Impacts of Environmental Changes on the Use of Cellular Storage Products in a Temperate Cnidarian-Dinoflagellate Symbiosis

<p>Members of the phylum Cnidaria, such as corals and sea anemones, often form mutualistic endosymbiotic relationships with photosynthetic dinoflagellates that are founded upon a reciprocal exchange of nutrients. In this exchange, the cnidarian host provides its symbionts with nutrients derived through respiration, heterotrophy, and the environment, while the symbionts provide their host with products of photosynthesis. The energy derived from this exchange is utilized for metabolism, growth, and reproduction; alternatively, it can be accumulated into storage bodies for use during nutritional shortages or stress. Cnidarian-algal symbioses can be found throughout the world and vary in their sensitivity to stress, with environmental changes playing a prominent role in inducing stress. Tropical cnidarian-dinoflagellate symbioses are particularly vulnerable to temperature change, with increases of just 1-2℃ above their upper thermal limit often resulting in bleaching (the breakdown of symbiosis via symbiont expulsion). In contrast, temperate cnidarian-dinoflagellate symbioses exhibit far greater tolerance to such environmental stressors, and are rarely seen to bleach in the field. It is unclear how temperate cnidarian-dinoflagellate symbioses achieve this resilience and stability. This thesis examines the effects of changes in temperature and irradiance on the content of energy-rich cellular storage products in the temperate sea anemone Anthopleura aureoradiata and its dinoflagellate endosymbionts (family: Symbiodiniaceae), in order to assess the potential of these compounds in contributing to the overall stability of the symbiosis. In particular, symbiont density and chlorophyll content (as well as photosynthetic efficiency, for experimental study only), in addition to both symbiont and host protein content, served as indicators of physiological health, and were then related to the accumulation of cellular storage products such as lipids and carbohydrates. A field study was conducted in which a population of A. aureoradiata was sampled from Wellington Harbor, New Zealand, at monthly intervals for one year. Despite monthly and seasonal variability in the physiological parameters measured, the symbiosis remained functional and stable (i.e. no signs of bleaching) throughout the year. The greatest inter-seasonal variation occurred in the symbiont cell-specific carbohydrate content, which decreased significantly between spring and summer. In contrast, host lipid content exhibited less variation than all other symbiont and host storage products. These observations suggest that symbiont carbohydrate stores are primarily utilized to sustain the symbiosis during times of seasonal environmental change (in this case, correlating with increased light and temperature during summer), while lipids may be kept in reserve. The robustness of this field population is expected; being a native species, A. aureoradiata is likely highly acclimated to the conditions that were observed throughout the year of this field study. A separate population of A. aureoradiata was subsequently acclimated to a moderate regime of temperature and irradiance, and then exposed to one of six treatments of different combined temperatures and irradiances (based on seasonal conditions in the Wellington Harbour), to establish their interactive effects on cellular storage product content. Specifically, three thermal regimes (low: 9±1°C, moderate: 14.5±1°C, high: 21±1°C), each at a low (70±10 µmol photons m-2 s-1) or high (145±15 µmol photons m-2 s-1) irradiance, were maintained for a total of sixteen weeks. Unlike in the field, a breakdown in symbiosis was observed; photo-physiological dysfunction of the symbiosis was observed within four weeks in all anemones exposed to low temperature at both irradiances, and bleaching was apparent by week eight. This response likely arose from a combination of the rapid decrease in temperature experienced upon distribution into the low-temperature tank, as well as the prolonged nature of the conditions in the experiment, which would not be experienced in the field. In contrast, the anemones maintained at both irradiances in the moderate and high temperature treatments maintained a stable symbiosis, suggesting that these conditions were not extreme enough to cause notable stress. In fact, anemones kept under both low and high irradiance within the moderate temperature treatment increased in symbiont density and exhibited the highest host lipid content relative to the other treatments, suggesting that this treatment was near-optimal for the symbiosis. Perhaps interestingly, both the moderate and high temperature treatments induced significant reductions in symbiont-specific protein, lipid, and carbohydrate content, while host storage products decreased less drastically. This observation suggests increased utilization of symbiont storage products to maintain a healthy symbiosis under these experimental conditions. My findings are consistent with previous reports of seasonal stability in temperate cnidarian-dinoflagellate symbioses; moreover, I provide experimental evidence for the utilization of symbiont storage products as a means of maintaining symbiosis stability, though this was less apparent in the field. Although recent studies have made great progress in identifying patterns of stability in temperate cnidarian-dinoflagellate symbioses, additional studies are required to build a more comprehensive picture of the mechanisms involved. Future studies would benefit from increased frequency of field sampling, including assessments of nutrient availability and host reproductive cycles, to better understand the monthly and seasonal variability in the intracellular storage product use observed in the field. Nevertheless, results of this study contribute to an improved understanding of the physiology and remarkable stability of temperate cnidarian-dinoflagellate symbioses, with implications for predictions of how they might respond to future climate change scenarios.</p>

Impacts of Environmental Changes on the Use of Cellular Storage Products in a Temperate Cnidarian-Dinoflagellate Symbiosis

<p>Members of the phylum Cnidaria, such as corals and sea anemones, often form mutualistic endosymbiotic relationships with photosynthetic dinoflagellates that are founded upon a reciprocal exchange of nutrients. In this exchange, the cnidarian host provides its symbionts with nutrients derived through respiration, heterotrophy, and the environment, while the symbionts provide their host with products of photosynthesis. The energy derived from this exchange is utilized for metabolism, growth, and reproduction; alternatively, it can be accumulated into storage bodies for use during nutritional shortages or stress. Cnidarian-algal symbioses can be found throughout the world and vary in their sensitivity to stress, with environmental changes playing a prominent role in inducing stress. Tropical cnidarian-dinoflagellate symbioses are particularly vulnerable to temperature change, with increases of just 1-2℃ above their upper thermal limit often resulting in bleaching (the breakdown of symbiosis via symbiont expulsion). In contrast, temperate cnidarian-dinoflagellate symbioses exhibit far greater tolerance to such environmental stressors, and are rarely seen to bleach in the field. It is unclear how temperate cnidarian-dinoflagellate symbioses achieve this resilience and stability. This thesis examines the effects of changes in temperature and irradiance on the content of energy-rich cellular storage products in the temperate sea anemone Anthopleura aureoradiata and its dinoflagellate endosymbionts (family: Symbiodiniaceae), in order to assess the potential of these compounds in contributing to the overall stability of the symbiosis. In particular, symbiont density and chlorophyll content (as well as photosynthetic efficiency, for experimental study only), in addition to both symbiont and host protein content, served as indicators of physiological health, and were then related to the accumulation of cellular storage products such as lipids and carbohydrates. A field study was conducted in which a population of A. aureoradiata was sampled from Wellington Harbor, New Zealand, at monthly intervals for one year. Despite monthly and seasonal variability in the physiological parameters measured, the symbiosis remained functional and stable (i.e. no signs of bleaching) throughout the year. The greatest inter-seasonal variation occurred in the symbiont cell-specific carbohydrate content, which decreased significantly between spring and summer. In contrast, host lipid content exhibited less variation than all other symbiont and host storage products. These observations suggest that symbiont carbohydrate stores are primarily utilized to sustain the symbiosis during times of seasonal environmental change (in this case, correlating with increased light and temperature during summer), while lipids may be kept in reserve. The robustness of this field population is expected; being a native species, A. aureoradiata is likely highly acclimated to the conditions that were observed throughout the year of this field study. A separate population of A. aureoradiata was subsequently acclimated to a moderate regime of temperature and irradiance, and then exposed to one of six treatments of different combined temperatures and irradiances (based on seasonal conditions in the Wellington Harbour), to establish their interactive effects on cellular storage product content. Specifically, three thermal regimes (low: 9±1°C, moderate: 14.5±1°C, high: 21±1°C), each at a low (70±10 µmol photons m-2 s-1) or high (145±15 µmol photons m-2 s-1) irradiance, were maintained for a total of sixteen weeks. Unlike in the field, a breakdown in symbiosis was observed; photo-physiological dysfunction of the symbiosis was observed within four weeks in all anemones exposed to low temperature at both irradiances, and bleaching was apparent by week eight. This response likely arose from a combination of the rapid decrease in temperature experienced upon distribution into the low-temperature tank, as well as the prolonged nature of the conditions in the experiment, which would not be experienced in the field. In contrast, the anemones maintained at both irradiances in the moderate and high temperature treatments maintained a stable symbiosis, suggesting that these conditions were not extreme enough to cause notable stress. In fact, anemones kept under both low and high irradiance within the moderate temperature treatment increased in symbiont density and exhibited the highest host lipid content relative to the other treatments, suggesting that this treatment was near-optimal for the symbiosis. Perhaps interestingly, both the moderate and high temperature treatments induced significant reductions in symbiont-specific protein, lipid, and carbohydrate content, while host storage products decreased less drastically. This observation suggests increased utilization of symbiont storage products to maintain a healthy symbiosis under these experimental conditions. My findings are consistent with previous reports of seasonal stability in temperate cnidarian-dinoflagellate symbioses; moreover, I provide experimental evidence for the utilization of symbiont storage products as a means of maintaining symbiosis stability, though this was less apparent in the field. Although recent studies have made great progress in identifying patterns of stability in temperate cnidarian-dinoflagellate symbioses, additional studies are required to build a more comprehensive picture of the mechanisms involved. Future studies would benefit from increased frequency of field sampling, including assessments of nutrient availability and host reproductive cycles, to better understand the monthly and seasonal variability in the intracellular storage product use observed in the field. Nevertheless, results of this study contribute to an improved understanding of the physiology and remarkable stability of temperate cnidarian-dinoflagellate symbioses, with implications for predictions of how they might respond to future climate change scenarios.</p>

Safety and Efficacy of 14-Day Cold Stored Platelets in Reversing Effects of Aspirin

Background: Aspirin is an antiplatelet therapy used to reduce the risk of vascular occlusive events. However, this therapy is associated with an increased risk of bleeding for which there is no antidote currently. Transfusion of 5-day room stored platelets (RSP) at 22°C can reverse the effect of aspirin but surprisingly, the recent randomized PATCH trial showed increased morbidity and mortality for patients who received RSP transfusion for intracranial hemorrhage while on aspirin. Prior studies have shown that cold stored platelets (CSP) at 4°C are mildly activated and may participate in clot formation immediately, thus may have the potential to reduce blood loss more rapidly than RSP in acutely bleeding patients. CSP also have the added advantages of decreased risk of bacterial contamination and longer shelf-life up to 14 days per current FDA variance. However, the function of 14-day CSP in plasma after transfusion is unclear and lacks high quality data. We aimed to evaluate the post-transfusion safety and efficacy of 14-day CSP in reversing the effects of aspirin therapy compared to that of 7-day RSP. Methods: Seven healthy human subjects were included in the analysis of this randomized, controlled, crossover study comparing transfusion of autologous 14-day CSP to 7-day RSP. Each subject participated in two study periods, which crossed over from one storage product to the other (CSP vs. RSP) according to randomization. For each study period, subjects underwent an apheresis platelet collection for autologous transfusion. Platelets were stored for either 14 days for CSP or 7 days for RSP. Subjects received a loading dose of aspirin 24 hours prior to transfusion. Blood was drawn at baseline, immediately pre-transfusion, at 1-hr, 4-hr, and 24-hr post-transfusion for an array of platelet function testing. After a washout period of 10-28 days, second study period commenced with similar sequence of events as the first study period using the other platelet storage product. The primary endpoint is the VerifyNOW Aspirin Reaction Units (ARU) at 1-hr after autologous transfusion. Secondary endpoints include ARU at 4-hr and 24-hr post transfusion, light transmission aggregometry in response to arachidonic acid and collagen, and the corrected count increment. Paired t-tests were used for statistical analysis between the two groups and, where appropriate, the change from pre-transfusion values were analyzed. Results: Transfusion of 14-day CSP and 7-day RSP units were well-tolerated by all subjects. Storage of platelets in the cold led to a non-significant trend for decreased platelet count, and the total platelet yield at the end of storage was significantly less in 14-day CSP compared to 7-day RSP (p=0.02). However, the corrected count increment did not differ significantly at 1-hr after transfusion. Platelet aggregation using the agonists, arachidonic acid 0.5mM and collagen 2.5ug/mL, did not reveal any significant difference between the two groups at any time points. The primary endpoint, platelet function testing by VerifyNOW, showed a larger change in platelet responsiveness at 1-hr post-transfusion in RSP than in CSP (p=0.03). Surprisingly, only RSP transfusion resulted in a significant change from the pre-transfusion VerifyNow measurements. Later time points showed a slight trend for improved platelet function as measured by VerifyNow with transfusion of both platelet products, but none were statistically significant. Conclusion: We report the first safety and efficacy data for 14-day cold stored platelets in in healthy humans. While prior in-vitro studies have demonstrated possible hemostatic superiority of cold stored over room temperature stored platelets, we observed inferior reversal of aspirin at early time points with CSP. This was in contrast to the results from our previous study, where 5 day-stored CSP were equivalent to RSP at early post transfusion time points. Further studies are needed to evaluate the maximal storage that provides functional equivalency between CSP and RSP. In addition, studies in actively bleeding patients are needed. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Preparation of Expanded Graphite and Fly Ash Base High Temperature Compound Phase Transition Heat Absorption Material and Its Application in Building Temperature Regulation

In this study, the product substance was pretreated with coal excrement as a raw product substance. Taking expanded graphite and paraffin as phase change product substances, expanded graphite and paraffin or coal excrement base high temperature compound phase transition thermal storage product substance were prepared by mixed sintering method. Firstly, the sintering mechanism of the composite should be deduced. In addition, XRD, TG-DSC and other analytical techniques were used to characterize the morphology and physical character of the product substances. In order to test the temperature regulation ability of the product substance, the composite product substance was fused with cement. The cement block added with composite product substance was taken as the research object, verify the performance of compound phase transition product substance on temperature regulation. In the experimental procedure, when phase change product substances' the proportion was less than 60%, the high temperature compound phase transition thermal storage product substances prepared were well-formed and had good crystallography, and the green body' various tissues were densely distributed. When the paraffin content was above 35 wt%, the external density, compressive strength, and cement blocks' thermal conductivity added with composite product substances increased significantly compared with ordinary expanded perlite cement blocks, which was suitable for building product substances' requirements. When the paraffin content reached 38 wt%, a temperature difference of 19.2 °C to 26.1 °C appeared in the composite product substance' corresponding cube space, indicating that room temperature can be adjusted by adding a certain phase change product substance.