Development and characterization of monoclonal antibodies against p37 protein of African swine fever virus.

African Swine Fever Virus (ASFV) is a highly contagious pathogen posing a serious threat to the global swine industry. Despite this, there is currently no effective vaccine against this virus. Within ASFV's core shell structure, p37, a product of polyprotein pp220, shares sequence similarity with SUMO-1 proteases. Localization studies show p37 in various nuclear regions during early infection, shifting to the cytoplasm later on. Research indicates active export of p37 from the nucleus, mediated by CRM1-dependent and -independent pathways. Hydrophobic amino acids in p37 are crucial for these pathways, highlighting their importance throughout the ASFV replication cycle. Additionally, p37 serves as the first nucleocytoplasmic shuttle protein encoded by ASFV, participating in the intranuclear material transport process during ASFV infection of host cells. In this study, we successfully screened five murine monoclonal antibodies targeting p37. Through the truncated expression method, we identified four dominant antigenic epitopes of p37 for the first time. Furthermore, utilizing alanine scanning technology, we determined the key amino acid residues for each epitope. This research not only provides essential information for a deeper understanding of the protein's function but also establishes a significant theoretical foundation for the design and development of ASFV vaccines.

Apolipoprotein A1 suppresses the hypoxia-induced angiogenesis of human retinal endothelial cells by targeting PlGF.

AIM: To investigate the anti-angiogenic effect of apolipoprotein A1 (apoA1) on primary human retinal vascular endothelial cells (HRECs) and explore the possible mechanism. METHODS: The primary HRECs were transfected with apoA1-GFP recombinant lentiviral and were compared with cells undergoing transfection with empty lentiviral vectors. Hypoxia chambers were used to simulate the anoxic environment of cells under pathological condition. The concentrations of secreted vascular endothelial growth factor (VEGF) and placental growth factor (PlGF) were measured by enzyme-linked immunosorbent assay (ELISA). Cell migration ability was detected by wound healing assay. The sprouting of HRECs was determined by tube formation assay. The protein levels of extracellular signal regulated kinase 1/2 (ERK1/2) and phosphorylated ERK1/2 (p-ERK1/2) were measured by Western blot. RESULTS: Overexpressed apoA1 in hypoxia-induced HRECs significantly suppressed PlGF (0.67±0.10 folds, P=0.007). Overexpressed apoA1 also attenuated hypoxia-induced cell migration (0.32±0.11 folds, P<0.0001), tube formation (0.66±0.01 folds, P<0.0001) and the phosphorylation levels of ERK (0.6±0.11 folds, P=0.025). Pretreatment of mitogen-activated protein kinase kinase (MEK) inhibitor (U0126) further reduced the PlGF and angiogenesis in hypoxia-induced HRECs. CONCLUSION: ApoA1 inhibits the angiogenesis at least in part by inactivating ERK1/2 in hypoxia-induced HRECs. Moreover, apoA1 suppresses the PlGF expression, which selectively associated with pathological angiogenesis.