repsox and Disease-Models--Animal

Epithelial-mesenchymal transition (EMT) of retinal pigment epithelial (RPE) cells is a key pathological event in proliferative retinal diseases such as proliferative vitreoretinopathy (PVR). This study aimed to explore a new method to reverse EMT in RPE cells to develop an improved therapy for proliferative retinal diseases.. In vitro, human embryonic stem cell-derived RPE cells were passaged and cultured at low density for an extended period of time to establish an EMT model. At different stages of EMT after treatment with known molecules or combinations of molecules, the morphology was examined, transepithelial electrical resistance (TER) was measured, and expression of RPE- and EMT-related genes were examined with RT-PCR, Western blotting, and immunofluorescence. In vivo, a rat model of EMT in RPE cells was established via subretinal injection of dispase. Retinal function was examined by electroretinography (ERG), and retinal morphology was examined.. EMT of RPE cells was effectively induced by prolonged low-density culture. After EMT occurred, only the combination of the Rho-associated coiled-coil containing protein kinase (ROCK) inhibitor Y27632 and the TGF-β receptor inhibitor RepSox (RY treatment) effectively suppressed and reversed the EMT process, even in cells in an intermediate state of EMT. In dispase-treated Sprague-Dawley rats, RY treatment maintained the morphology of RPE cells and the retina and preserved retinal function.. RY treatment might promote mesenchymal-epithelial transition (MET), the inverse process of EMT, to maintain the epithelial-like morphology and function of RPE cells. This combined RY therapy could be a new strategy for treating proliferative retinal diseases, especially those involving EMT of RPE cells.

Topics: Amides; Animals; Cells, Cultured; Disease Models, Animal; Enzyme Inhibitors; Epithelial-Mesenchymal Transition; Humans; Mice; Pyrazoles; Pyridines; Rats; Rats, Sprague-Dawley; Retinal Pigment Epithelium; Transforming Growth Factor beta1; Vitreoretinopathy, Proliferative

When Zika virus emerged as a public health emergency there were no drugs or vaccines approved for its prevention or treatment. We used a high-throughput screen for Zika virus protease inhibitors to identify several inhibitors of Zika virus infection. We expressed the NS2B-NS3 Zika virus protease and conducted a biochemical screen for small-molecule inhibitors. A quantitative structure-activity relationship model was employed to virtually screen ∼138,000 compounds, which increased the identification of active compounds, while decreasing screening time and resources. Candidate inhibitors were validated in several viral infection assays. Small molecules with favorable clinical profiles, especially the five-lipoxygenase-activating protein inhibitor, MK-591, inhibited the Zika virus protease and infection in neural stem cells. Members of the tetracycline family of antibiotics were more potent inhibitors of Zika virus infection than the protease, suggesting they may have multiple mechanisms of action. The most potent tetracycline, methacycline, reduced the amount of Zika virus present in the brain and the severity of Zika virus-induced motor deficits in an immunocompetent mouse model. As Food and Drug Administration-approved drugs, the tetracyclines could be quickly translated to the clinic. The compounds identified through our screening paradigm have the potential to be used as prophylactics for patients traveling to endemic regions or for the treatment of the neurological complications of Zika virus infection.

Topics: Animals; Antiviral Agents; Artificial Intelligence; Chlorocebus aethiops; Disease Models, Animal; Drug Evaluation, Preclinical; High-Throughput Screening Assays; Immunocompetence; Inhibitory Concentration 50; Methacycline; Mice, Inbred C57BL; Protease Inhibitors; Quantitative Structure-Activity Relationship; Small Molecule Libraries; Vero Cells; Zika Virus; Zika Virus Infection

Cultured epidermal autografts have been used worldwide since 1981 for patients with extensive third-degree burn wounds and limited skin donor sites. Despite significant progress in techniques toward improving clinical outcome of skin grafts, the long in vitro preparation time of cultured autografts has remained a major factor limiting its widespread use. Here, we show that pharmacological inhibition of TGF-β signaling promotes the expansion of human epidermal keratinocytes (HEKs) with high proliferative potential in co-cultures with both murine 3T3-J2 cells and human feeder cells, including dermal fibroblasts and preadipocytes. In contrast, TGF-β signaling inhibition does not enhance the growth of HEKs in a serum- and feeder-free condition, an alternative approach to propagate HEKs for subsequent autograft production. Our results have important implications for the use of TGF-β signaling inhibition as a viable therapeutic strategy for improving Green's methodology and for more efficient production of customized skin autografts with human feeder cells.

Topics: 3T3 Cells; Animals; Coculture Techniques; Disease Models, Animal; Epidermal Cells; Epidermis; Feeder Cells; Humans; Keratinocytes; Mice; Pyrazoles; Pyridines; Signal Transduction; Skin Transplantation; Transforming Growth Factor beta