This study investigated the effect of nanoparticle size (50 and 100 nm) and surface charge on their interaction with Caco-2 monolayers as a model of the intestinal epithelium, including cell internalization pathways and the level of transepithelial transport. Initially, toxicity assays showed that cell viability and cell membrane integrity were dependent on the surface charge and applied mass, number, and total surface area of nanoparticles, as tested in two epithelial cell lines, colon carcinoma Caco-2 and airway Calu-3. This also identified suitable nanoparticle concentrations for subsequent cell uptake experiments. Nanoparticle application at doses below half maximal effective concentration (EC₅₀) revealed that the transport efficiency (ratio of transport to cell uptake) across Caco-2 cell monolayers is significantly higher for negatively charged nanoparticles compared to their positively charged counterparts (of similar size), despite the higher level of internalization of positively charged systems. Cell internalization pathways were hence probed using a panel of pharmacological inhibitors aiming to establish whether the discrepancy in transport efficiency is due to different uptake and transport pathways. Vesicular trans-monolayer transport for both positively and negatively charged nanoparticles was confirmed via inhibition of dynamin (by dynasore) and microtubule network (via nocodazole), which significantly reduced the transport of both nanoparticle systems. For positively charged nanoparticles a significant decrease in internalization and transport (46% and 37%, respectively) occurred in the presence of a clathrin pathway inhibitor (chlorpromazine), macropinocytosis inhibition (42%; achieved by 5-(N-ethyl-N-isopropyi)-amiloride), and under cholesterol depletion (38%; via methyl-β-cyclodextrin), but remained unaffected by the inhibition of lipid raft associated uptake (caveolae) by genistein. On the contrary, the most prominent reduction in internalization and transport of negatively charged nanoparticles (51% and 48%, respectively) followed the inhibition of lipid raft-associated pathway (caveolae inhibition by genistein) but was not significantly affected by the inhibition of clathrin pathway.
Despite the development of progressive policies in forestry and wildlife management sectors in Nepal, over the last ten years it has become clear that protected areas in the country lack the landscape connectivity required to support viable populations of endangered species such as rhino (Rhinoceros unicornis) and tiger (Panthera tigris tigris). Forest resources that provide essential habitat for a diverse array of species, and forest products to many human populations continue to decline. The present study, focusing upon the northern Barandabhar Forest Corridor (BFC) aimed to begin to address these problems by establishing preliminary data concerning the extent, biological diversity and resources available for local communities of the forest in order to promote a community-based management strategy. Results established that the northern BFC covers an area of 10,644 ha between the East-West Mahendra highway and the Mahabharat range, incorporating 15 community forestry areas (3,184 ha). It was found to support a number of IUCN listed threatened and endangered species including rhino and tiger; and contains suitable habitat for a number of other endangered species. Owing to its connectivity, linking Chitwan National Park with the Mahabharat range, BFC has the potential to make an important contribution to improving the ecological integrity in Nepal. Different institutions and governance structures currently exist to manage the northern BFC. Although these institutions differ in resource utilization and benefit sharing mechanisms, they are united to safeguard the habitat of key wildlife species including rhino and tiger. We propose that the northern BFC should be managed through a new participatory scheme, the Barandabhar Forest Management Council, to foster ecological integrity of the area while providing forest products to communities.
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