult Liver Transplantation: A Case Series and Review of the Literature.

BACKGROUND: Portal vein thrombosis (PVT) after adult liver transplantation (LT) is a rare but serious complication with no consensus on the ideal treatment. We report a case series and a comprehensive review of the literature on PVT after LT to discuss the therapeutic options. METHODS: The clinical data of 360 adult patients (≥18 years of age) who underwent LT from January 2017 to January 2020 were reviewed, and a comprehensive search of PubMed and Web of Science was conducted. Patients diagnosed with PVT after LT were identified, and relevant risk factors and therapies were analyzed. RESULTS: Among the 360 patients, 7 (1.94%) developed PVT after LT. Onset of PVT within one week after LT was found in six patients (85.71%). Four of the seven patients with PVT received systemic anticoagulation (low molecular weight heparin and warfarin) therapy. Minimally invasive interventional therapies combined with systemic anticoagulation (heparin and warfarin) were applied for three patients, two of whom died because of severe abdominal hemorrhage and liver failure. Of the 33 cases reported in the literature, minimally invasive interventional therapy combined with systematic anticoagulation or sclerotherapy were the most-used methods (20/33). Systemic anticoagulation was administered to four patients, and surgical operation (thrombectomy; portosystemic shunt and retransplantation) was performed for nine patients. Among these 33 patients, 4 eventually died. CONCLUSIONS: Interventional therapy combined with systemic anticoagulation is a good choice for the management of PVT after LT, and in our experience, systemic anticoagulation alone can also have a positive effect for early PVT patients.

Oct4 Regulates the Transition of Cancer Stem-Like Cells to Tumor Endothelial-Like Cells in Human Liver Cancer.

Octamer-binding transcription factor 4 (Oct4) has been recently implicated as a proangiogenic regulator in several induced pluripotent stem cells (iPSCs), however, its role in cancer stem-like cells (CSCs) remain unclear. We report here that Oct4 participates in tumor vasculogenesis in liver CSCs (LCSCs). We identify that LCSCs possess the potential of endothelial trans-differentiation under endothelial induction, present endothelial specific markers and their functions in vitro, and participate in neovasculogenesis in vivo. The knockdown of the Oct4A by short hairpin RNA (shRNA) in LCSCs represses endothelial trans-differentiation potential, but induces endothelial lineage-restricted differentiation, the latter is positively regulated by Oct4B1. Furthermore, Oct4 regulates vasculogenesis in LCSCs may be via the AKT-NF-κB-p65 signaling pathway. This work reveals Oct4, which is a crucial regulator, plays a critical role in tumor endothelial-like cells transition of LCSCs through Oct4A and Oct4B1 by different ways. The simultaneous inhibition of both the isoforms of Oct4 is hence expected to help regress neovascularization derived from CSCs. Our findings may provide insights to the possible new mechanisms of tumor vasculogenesis for primary liver cancer.

Detection of OPCML methylation, a possible epigenetic marker, from free serum circulating DNA to improve the diagnosis of early-stage ovarian epithelial cancer.

The aim of the present study was to identify the appropriate DNA sequence and design high-quality primers for methylation-specific polymerase chain reaction (MSP). These primers may be used to examine and identify patients with early-stage epithelial ovarian carcinoma (EOC). Opioid binding protein/cell adhesion molecule like (OPCML), Runt-related transcription factor 3 and tissue factor pathway inhibitor 2 were selected as possible molecular markers. MSP primer sets were designed to monitor the methylation of the three markers. Free circulating DNA (fcDNA) from 194 patients with epithelial ovarian carcinoma and healthy donors were templates in the nested MSP. OPCML MSP was effective with respect to screening methylated fcDNA. One-way ANOVA P-values indicated that the difference in cancer antigen 125 (CA125), a biomarker for EOC diagnosis, level between early EOC and healthy donors was not significant. The methylation of OPCML was significantly altered in early-stage EOC compared with healthy donors (P<0.0001), and this supported the hypothesis that specific fcDNA methylation was able to distinguish patients with early-stage EOC from healthy donors. With respect to detecting early EOC, compared with the results of the CA125 test, MSP increased the κ coefficient from 0.140 to 0.757. Therefore, OPCML combined with fcDNA may be used to establish an improved clinical assay compared with the current CA125 test.

[Identification of proteins interacted with Bat3 using tandem affinity purification].

OBJECTIVE: To identify the specific protein interactions involved in Bat3-mediated apoptosis. METHODS: Tandem affinity purification (TAP) was utilized to investigate Bat3-protein interactions, during which full-length human Bat3 fused with Strep2 and FLAG tag as a bait was used to screen the specific protein-protein interactions. The isolated proteins were identified with mass spectrometry. RESULTS: TAP studies showed that Ubl4A was identified as a Bat3-binding partner. Further investigation using co-immunoprecipitation confirmed that Bat3 was associated with Ubl4A. CONCLUSION: TAP was successfully established and is suitable for isolating the binding partners of Bat3.

The molecular mechanism of apoptosis of human umbilical vein endothelial cells induced by monocyte chemotacitic protein-1.

The present study was aimed to investigate whether Bcl-2, Fas and Bax are involved in monocyte chemotacitic protein-1 (MCP-1)-induced apoptosis of human umbilical vein endothelial cells (hUVECs). hUVECs were cultured, and the purity was identified by immunofluorescence and immunohistochemistry with specific anti-von Willebrand factor (vWF) and anti-VEGF receptor-2 (KDR) antibodies. With 90% confluence hUVECs were serum-starved for 12 h, and then treated with different concentrations of MCP-1 (0.1, 1.0, 10, 100 ng/mL) for 24 and 48 h respectively. The expressions of apoptosis related proteins Fas, Bcl-2, Bax were detected by flow cytometry (FACS) and Western blot. As shown in our preliminary study, MCP-1 induced apoptosis of hUVECs in a dose-dependent manner at both 24 h and 48 h. FACS and Western blot analysis results in the present study indicated that MCP-1 promoted the expression of proapoptotic proteins Bax and Fas and inhibited the expression of antiapoptotic protein Bcl-2. These results suggest that MCP-1 may induce the apoptosis of hUVECs through evoking the imbalance between proapoptotic Fas/Bax and antiapoptotic Bcl-2 protein.

[Effect of hesperidin and rutin on oxidative modification of high density lipoprotein in vitro].

OBJECTIVE: To study the effect of hesperidin and rutin on oxidative modification of high density lipoprotein (HDL) in vitro. METHODS: HDL was isolated from healthy human plasma by sequential ultracentrifugation, and was oxidized by copper ions. The inhibitory effects of hesperidin and rutin on HDL oxidative modification were valued by the formation of malondialdehyde (MDA). RESULTS: Hesperidin and rutin significantly inhibited copper-induced oxidation of HDL in a dose-dependent manner. CONCLUSION: Both hesperidin and rutin can prevent HDL from copper-induced oxidative modification in vitro. This result suggests that they might have antiatherogenic effect.