胰蛋白酶-EDTA溶液(0.05%:0.02%)

Parkinson's disease (PD) is a neurodegenerative disorder that gets exacerbated by vascular injury. Neural stem cell-derived exosomes (NSC-Exos) display effective neuroprotective properties in PD models. Cell division control protein 42 (CDC42) is connected to angiogenesis, but its effects in PD remain undefined. This research aims to reveal the role of CDC42 in PD. First, we applied 1-methyl-4-phenylpyridinium (MPP + ) to induce the human cerebral microvascular endothelial cells (HCMECs) model and evaluated cell viability and ferroptosis. Then, we characterized NSC-Exos. Next, to appraise the effect of hypoxia-pretreated NSC-Exos (H-NSC-Exos) on the MPP + -induced cells model, we examined angiogenesis and ferroptosis in HCMECs. Moreover, we constructed the PD mice model using 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and tested the behavioral experiments and vascular injury of mice. Furthermore, we examined cellular ferroptosis and angiogenesis after knockdown of CDC42. Additionally, we investigated the interaction of CDC42 with Acyl-CoA synthetase long-chain family member 4 (ACSL4) and detected cellular ferroptosis and angiogenesis after overexpression of ACSL4. We found that H-NSC-Exos reversed the MPP + -induced decrease in HCMECs viability and migration, lowered lipid-reactive oxygen species (lipid-ROS) levels, suppressed ferroptosis, and facilitated angiogenesis. Moreover, H-NSC-Exos attenuated MPTP-induced PD development, vascular injury, and ferroptosis in mice. H-NSC-Exos with the knockdown of CDC42 reduced cell viability and angiogenesis and raised ferroptosis and lipid-ROS levels, which were reversed by ferrostatin-1 and liproxstatin-1. CDC42 interacted with ACSL4. Furthermore, overexpression of ACSL4 aggravated the above effects of H-NSC-Exos in which CDC42 was knocked down. Our study reveals that H-NSC-Exos-derived CDC42 inhibited ACSL4-related ferroptosis to alleviate vascular injury in PD mice models. CDC42 may serve as a potent therapeutic target for PD treatment.

Melanoma is a malignant tumor with limited treatment option in advanced stages. Apolipoprotein L3 (APOL3), a protein implicated in immune regulation, has recently emerged as a potential player in tumor immunity. This research aims to explore the potential efficacy of APOL3 in melanoma. Using data from the Cancer Genome Atlas-Skin Cutaneous Melanoma (TCGA-SKCM), we identified two clusters based on 56 prognostic antigen presentation-related genes. Differential expression analysis revealed 185 genes between these two clusters, which were further narrowed down to 34 genes using univariate analysis and random survival forest dimensionality reduction. Among them, APOL3 was found to be the top-ranked gene. Afterward, the effect of APOL3 on melanoma cells was evaluated using CCK-8, EdU, and Transwell experiment. The results showed that overexpression of APOL3 decreases melanoma cell viability, clonogenicity, proliferation, migration, and invasion. Bioinformatics analysis showed the association of high/low APOL3 expression with genomic mutations characterizing melanoma. APOL3 was also found associated with T-cell infiltration levels, immune checkpoints (CD274, PDCD1, CD247, PDCD1LG2, CTLA4, TNFRSF9, TNFRSF4, and TLR9), and some immune pathways. To validate the role of APOL3 on T cell immunity, we applied B16 melanoma cells to construct the mice tumor models. The model showed that APOL3 overexpression markedly reduces melanoma tumor volume and weight while increasing interferon-γ (IFN-γ), granzyme B production, and CD3+ T cell infiltration. In conclusion, antigen presentation-related APOL3 promotes anti-tumor T-cell immunity and suppresses melanoma cell growth in-vitro and in-vivo in a murine model. These results suggested that APOL3 may serve as a promising immunotherapeutic target for the treatment of melanoma.

Background Hirschsprung’s-associated enterocolitis (HAEC) is a prevalent complication of Hirschsprung’s disease (HSCR). Zinc finger E-box binding homeobox 2 (ZEB2) and Notch-1/Jagged-2 are dysregulated in HSCR, but their role in HAEC progression remains poorly understood. We aimed to explore the role and underlying mechanism of enteric neural precursor cells (ENPCs) and the ZEB2/Notch-1/Jagged-2 pathway in HAEC development. Methods Colon tissues were collected from HSCR and HAEC patients. ENPCs were isolated from the HAEC group and stimulated by lipopolysaccharide (LPS). The expressions of ZEB2/Notch-1/Jagged-2 were measured using RT-qPCR and Western blot. Immunofluorescence and cell counting kit-8 assays were performed to assess the differentiation and proliferation of ENPCs. Inflammatory factors were measured by ELISA kits. Co-immunoprecipitation and bioinformatic analysis were used to explore the interaction between ZEB2 and Notch-1. Small interfering RNA and overexpression vectors were used to investigate the role and mechanism of ZEB2 and Notch-1 in regulating ENPCs' proliferation and differentiation during HAEC progression. Results We observed increased LPS in the colon tissues of HAEC, with downregulated ZEB2 expression and upregulated Notch-1/Jagged-2 expression. ZEB2 interacts with Notch-1. LPS treatment downregulated ZEB2 expression, upregulated Notch-1/Jagged-2 expression, and induced proliferation and differentiation disorders in ENPCs, which were reversed by the knockdown of Notch-1. Furthermore, overexpression of ZEB2 inhibited Notch-1/Jagged-2 signaling and ameliorated inflammation and dysfunction in LPS-induced ENPCs. Notch-1 overexpression enhanced LPS-induced dysfunction, but this effect was antagonized by the overexpression of ZEB2. Conclusion Overexpression of ZEB2 ameliorates LPS-induced ENPCs' dysfunction via the Notch-1/Jagged-2 pathway, thus playing a role in HAEC.

This study proposes to investigate the therapeutic efficacy and mechanism of combining tibial transverse transport (TTT) with platelet-rich plasma (PRP) for diabetic foot ulcer (DFU). The diabetic rabbit model was constructed with Streptozotocin, which was intervened with TTT and PRP. PRP injection combined with TTT significantly promoted vascularisation and enhanced CD31, VEGFA, and VEGFR2 expressions compared to traditional TTT. However, the VEGFR2 inhibitor suppressed these phenomena. In the in vitro injury model, PRP reversed the diminished human umbilical vein endothelial cells (HUVECs) function and vascularisation caused by high-glucose damage. Additionally, PRP reduced inflammation and oxidative stress (approximately 47% ROS level) and enhanced VEGFA and VEGFR2 expression in HUVECs. However, the knockdown of VEGFR2 reversed the effect of PRP. In conclusion, TTT combined with intraosseous flap injection of PRP sustained-release microspheres activated the VEGFA/VEGFR2 pathway to promote microcirculatory reconstruction in DFU. These findings may provide new potential therapeutic strategies for DFU.

Colorectal cancer (CRC) is an extremely harmful malignant tumor. Optic atrophy 3 (OPA3) is highly expressed in multiple tumors, but its action in CRC is still unknown. This research aims to explore the role of OPA3 and its related molecular mechanisms for CRC. Firstly, we overexpressed and knocked down OPA3 to examine its effect on CRC cell (HT29 cell) growth. CRC cell viability, migration, invasion, and levels of proliferation markers and cell cycle-associated proteins were measured. Then, we treated cells with carbonyl cyanide m-chlorophenyl hydrazone (CCCP) to explore mitochondrial dysfunction and mtDNA stress in HT29 cells. Next, we overexpressed cGAS and STING to examine their correlation with OPA3. The results showed that OPA3 overexpression enhanced CRC cell viability, migration, invasion, and the levels of PCNA, Cyclin A2, and Cyclin B1. Knockdown of OPA3 had the opposite effects. Moreover, OPA3 knockdown facilitated mitochondrial dysfunction and mtDNA stress in CRC cells. OPA3 overexpression also inhibited CCCP-induced mitochondrial stress disorder. Additionally, OPA3 knockdown elevated the protein levels of p-STING and cGAS and the mRNA level of STING target genes. Furthermore, overexpression of either cGAS or STING partially alleviated the enhancement of HT29 cell proliferation, migration, and invasion mediated by OPA3 overexpression. In conclusion, OPA3 promotes CRC progression via inhibiting the cGAS-STING pathway, which is mediated by mtDNA stress. OPA3 may be a new potential target for CRC.