Antibody Lineages with Vaccine-Induced Antigen-Binding Hotspots Develop Broad HIV Neutralization.

The vaccine-mediated elicitation of antibodies (Abs) capable of neutralizing diverse HIV-1 strains has been a long-standing goal. To understand how broadly neutralizing antibodies (bNAbs) can be elicited, we identified, characterized, and tracked five neutralizing Ab lineages targeting the HIV-1-fusion peptide (FP) in vaccinated macaques over time. Genetic and structural analyses revealed two of these lineages to belong to a reproducible class capable of neutralizing up to 59% of 208 diverse viral strains. B cell analysis indicated each of the five lineages to have been initiated and expanded by FP-carrier priming, with envelope (Env)-trimer boosts inducing cross-reactive neutralization. These Abs had binding-energy hotspots focused on FP, whereas several FP-directed Abs induced by immunization with Env trimer-only were less FP-focused and less broadly neutralizing. Priming with a conserved subregion, such as FP, can thus induce Abs with binding-energy hotspots coincident with the target subregion and capable of broad neutralization.

Epitope-based vaccine design yields fusion peptide-directed antibodies that neutralize diverse strains of HIV-1.

A central goal of HIV-1 vaccine research is the elicitation of antibodies capable of neutralizing diverse primary isolates of HIV-1. Here we show that focusing the immune response to exposed N-terminal residues of the fusion peptide, a critical component of the viral entry machinery and the epitope of antibodies elicited by HIV-1 infection, through immunization with fusion peptide-coupled carriers and prefusion stabilized envelope trimers, induces cross-clade neutralizing responses. In mice, these immunogens elicited monoclonal antibodies capable of neutralizing up to 31% of a cross-clade panel of 208 HIV-1 strains. Crystal and cryoelectron microscopy structures of these antibodies revealed fusion peptide conformational diversity as a molecular explanation for the cross-clade neutralization. Immunization of guinea pigs and rhesus macaques induced similarly broad fusion peptide-directed neutralizing responses, suggesting translatability. The N terminus of the HIV-1 fusion peptide is thus a promising target of vaccine efforts aimed at eliciting broadly neutralizing antibodies.

Prognostic significance of Notch 3 gene expression in ovarian serous carcinoma.

The Notch signaling pathway is an important cell signaling system, which regulates cell differentiation, proliferation, and apoptosis, and is aberrantly activated in a wide range of cancer, including ovarian cancers. However, it remains unclear as to whether Notch signaling plays a role in the progression and prognosis of ovarian cancer. We examined the mRNA and protein expression of Notch 3, Jagged 1, and Jagged 2 in 98 ovarian epithelial tumors via real-time PCR and in 175 tumors with immunohistochemical analysis, and then correlated their expression levels with clinicopathological parameters and patient survival. In this study, we detected high levels of Notch3 mRNA and protein expression especially in serous ovarian carcinomas compared to their benign counterparts, accompanied by a positive correlation with the expressions of Jagged 1 and Jagged 2. High levels of Notch 3 mRNA expression (>2-fold than that of benign tumor) were noted in 63% of the serous carcinomas (mean level: 17-fold, P = 0.032). Additionally, Notch 3 protein overexpression was significantly associated with advanced stage (P = 0.0008), lymph node (P = 0.001), and distant metastasis (P = 0.003). Notably, high Notch 3 mRNA and protein expressions were correlated with chemoresistance (P = 0.033) and poor overall survival (P = 0.027, P = 0.042) in these patients. Our results indicate that the Notch 3 signaling pathway is involved in the tumor progression of ovarian serous carcinoma, and higher Notch 3 expression may be an independent poor prognostic factor in this subset of tumors.

The use of green fluorescence gene (GFP)-modified rabbit mesenchymal stem cells (rMSCs) co-cultured with chondrocytes in hydrogel constructs to reveal the chondrogenesis of MSCs.

This study was conducted to reveal the chondrogenesis of mesenchymal stem cells that had been genetically modified with the green fluorescence protein (GFP) gene and then co-cultured with chondrocytes in vitro and in vivo. Subsequent mixing of chondrocytes in the hydrogel constructs induced increased chondrogenic differentiation of the transfected hMSCs. The proliferation and differentiation of MSCs that were transfected with the GFP gene and co-cultured with chondrocytes (1:1 and 1:3) or chondrocytes alone were evaluated by a live/dead assay, MTT assay, GAG & DNA assay, RT-PCR, real time-PCR, and histological and immunochemical analysis in vitro and in vivo. Real-time PCR revealed that the expression of aggrecan and COMP by genetically modified hMSCs co-cultured with chondrocytes was 2 or 3 times greater than that of genetically modified MSCs alone. Moreover, the expression of collagen type II was more than 3.5 times greater than that of genetically modified MSCs alone. 3-D hydrogel constructs co-cultured with chondrocytes and genetically modified MSCs showed a significantly higher number of specific lacunae phenotypes at the end of the 4 week study, regardless of whether they were co-cultured in the presence of chondrocytes. These findings indicate that co-culture with chondrocytes and genetically modified MSCs can be used to engineer well designed implants for the formation of neocartilage by transplanted genetically modified MSCs.