A Unique strategy for probing in-situ NO for screening the neuroinflammatory phenotypes against SARS-CoV-2 RNA in phagocytotic microglia

The severe acute respiratory (SARS-CoV-2) jeopardized public health by causing significant morbidity and mortality among people with pre-existing physiological dysfunction caused by aging, diabetes, hypertension, and obesity. Besides, patients with severe infections are more likely to have neuro-inflammation and higher mortality risk. Neuroinflammation is majorly caused by activating the brain’s residential macrophage cells called microglia, which increases the generation of reactive species including nitric oxide via the iNOS pathway. In addition, NO regulates lysosomal functions and exhibits complex effects on lysosomal machinery to neutralize foreign pathogens through phagocytosis and improve host-defense inflammatory response. To date, lack of efficient probes to monitor lysosomal NO and phagocytosis processes in the least explored human microglia during SARS-CoV-2 infection. Herein, a unique design strategy was adopted for the first time by avoiding the conventional control amination reaction approach to develop lysosomal specific-NO probe, PDM-NO, which can discriminate activated microglia from its resting state. The non-fluorescent probe at physiological pH exhibits turn-on response towards NO only at lysosomal pH (4.5-5.5). PDM-NO demonstrated lysosomal specificity in activated HMC3 cells and enabled monitoring phagocytosis process with a several advantages compared to commercial E. coli bio-particles. Moreover, this probe can effectively map the overexpression and dynamics of lysosomal NO levels against SARS-CoV-2 RNA virus-induced neuroinflammation in HMC3. Thus, lysosome-specific PDM-NO is a potential fluorescent marker for detecting RNA virus infection in human microglia and excellent molecular probe for monitoring phagocytosis during neuroinflammation. It could be a useful commercial probe in future for screening viral activity and neuro-inflammation for diagnosis of neurological diseases.