To reduce the pain caused by subcutaneous injections, microneedle patches as the new transdermal drug delivery method are gaining increased attention. In this study, we fabricated a composite insulin-loaded microneedle patch. Silk fibroin, a natural polymer material, was used as the raw material. The tip of the microneedle had good dissolving property and was able to dissolve rapidly to promote the release of insulin. The pedestal had the property of swelling without dissolving and was carrying insulin as a drug store. The insulin carried by the pedestal could release continuously through the micropore channels created by the microneedles. This kind of microneedle could achieve a sustained release effect. It was observed that the insulin had good storage stability in this kind of microneedle, and it maintained more than 90% of its biological activity after 30 days. The results of transdermal delivery to diabetic rats showed that the microneedle patches displayed an apparent hypoglycemic effect and indicated a sustained release effect. These drug-loaded silk microneedle patches may act as potential delivery systems for the treatment of diabetes.

Background Toxoplasma gondii is a zoonotic pathogen that causes toxoplasmosis and leads to serious public health problems in developing countries. However, current clinical therapeutic drugs have some disadvantages, such as serious side effects, a long course of treatment and the emergence of drug-resistant strains. The urgent need to identify novel anti-Toxoplasma drugs has initiated the effective strategy of repurposing well-characterized drugs. As a principled screening for the identification of effective compounds against Toxoplasma gondii, in the current study, a collection of 666 compounds were screened for their ability to significantly inhibit Toxoplasma growth. Methods The inhibition of parasite growth was determined using a luminescence-based β-galactosidase activity assay. Meanwhile, the effect of compounds on the viability of host cells was measured using CCK8. To assess the inhibition of the selected compounds on discrete steps of the T. gondii lytic cycle, the invasion, intracellular proliferation and egress abilities were evaluated. Finally, a murine infection model of toxoplasmosis was used to monitor the protective efficacy of drugs against acute infection of a highly virulent RH strain. Results A total of 68 compounds demonstrated more than 70% parasite growth inhibition. After excluding compounds that impaired host cell viability, we further characterized two compounds, NVP-AEW541 and GSK-J4 HCl, which had IC50 values for parasite growth of 1.17 μM and 2.37 μM, respectively. In addition, both compounds showed low toxicity to the host cell. Furthermore, we demonstrated that NVP-AEW541 inhibits tachyzoite invasion, while GSK-J4 HCl inhibits intracellular tachyzoite proliferation by halting cell cycle progression from G1 to S phase. These findings prompted us to analyse the efficacy of the two compounds in vivo by using established mouse models of acute toxoplasmosis. In addition to prolonging the survival time of mice acutely infected with T. gondii, both compounds had a remarkable ability to reduce the parasite burden of tissues. Conclusions Our findings suggest that both NVP-AEW541 and GSK-J4 could be potentially repurposed as candidate drugs against T. gondii infection.

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Toxoplasmosis is caused by an obligate intracellular parasite, the protozoan Discovery of novel drugs against infection could circumvent the toxicity of existing drugs and resistance to current treatments. The autophagy-related protein 8 (Atg8)-Atg3 interaction in is a promising drug target because of its importance for regulating Atg8 lipidation. We reported previously that TgAtg8 and TgAtg3 interact directly. Here we validated that substitutions of conserved residues of TgAtg8 interacting with the Atg8 family-interacting motif (AIM) in Atg3 disrupt the TgAtg8-TgAtg3 interaction and reduce TgAtg8 lipidation and autophagosome formation. These findings were consistent with results reported previously for Atg8, suggesting functional conservation of Atg8 in and Moreover, using peptide and AlphaScreen assays, we identified the AIM sequence in TgAtg3 that binds TgAtg8. We determined that the core TgAtg3 AIM contains a Phe-Ala-Asp-Ile (FADI) signature distinct from the WLLP signature in the AIM of Atg3. Furthermore, an alanine-scanning assay revealed that the TgAtg8-TgAtg3 interaction in also depends strongly on several residues surrounding the core TgAtg3 AIM, such as Asn, Asp, and Cys These results indicate that distinct AIMs in Atg3 contribute to differences between and Atg8-Atg3 interactions. By elucidating critical residues involved in the TgAtg8-TgAtg3 interaction, our work paves the way for the discovery of potential anti-toxoplasmosis drugs. The quantitative and straightforward AlphaScreen assay developed here may enable high-throughput screening for small molecules disrupting the TgAtg8-TgAtg3 interaction.

As a patient-friendly technology, drug-loaded microneedles can deliver drugs through the skin into the body. This system has broad application prospects and is receiving wide attention. Based on the knowledge acquired in this work, we successfully developed a melatonin-loaded microneedle prepared from proline/melatonin/silk fibroin. The engineered microneedles’ morphological, physical, and chemical properties were characterized to investigate their structural transformation mechanism and transdermal drug-delivery capabilities. The results indicated that the crystal structure of silk fibroin in drug-loaded microneedles was mainly Silk I crystal structure, with a low dissolution rate and suitable swelling property. Melatonin-loaded microneedles showed high mechanical properties, and the breaking strength of a single needle was 1.2 N, which could easily be penetrated the skin. The drug release results in vitro revealed that the effective drug concentration was obtained quickly during the early delivery. The successful drug concentration was maintained through continuous release at the later stage. For in vivo experimentation, the Sprague Dawley (SD) rat model of insomnia was constructed. The outcome exhibited that the melatonin-loaded microneedle released the drug into the body through the skin and maintained a high blood concentration (over 5 ng/mL) for 4–6 h. The maximum blood concentration was above 10 ng/mL, and the peak time was 0.31 h. This system indicates that it achieved the purpose of mimicking physiological release and treating insomnia.