hTERT-immortalized mesenchymal stem cell-derived EV treatment reduces ZIKV-induced cortical neuronal death, infection, and exosome-mediated transmission

The application of stem cells, mesenchymal stem cells (MSCs), and exosomes/extracellular vesicles (EVs) derived from these stem cells has shown potential advancement in solving several major medical complications. Recent studies on MSC-EVs have gained focus on tissue repair, inflammation, apoptosis, fibrosis, and regenerative medicine. MSC-EVs modulate and play potential therapeutic roles in the treatment of various diseases such as cancer, neurological disorders, and viral infections. In the current study, we provide evidence that human telomerase reverse transcriptase (hTERT)-immortalized MSC-EVs possess a neuroprotective role in Zika virus (ZIKV)-infected murine cortical neurons. The American Type Culture Collection (ATCC) 2022 challenge award (call for proposals to use the product "ATCC SCRC-4000-EXM," hTERT-immortalized MSC-EVs in new, interesting, or daring applications) afforded us the opportunity to test these hTERT-MSC-EV in ZIKV-infected neurons. We found that hTERT-MSC-EV treatment increased the cell viability of ZIKV-infected neurons and decreased the expression of apoptotic markers (Bax, caspases 3/9, and Bcl-2). In addition, hTERT-MSC-EV treatment reduced ZIKV loads in cortical neurons that correlated with increased interferon-beta and decreased TNF-alpha transcripts. Poly I:C (polyinosinic:polycytidylic acid) stimulation and treatment of uninfected cortical neurons with hTERT-MSC-EV showed an increased trend in IFN-alpha levels, but IFN-beta transcript levels were reduced. Furthermore, hTERT-MSC-EV treatment continued to reduce viral loads, even upon Poly I:C stimulation and ZIKV infection. GW4869 inhibition followed by hTERT-MSC-EV treatment revealed that exosome-mediated viral transmission is also affected in ZIKV-infected cortical neurons. Overall, this study suggests a potential therapeutic effect of hTERT-MSC-EV in ZIKV-induced neuronal death and perhaps in controlling exosome biogenesis during viral infections. This study opens new thoughts on how hTERT-MSC-EV could be proposed as a therapeutic potential in ZIKV-mediated microcephaly by enhancing cell viability of ZIKV-infected cortical neurons, inhibiting apoptosis, and reducing viral infection and exosome-mediated transmission in fetal and newborn brains. IMPORTANCE: Zika virus (ZIKV), a mosquito-borne flavivirus that vertically transmits from pregnant women to her fetus, causes microcephaly, a birth defect where newborns show smaller head circumference and brain size compared to normal healthy babies. Microcephalic newborns exhibit several developmental delays and neurological complications. There is no cure or potential therapeutics available to treat ZIKV-caused microcephaly. Human telomerase reverse transcriptase (hTERT)-immortalized mesenchymal stem cell (MSC)-derived extracellular vesicles (MSC-EVs) have been recently proposed as potential therapeutics in the treatment of various diseases, including cancer, neurological disorders, and viral infections. We hypothesized an important role of hTERT MSC-EVs in providing neuroprotective effects upon ZIKV infection in murine cortical neurons. Our study opens new thoughts on how hTERT-MSC-EV could be proposed as potential therapeutics in ZIKV-caused microcephaly, where they enhance cell viability, inhibit apoptosis, and reduce viral infection and exosome-mediated transmission/dissemination of ZIKV-infected murine cortical neurons in embryonic brains.