CRISPR-Cas9/gRNA exhibits therapeutic efficacy against latent human immunodeficiency virus (HIV) genome but the delivery of this therapeutic cargo to the brain remains as a challenge. In this research, for the first time, we demonstrated magnetically guided non-invasive delivery of a nano-formulation (NF), composed of Cas9/gRNA bound with magneto-electric nanoparticles (MENPs), across the blood-brain barrier (BBB) to inhibit latent HIV-1 infection in microglial (hμglia)/HIV (HC69) cells. An optimized ac-magnetic field of 60 Oe was applied on NF to release Cas9/gRNA from MENPs surface and to facilitate NF cell uptake resulting in intracellular release and inhibition of HIV. The outcomes suggested that developed NF reduced HIV-LTR expression significantly in comparison to unbound Cas9/gRNA in HIV latent hμglia/HIV (HC69) cells. These findings were also validated qualitatively using fluorescence microscopy to assess NF efficacy against latent HIV in the microglia cells. We believe that CNS delivery of NF (CRISPR/Cas9-gRNA-MENPs) across the BBB certainly will have clinical utility as future personalized nanomedicine to manage neuroHIV/AIDS.

In order to establish infection, intra-macrophage parasite Leishmania donovani needs to inhibit host defense parameters like inflammatory cytokine production and apoptosis. In the present study, we demonstrate that the parasite achieves both by exploiting a single host regulator AKT for modulating its downstream transcription factors, β-catenin and FOXO-1. L. donovaniinfected RAW264.7 and bone marrow-derived macrophages (BMDM) treated with AKT inhibitor or dominant negative AKT constructs showed decreased anti-inflammatory cytokine production and increased host cell apoptosis resulting in reduced parasite survival. Infection-induced activated AKT triggered phosphorylation-mediated deactivation of its downstream target, GSK-3β. Inactivated GSK-3β, in turn, could no longer sequester cytosolic β-catenin, an anti-apoptotic transcriptional regulator, as evidenced from its nuclear translocation during infection. Constitutively active GSK-3β-transfected L. donovani-infected cells mimicked the effects of AKT inhibition and siRNA-mediated silencing of β-catenin led to disruption of mitochondrial potential along with increased caspase-3 activity and IL-12 production leading to decreased parasite survival. In addition to activating antiapoptotic β-catenin, phospho-AKT inhibits activation of FOXO-1, a pro-apoptotic transcriptional regulator. Nuclear retention of FOXO-1, inhibited during infection, was reversed when infected cells were transfected with dominant negative AKT constructs. Overexpression of FOXO-1 in infected macrophages not only documented increased apoptosis but promoted enhanced TLR4 expression and NF-κB activity along with an increase in IL-1β and decrease in IL-10 secretion. In vivo administration of AKT inhibitor significantly decreased liver and spleen parasite burden and switched cytokine balance in favor of host. In contrast, GSK-3β inhibitor did not result in any significant change in infectivity parameters. Collectively our findings revealed that L. donovani triggered AKT activation to regulate GSK-3β/β-catenin/FOXO-1 axis, thus ensuring inhibition of both host cell apoptosis and immune response essential for its intra-macrophage survival.

This study reports the self-assembly and application of a naphthalene diimide (NDI)-appended peptide amphiphile (PA). H-bonding among the peptide moiety in conjunction with π-stacking between NDI and hydrophobic interactions within the alkyl chain are the major driving forces behind the stepwise aggregation of the PA to form hydrogels. The PA produced efficient self-assemblies in water, forming a nanofibrous network that further formed a self-supportive hydrogel. The molecule followed a three-step self-assembly mechanism. At a lower concentration (50 μM), it forms extremely small aggregates with a very low population of the molecules. With an increase in concentration, spherical aggregates are formed above 450 μM concentration. Importantly, this water-soluble conjugate was found to be nontoxic, cell permeable, and usable for cell imaging. Moreover, the aggregation process and consequently the emission behavior are highly responsive to the pH of the medium. Thus, the pH responsive aggregation and emission behavior has an extended biological application for assessing intracellular pH. The biocompatibility and intracellular pH determining capability suggest it is a promising candidate for use as a supramolecular material in biomedical applications.

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Background: Leishmania inhibits oxidative burst-mediated apoptosis of macrophages during phagocytosis. Results: L. donovani induces (SOCS) 1 and 3, which suppress macrophage apoptosis through thioredoxin-mediated stabilization of protein-tyrosine phosphatases. Conclusion: Leishmania exploits macrophage SOCS proteins for inhibition of apoptosis, thus protecting its niche for survival and replication. Significance: This study demonstrates a novel anti-apoptotic mediator for parasite infection.