ABSTRACT
Oleanolic acid (OA), a biologically active pentacyclic triterpenoid compound, has been implicated in a number of clinical benefits including antioxidant, and anti-inflammatory properties. OA has been previously shown to ameliorate the toxic effects of 6-hydroxydopamine (6-OHDA), however, the mechanism by which this effect is exhibited is not clearly understood. In the present study, we investigated the role of OA in attenuation of microglial activation in 6-OHDA induced Parkinsonian rat model. We also explored the ability of OA to attenuate 6-OHDA-induced intracellular reactive oxygen species (ROS), and thus prevent cell death in PC12 cells. We accessed the utility of immunohistochemistry to assess striatal microglial activation, where shape descriptors such as area, perimeter, Feret's diameter, aspect ratio and solidity were determined using the Fiji ImageJ software. Intracellular ROS and cell viability were assessed in PC12 cells using the OxiSelectTM Intracellular ROS Assay Kit and MTT assay, respectively. We found that microglial activation was decreased in rats pre-treated with OA prior to 6-OHDA insult as well as in rats treated with OA 1 day post 6-OHDA exposure when compared to untreated rats, as determined by shape descriptors. This finding was in correlation with significantly improved motor symptoms and increased striatal dopamine in treated rats as compared to non-treated rats. Flow cytometry assessment of PC12 cells revealed a decreased amount of intracellular ROS in cells pre-treated with OA 6 h prior to 6-OHDA exposure and cells treated with OA 1 h post 6-OHDA exposure, suggesting that OA provides neuroprotection in PC12 cells by removing intracellular ROS, thereby reducing oxidative stress. Our finding suggest that OA exhibits its neuroprotective effect by attenuating striatal microglial activation, which results in neuroinflammation that is implicated in Parkinson's disease pathology. Further studies detailing the mechanism by which OA interacts with microglia may be useful in understanding the role of OA in attenuating neuroinflammation.
ABSTRACT
UNLABELLED: Newly emerging human viruses such as Ebola virus, severe acute respiratory syndrome (SARS) virus, and HIV likely originate within an extant population of viruses in nonhuman hosts and acquire the ability to infect and cause disease in humans. Although several mechanisms preventing viral infection of particular hosts have been described, the mechanisms and constraints on viral host expansion are ill defined. We describe here mycobacteriophage Patience, a newly isolated phage recovered using Mycobacterium smegmatis mc(2)155 as a host. Patience has genomic features distinct from its M. smegmatis host, including a much lower GC content (50.3% versus 67.4%) and an abundance of codons that are rarely used in M. smegmatis. Nonetheless, it propagates well in M. smegmatis, and we demonstrate the use of mass spectrometry to show expression of over 75% of the predicted proteins, to identify new genes, to refine the genome annotation, and to estimate protein abundance. We propose that Patience evolved primarily among lower-GC hosts and that the disparities between its genomic profile and that of M. smegmatis presented only a minimal barrier to host expansion. Rapid adaptions to its new host include recent acquisition of higher-GC genes, expression of out-of-frame proteins within predicted genes, and codon selection among highly expressed genes toward the translational apparatus of its new host. IMPORTANCE: The mycobacteriophage Patience genome has a notably lower GC content (50.3%) than its Mycobacterium smegmatis host (67.4%) and has markedly different codon usage biases. The viral genome has an abundance of codons that are rare in the host and are decoded by wobble tRNA pairing, although the phage grows well and expression of most of the genes is detected by mass spectrometry. Patience thus has the genomic profile of a virus that evolved primarily in one type of host genetic landscape (moderate-GC bacteria) but has found its way into a distinctly different high-GC environment. Although Patience genes are ill matched to the host expression apparatus, this is of little functional consequence and has not evidently imposed a barrier to migration across the microbial landscape. Interestingly, comparison of expression levels and codon usage profiles reveals evidence of codon selection as the genome evolves and adapts to its new environment.
