PUM2 promoted osteoarthritis progression through PTEN-mediated chondrocyte ferroptosis by facilitating NEDD4 mRNA degradation.

Osteoarthritis (OA) is a prevalent degenerative joint disease with a lack of effective therapeutic. Chondrocyte ferroptosis contributes to the progression of OA. PUM2 is shown to exacerbate ischemia-reperfusion-induced neuroinflammation by promoting ferroptosis, but its role in OA remains unexplored. Here, primary mouse chondrocytes were stimulated with IL-1ß to mimic OA chondrocyte injury in vitro. And PUM2 was upregulated in OA cartilage tissues and IL-1ß-induced chondrocytes. Silencing PUM2 alleviated IL-1ß-induced chondrocyte inflammation and ECM degradation. Mechanistically, PUM2 facilitated the degradation of NEDD4 mRNA by binding to the 3'UTR of NEDD4 mRNA, which in turn inhibited NEDD4 induced PTEN ubiquitination and degradation. Consistently, NEDD4 silencing reversed the ameliorative effect of PUM2 knockdown on chondrocyte injury, and overexpression of PTEN abolished the improved role of NEDD4 in chondrocyte injury. Moreover, PTEN aggravated IL-1ß-induced ferroptosis in chondrocytes through the Nrf2/HO-1 pathway by increasing the levels of Fe2+, ROS, MDA, and ACSL4 protein, decreasing the activity of SOD and the levels of GSH and GPX4 protein, and aggravating mitochondrial damage. Additionally, destabilized medial meniscus (DMM) were conducted to establish the OA mouse model, and adenovirus-mediated PUM2 shRNA was administered intra-articularly. Silencing PUM2 attenuated OA-induced cartilage damage in vivo. In conclusion, PUM2 promoted OA progression through PTEN-mediated chondrocyte ferroptosis by facilitating NEDD4 mRNA degradation.

Characterization and comparison site-specific N-glycosylation profiling of milk fat globule membrane proteome in donkey and human colostrum and mature milk.

Milk fat globule membrane (MFGM) proteins are highly glycosylated and involved in various biological processes within the body. However, information on site-specific N-glycosylation of MFGM glycoproteins in donkey and human milk remains limited. This study aimed to map the most comprehensive site-specific N-glycosylation fingerprinting of donkey and human MFGM glycoproteins using a site-specific glycoproteomics strategy. We identified 1,360, 457, 2,617, and 986 site-specific N-glycans from 296, 77, 214, and 196 N-glycoproteins in donkey colostrum (DC), donkey mature milk (DM), human colostrum (HC), and human mature milk (HM), respectively. Bioinformatics was used to describe the structure-activity relationships of DC, DM, HC, and HM MFGM N-glycoproteins. The results revealed differences in the molecular composition of donkey and human MFGM N-glycoproteins and the dynamic changes to site-specific N-glycosylation of donkey and human MFGM glycoproteins during lactation, deepening our understanding of the composition of donkey and human MFGM N-glycoproteins and their potential physiological roles.

Glycoproteomics analysis reveals differential site-specific N-glycosylation of donkey milk fat globule membrane protein during lactation.

N-glycosylation is a prevalent and complex post-translational modification of milk proteins with significant biological importance. However, the systematic characterisation of donkey milk fat globule membrane (MFGM) N-glycoproteins remains largely ill-defined. Here, 1443 intact N-glycopeptides from 336 MFGM glycoproteins in donkey colostrum (DC) and 489 intact N-glycopeptides from 86 MFGM glycoproteins in donkey mature milk (DM) were identified via label-free site-specific glycoproteomics. Mannosylation and fucosylation were predominant in DC MFGM N-glycoproteins compared to sialylation and mannosylation in DM. Among them, 22 site-specific N-glycans attached to 14 glycosites of eight glycoproteins were significantly increased, whereas 30 site-specific N-glycans attached to 19 glycosites of 16 glycoproteins were significantly decreased. Furthermore, the site-specific N-glycans with Neu5Gc moieties or simultaneous fucosylation and sialylation were not significantly increased, exhibiting significant site specificity. We provide new insights into the composition of donkey MFGM N-glycoproteins and their roles in donkey milk-related biological functions.

A new sight to explore site-specific N-glycosylation in donkey colostrum milk fat globule membrane proteins with glycoproteomics analysis.

Donkey colostrum milk fat globule membrane (DCMFGM) proteins are involved in multiple biological functions. However, the effect of N-glycosylation on their physiological properties are unknown. The aim of this study was to map the DCMFGM protein site-specific N-glycosylation landscape using a label-free glycoproteomic approach. A total of 1,443 unique intact N-glycopeptides mapping to 453 unique N-glycosites on 336 N-glycoproteins were identified. The macro- and microheterogeneity of DCMFGM glycoproteins were explored at the N-glycosite level and the site-specific N-glycan level, respectively, and it was found that the N-glycosylation profiles of the DCMFGM proteins varied based on subcellular localisation and protein domain types. Our findings reveal the heterogeneity and functional diversity of N-glycosylation of DCMFGM proteins and provide theoretical support for the promotion of DCMFGM proteins as a functional food ingredient.

The anti-metastatic effect of baicalein on colorectal cancer.

Baicalein, a naturally occurring flavonoid isolated from the roots of Scutellaria baicalensis, is historically and widely used as anti-inflammatory and anticancer therapy. Nevertheless, the anti-metastatic effect and underlying molecular mechanisms of baicalein on colorectal carcinoma (CRC) remain unclear. The aim of the present study was, therefore, to invastigate the anti-metastatic activity of baicalein and related mechanism(s) on CRC cells. In this study, we observed that baicalein treatment inhibited proliferation, as well as migration and invasion of HT-29 and DLD1 cells. Baicalein decreased the expression of the matrix metalloproteinases-2 (MMP-2) and MMP-9 in a dose-dependent manner. Also, baicalein treatment significantly reduced phosphorylation of extracellular signal regulated kinases (ERK). Furthermore, in DLD1 cells, MEK1 overexpression partially blocked the anti-metastatic effects of baicalein. Combined treatment with an ERK inhibitor (U0126) and baicalein led to the synergistic reduction of MMP-2/9 expression; and the invasive capabilities of DLD1 cells were also inhibited markedy. Finally, intragastric administration of baicalein inhibited CRC xenograft growth in vivo and suppressed the phosphorylation of ERK and the expression of MMP-2/9 in tumor tissues. Consequently, baicalein suppresses CRC cell invasion via inhibition of the ERK signaling pathways, indicating that baicalein is a potential agent for CRC treatment.

[Expression of EphA7 protein in primary hepatocellular carcinoma and its clinical significance].

OBJECTIVE: To investigate the expression and clinical significance of EphA7 protein in primary hepatocellular carcinoma. METHODS: Immunohistochemistry and Western blot were used to detect the expression of EphA7 protein in 40 cases of primary hepatocellular carcinoma, their corresponding adjacent liver tissues and 10 cases of normal liver tissues. The relations with its clinical pathological parameters were analyzed too. RESULTS: Expression of EphA7 protein was mainly located in the cytoplasm and the blood vessels of the septa, which was found in hepatocellular carcinoma tissues, their corresponding adjacent liver tissues and normal liver tissues. Western blot analysis showed that the expression level of EphA7 protein in hepatocellular carcinoma (0.58 +/- 0.26) was greater than that in corresponding adjacent liver tissues (0.40 +/- 0.22, P < 0.05) and normal liver tissues (0.32 +/- 0.16, P < 0.05). But it had no significant difference between corresponding adjacent liver tissues and normal liver tissues (P > 0.05). EphA7 protein expression was correlated with histological differentiation, tumor thrombi in portal vein, lymph node metastasis and high AFP level (P < 0.05). CONCLUSIONS: EphA7 protein expression is significantly correlated with the biological behavior of primary hepatocellular carcinoma. The high expression of EphA7 protein may play an important role in the malignancy transformation, invasion progression and metastasis of primary hepatocellular carcinoma.