Biomedical subjects

Youjun Wu

Publications and source records attributed to Youjun Wu.

3 recordsLinked to original sources

PubMed2026

Suppression of AAV-Delivered Transgene Expression Using Artificial MicroRNAs Delivered by an Alternative AAV Serotype.

Adeno-associated virus (AAV) gene transfer vectors mediate long-term expression in nondividing cells, an advantage for treating chronic disorders. However, current platforms lack a way to selectively shut down transgene expression if adverse effects arise. To create an "off switch," we hypothesized that incorporating unique artificial microRNA (amiRNA) target sequences into an AAV expression cassette would allow subsequent suppression of transgene expression using a second AAV vector encoding the cognate amiRNA. We introduced 22-nt sequences absent from human and mouse transcriptomes into the 3' untranslated region (UTR) of a therapeutic AAV cassette. To identify optimal amiRNAs, two tandem copies of each amiRNA were cloned into the 3'UTR of an mCherry reporter gene. In vitro assessment of six amiRNA/target pairs using a dual luciferase assay identified four amiRNAs that efficiently suppressed reporter expression. Cells cotransfected with target site 3 (TS3) and amiRNA-T3B showed the greatest reduction in luciferase activity (80%, p < 0.0001) and were selected for further study. The "off-switch" system was then evaluated using an AAV5 therapeutic vector expressing a recombinant humanized anti-IgE monoclonal antibody (AAV5-TBG-anti-IgE-TS3), designed for long-term suppression of allergen-induced reactions. Co-transfection of HEK293T cells with anti-IgE-TS3 and amiRNA-T3B significantly reduced anti-IgE mRNA and protein levels relative to a control amiRNA (p < 0.0001). In vivo testing in Balb/c mice (n = 5) involved intravenous administration of AAV5-anti-IgE-TS3 (3.2 × 1010 gc), followed 4 weeks later by an AAVrh.10 amiRNA vector (AAVrh.10-TBG-amiRNA-T3B; 1 × 1011 gc). Control mice receiving only the therapeutic vector expressed 18.4 ± 13.8 µg/mL serum anti-IgE at 10 weeks. In contrast, mice receiving the amiRNA "off" vector showed marked suppression of anti-IgE (0.3 ± 0.15 µg/mL, p < 0.0001). These findings provide proof-of-concept that AAV-delivered amiRNAs can selectively switch off transgene expression, offering a strategy to improve the safety of AAV-mediated gene therapies.

Dependovirus↗

PubMed2025

Transcriptional signature of induced neurons differentiates virologically suppressed people with HIV from people without HIV.

Neurocognitive impairment is a prevalent comorbidity in virologically suppressed people living with HIV (PLWH), yet the underlying mechanisms remain elusive and treatments lacking. We explored use of participant-derived directly induced neurons (iNs) to model neuronal biology and injury in PLWH. iNs retain age- and disease-related donor features, providing unique opportunities to reveal important aspects of neurological disorders. We obtained primary dermal fibroblasts from 6 virologically suppressed PLWH and 7 matched people without HIV (PWOH). iNs were generated using transcription factors NGN2 and ASCL1 and validated by immunocytochemistry, single-cell RNA-Seq, and electrophysiological recordings. Transcriptomic aging analyses confirmed retention of donor age-related signatures. Bulk RNA-Seq identified 29 significantly differentially expressed genes between PLWH and PWOH iNs. Of these, 16 were downregulated and 13 upregulated in PLWH iNs. Protein-protein interaction network mapping indicated iNs from PLWH exhibited differences in extracellular matrix organization and synaptic transmission. IFI27 was upregulated in PLWH iNs, complementing independent postmortem studies demonstrating elevated IFI27 expression in PLWH-derived brain tissue. FOXL2NB-FOXL2-LINC01391 expression was reduced in PLWH iNs and negatively correlated with neurocognitive impairment. Thus, we identified an iN gene signature of HIV revealing mechanisms of neurocognitive impairment in PLWH.

Humans↗

PubMed2024

A Transcriptional Signature of Induced Neurons Differentiates Virologically Suppressed People Living With HIV from People Without HIV.

Neurocognitive impairment is a prevalent and important co-morbidity in virologically suppressed people living with HIV (PLWH), yet the underlying mechanisms remain elusive and treatments lacking. Here, we explored for the first time, use of participant-derived directly induced neurons (iNs) to model neuronal biology and injury in PLWH. iNs retain age- and disease-related features of the donors, providing unique opportunities to reveal novel aspects of neurological disorders. We obtained primary dermal fibroblasts from six virologically suppressed PLWH (range: 27 - 64 years, median: 53); 83% Male; 50% White) and seven matched people without HIV (PWOH) (range: 27 - 66, median: 55); 71% Male; 57% White). iNs were generated using transcription factors NGN2 and ASCL1, and validated by immunocytochemistry and single-cell-RNAseq. Transcriptomic analysis using bulk-RNAseq identified 29 significantly differentially expressed genes between iNs from PLWH and PWOH. Of these, 16 genes were downregulated and 13 upregulated in PLWH iNs. Protein-protein interaction network mapping indicates that iNs from PLWH exhibit differences in extracellular matrix organization and synaptic transmission. IFI27 was upregulated in iNs from PLWH, which complements independent post-mortem studies demonstrating elevated IFI27 expression in PLWH-derived brain tissue, indicating that iN generation reconstitutes this pathway. Finally, we observed that expression of the FOXL2NB-FOXL2-LINC01391 genome locus is reduced in iNs from PLWH and negatively correlates with neurocognitive impairment. Thus, we have identified an iN gene signature of HIV through direct reprogramming of skin fibroblasts into neurons revealing novel mechanisms of neurocognitive impairment in PLWH.

HAND↗