Cytotoxicity assessment, inflammatory properties, and cellular uptake of Neutraplex lipid-based nanoparticles in THP-1 monocyte-derived macrophages.

Current antiretroviral drugs used to prevent or treat human immunodeficiency virus type 1 (HIV-1) infection are not able to eliminate the virus within tissues or cells where HIV establishes reservoirs. Hence, there is an urgent need to develop targeted delivery systems to enhance drug concentrations in these viral sanctuary sites. Macrophages are key players in HIV infection and contribute significantly to the cellular reservoirs of HIV because the virus can survive for prolonged periods in these cells. In the present work, we investigated the potential of the lipid-based Neutraplex nanosystem to deliver anti-HIV therapeutics in human macrophages using the human monocyte/macrophage cell line THP-1. Neutraplex nanoparticles as well as cationic and anionic Neutraplex nanolipoplexes (Neutraplex/small interfering RNA) were prepared and characterized by dynamic light scattering. Neutraplex nanoparticles showed low cytotoxicity in CellTiter-Blue reduction and lactate dehydrogenase release assays and were not found to have pro-inflammatory effects. In addition, confocal studies showed that the Neutraplex nanoparticles and nanolipoplexes are rapidly internalized into THP-1 macrophages and that they can escape the late endosome/lysosome compartment allowing the delivery of small interfering RNAs in the cytoplasm. Furthermore, HIV replication was inhibited in the in vitro TZM-bl infectivity assay when small interfering RNAs targeting CXCR4 co-receptor was delivered by Neutraplex nanoparticles compared to a random small interfering RNA sequence. This study demonstrates that the Neutraplex nanosystem has potential for further development as a delivery strategy to efficiently and safely enhance the transport of therapeutic molecules into human monocyte-derived macrophages in the aim of targeting HIV-1 in this cellular reservoir.

Preparation, characterization, and safety evaluation of poly(lactide-co-glycolide) nanoparticles for protein delivery into macrophages.

Following infection, HIV establishes reservoirs within tissues that are inaccessible to optimal levels of antiviral drugs or within cells where HIV lies latent, thus escaping the action of anti-HIV drugs. Macrophages are a persistent reservoir for HIV and may contribute to the rebound viremia observed after antiretroviral treatment is stopped. In this study, we further investigate the potential of poly(lactic-co-glycolic) acid (PLGA)-based nanocarriers as a new strategy to enhance penetration of therapeutic molecules into macrophages. We have prepared stable PLGA nanoparticles (NPs) and evaluated their capacity to transport an active molecule into the human monocyte/macrophage cell line THP-1 using bovine serum albumin (BSA) as a proof-of-concept compound. Intracellular localization of fluorescent BSA molecules encapsulated into PLGA NPs was monitored in live cells using confocal microscopy, and cellular uptake was quantified by flow cytometry. In vitro and in vivo toxicological studies were performed to further determine the safety profile of PLGA NPs including inflammatory effects. The size of the PLGA NPs carrying BSA (PLGA-BSA) in culture medium containing 10% serum was ~126 nm in diameter, and they were negatively charged at their surface (zeta potential =-5.6 mV). Our confocal microscopy studies and flow cytometry data showed that these PLGA-BSA NPs are rapidly and efficiently taken up by THP-1 monocyte-derived macrophages (MDMs) at low doses. We found that PLGA-BSA NPs increased cellular uptake and internalization of the protein in vitro. PLGA NPs were not cytotoxic for THP-1 MDM cells, did not modulate neutrophil apoptosis in vitro, and did not show inflammatory effect in vivo in the murine air pouch model of acute inflammation. In contrast to BSA alone, BSA encapsulated into PLGA NPs increased leukocyte infiltration in vivo, suggesting the in vivo enhanced delivery and protection of the protein by the polymer nanocarrier. We demonstrated that PLGA-based nanopolymer carriers are good candidates to efficiently and safely enhance the transport of active molecules into human MDMs. In addition, we further investigated their inflammatory profile and showed that PLGA NPs have low inflammatory effects in vitro and in vivo. Thus, PLGA nanocarriers are promising as a drug delivery strategy in macrophages for prevention and eradication of intracellular pathogens such as HIV and Mycobacterium tuberculosis.

The effects of in utero and lactational exposure to chloroform on postnatal growth and glucose tolerance in male Wistar rats.

Water chlorination results in the formation of trihalomethanes (THMs) including chloroform. In human stud-ies, fetal growth restriction has been associated with exposure to THMs during pregnancy and impaired fetal growth has been associated with an increased risk of type 2 diabetes. Therefore, the objective of this study was to determine the effect of in utero and lactational exposure to chloroform on birthweight and postnatal indicators of type 2 diabetes. Female Wistar rats were given chloroform (0 microg/L, 75 microg/L) in their drinking water for 2 wk prior to mating until parturition (in utero exposure only) or until weaning (in utero+lactational exposure). At postnatal d 1 (PND1) pups of dams exposed to chloroform had significantly higher serum glucose levels and lower insulin levels, but this effect was not due to b-cell depletion in the neonatal pancreas. Glucose homeostasis in response to a glucose challenge was not changed by chloroform treatment. Chloroform exposure did not affect birthweight; however, offspring of dams exposed to chloroform had significantly impaired postnatal growth. Although fetal and neonatal exposure to chloroform did not elicit physiological changes associated with the onset of type 2 diabetes, there were physiological changes resulting in impaired postnatal growth.