For patients with proliferative diabetic retinopathy (PDR) who do not respond adequately to pan-retinal laser photocoagulation (PRP) or anti–vascular endothelial growth factor (VEGF) therapies, we hypothesized that vascular cells within neovascular tissue secrete autocrine/paracrine angiogenic factors that promote disease progression. To identify these factors, we performed multiplex ELISA angiogenesis arrays on aqueous fluid from PDR patients who responded inadequately to anti-VEGF therapy and/or PRP and identified plasminogen activator inhibitor-1 (PAI-1). PAI-1 expression was increased in vitreous biopsies and neovascular tissue from PDR eyes, limited to retinal vascular cells, regulated by the transcription factor hypoxia-inducible factor (HIF)-2α, and necessary and sufficient to stimulate angiogenesis. Using a pharmacologic inhibitor of HIF-2α (PT-2385) or nanoparticle-mediated RNA interference targeting Pai1 , we demonstrate that the HIF-2α/PAI-1 axis is necessary for the development of retinal neovascularization in mice. These results suggest that targeting HIF-2α/PAI-1 will be an effective adjunct therapy for the treatment of PDR patients.

Most patients with neovascular age-related macular degeneration (nvAMD), the leading cause of severe vision loss in elderly US citizens, respond inadequately to current therapies targeting a single angiogenic mediator, vascular endothelial growth factor (VEGF). Here, we report that aqueous fluid levels of a second vasoactive mediator, angiopoietin-like 4 (ANGPTL4), can help predict the response of patients with nvAMD to anti-VEGF therapies. ANGPTL4 expression was higher in patients who required monthly treatment with anti-VEGF therapies compared with patients who could be effectively treated with less-frequent injections. We further demonstrate that ANGPTL4 acts synergistically with VEGF to promote the growth and leakage of choroidal neovascular (CNV) lesions in mice. Targeting ANGPTL4 expression was as effective as targeting VEGF expression for treating CNV in mice, while simultaneously targeting both was more effective than targeting either factor alone. To help translate these findings to patients, we used a soluble receptor that binds to both VEGF and ANGPTL4 and effectively inhibited the development of CNV lesions in mice. Our findings provide an assay that can help predict the response of patients with nvAMD to anti-VEGF monotherapy and suggest that therapies targeting both ANGPTL4 and VEGF will be a more effective approach for the treatment of this blinding disease.

Wiskott Aldrich Syndrome protein (WASP) has a unique regulatory role in sealing ring formation and bone resorption in osteoclasts. Here, using the TAT-transduction method, we show the possible role of WASP domain(s) in sealing ring formation and bone resorption. Transduction of TAT-fused full-length WASP peptide induced Arp2/3 complex formation, F-actin content, sealing ring formation and bone resorption. Transduction of WASP peptides containing basic, verpolin-central, pTyr294, and proline-rich regions inhibited the processes listed above at various levels. The ability to resorb bone by WASP peptides containing basic, verpolin-central, and proline-rich regions was reduced and the resorbed area matched the size of the sealing ring. However, osteoclasts transduced with WASP peptide containing pTyr294aa demonstrated the following: a) a considerable decrease in the interaction and phosphorylation of c-Src with endogenous WASP; b) total loss of sealing ring-like structures; c) formation of actin-rich patches at the peripheral edge that contains filopodia-like projections; d) reduced capacity for bone resorption in vitro. These findings suggest that modulation of phosphorylation state of pTyr294aa assists in integrating multiple signaling molecule and pathways that partake in the assembly of sealing ring.

Traditional pulp-capping materials like mineral trioxide aggregate (MTA) offer excellent biocompatibility and sealing, but limitations such as prolonged setting time, low bioactivity, and high costs persist. Metformin, with its potential in craniofacial regeneration, could enhance dentin synthesis by targeting pulp cells. This study aimed to: (1) develop a calcium phosphate cement with chitosan (CPCC) with improved physio-mechanical properties; (2) incorporate metformin (CPCC-Met) to assess release; and (3) evaluate human dental pulp stem cells (hDPSCs) response. CPCC was mixed at different powder-to-liquid ratios to evaluate physio-mechanical properties compared to MTA. The optimized CPCC formulation was loaded with 0, 50, 100, and 150 µg of metformin to measure release and assess hDPSCs attachment and proliferation (1, 4, and 7 d) via live/dead imaging and SEM. One-way ANOVA was used for statistical analysis. Results showed CPCC at a 3.25:1 ratio significantly reduced setting time to 41.5 min versus 123 min for MTA (p < 0.05). Metformin release correlated with concentration, and SEM confirmed the presence of a porous, hydroxyapatite-rich surface. Cell viability was consistently high across groups (>93% at 1 d, >95% at 4 d, ≈98% at 7 d), with no significant differences (p > 0.05). These findings suggest that the novel CPCC-Met demonstrates promise as a fast-setting, cost-effective pulp-capping material, offering metformin delivery to enhance dentin repair.