Browsing Bachelor's Essays (Embargoed) by Issue Date "2022-03-29T19:01:21Z"
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- ItemMKP‐1 LPS-Driven Osteoclastogenesis More Prevalent in Dusp‐1‐/‐ Females Than Males(2013-06-05) Browne, Courtney G; Valerio, Michael S; Kirkwood, Keith L; Morrison, Susan$Periodontal disease is the result of chronic inflammatory bone loss due to a sustained immune system response to pathogenic invasion. Osteoclasts (OC) are multinucleated, bone resorbing cells that play a vital role in bone turnover. Receptor activator of nuclear factor kappa B ligand (RANKL) is the physiological regulator of osteoclastogenesis (OCgen) in naïve progenitor populations. Gram-negative bacteria involved in pathogenesis of periodontal disease contain the endotoxin, lipopolysaccharide (LPS), which increases RANKL expression by activating innate immune inflammatory cascades including mitogen activated protein kinase (MAPK). MAPK activity is negatively regulated by MAPK phosphatase (MKP)-1 which is encoded by the gene Dusp-1. The current study aims to investigate the role of Dusp-1 when driven by LPS and primed with RANKL OC progenitors (OCP). Based on the role of MKP-1 as a negative regulator of activated MAPK, we hypothesize that Dusp-1 deficient OCP will form more OCs in response to LPS stimuli. OCPs were isolated from male and female hematopoietic stem cells via magnet-activated cell sorting (MACS) and plated (5x104/well) based on expression of CD11b (CD11bhigh, low and neg), from Dusp-1+/+ and Dusp-1-/- mice. Cells were primed with macrophage colony stimulating factor (M-CSF) and RANKL for 48 hours and were then stimulated with 10ng/ml of A. actinomycetemcomitans LPS for up to 4 days. A tartrate resistant acid phosphatase (TRAP) assay was used to visualize formation of OCs at day 0 (M-CSF +RANKL only) and days 2 and 4 post-LPS. Images generated from TRAP were used to determine the number of OCs, number of nuclei per OC, and area per OC. Day 0 cells lacked any exposure to LPS, resulting in minimal variation between Dusp-1+/+ and Dusp-1-/- for all CD11b populations. By Day 2 there was a significant trend of increased female Dusp-1-/- OCgen, with CD11bhigh having the greatest difference (p<0.05). Male data followed an opposite trend, with Dusp-1-/- having less OCgen for all CD11b sorted populations. By Day 4, female CD11bhigh (p<0.0001) and CD11blow (p<0.05) had significantly more OCs for Dusp-1-/-. Day 4 of the males showed a large decrease in Dusp-1+/+ cells, specifically in CD11blow. The data show that LPS induced more osteoclast formation in female Dusp-1-/- mice than Dusp-1+/+ mice. The male Dusp-1+/+ populations show a preliminary increase in OCgen. However, by day 4 there are substantially more Dusp-1-/- cells that are larger than their Dusp1+/+ counterparts. Therefore, these studies indicate that MKP-1 is key negative regulator of MAPK and the subsequent increased OC differentiation promotes inflammatory bone resorption.
- ItemThe effects of pesticide and salinity on early life stages of the green tree frog (<i>Hyla cinerea</i>)(2013-11-05) Wilder, Anneke E; Welch, AllisonIncreased salinity in freshwater habitats can result from anthropogenic factors such as climate change and sea level rise, use of road salts, over-irrigation and groundwater depletion. This emerging threat can affect freshwater species in both coastal and inland environments. Pesticides have also been found in freshwater environments around the globe, and can be harmful to many different aquatic organisms. Amphibians are highly sensitive to changes in their surroundings, making them good indicators of environmental quality, especially in freshwater habitats. This study examined the effects of increased salinity and a common insecticide, carbaryl, on early life history stages of the green tree frog (<i>Hyla cinerea</i>). To test effects of salinity on sperm activity, we subjected <i>H. cinerea</i> sperm to levels of salinity ranging from freshwater to moderately brackish and analyzed activity using computer assisted sperm analysis (CASA). Effects of carbaryl on sperm activity were measured in a similar fashion, using concentrations ranging from zero to a dose that is beyond the expected field concentration. We also examined female oviposition site selection in response to salinity and the presence of carbaryl in artificial pools. Mean sperm motility and velocity were both found to decrease as salinity concentration increased, butcarbaryl had no significant effect on sperm activity. Females tended to avoid ovipositing in pools with increased salinity. Although no significant difference was observed in oviposition between pools with and without carbaryl, pools received no oviposition when freshly dosed with carbaryl. These findings suggest that increased salinity may negatively affect reproductive success in <i>H. cinerea</i>, but that females may be able to avoid these effects through depositing eggs in freshwater sites. Selective oviposition may increase the ability of amphibian populations to persist in degraded habitats.