Interleukin 2 and stimulator lymphoblastoid cells will induce human thymocytes to bind and kill K562 targets.

Human thymocytes cultured in the presence of IL-2 and an irradiated B cell line became cytotoxic to K562 target cells. Thymocytes cultured alone or with only IL-2 exhibited almost no killing, but thymocytes cultured in the presence of stimulator cells alone exhibited low levels of cytotoxic activity. Removal of Fc gamma receptor-bearing cells from the activated thymocyte population almost completely abolished the binding and lytic activity. Separation of thymocytes into Fc microns+ and Fc microns-cells before culturing with IL-2 and stimulator cells revealed that only the Fc microns+ subpopulation developed into K562 killer cells. These findings indicate that modulation of Fc microns to Fc gamma receptors on the thymocyte cell surface is part of the maturation process of this particular subset of cytotoxic cells. Morphologically, most of the activated Fc gamma+ K562-binding cells were large, granulated lymphocytes. Only very few of the round, nongranulated small thymocytes were bound to K562 target cells

Cold Body Temperature as an Evolutionary Shaping Force in the Physiology of Antarctic Fishes

Notothenioid fishes that dominate the fish fauna surrounding Antarctica have been evolving for 10-14 million years at a nearly constant body temperature of ~0C throughout their life histories. As a result, this group of animals is uniquely suited to studies aimed at understanding and identifying features of physiology and biochemistry that result from the process of evolution at cold body temperature. This project has three major objectives aimed at examining adaptations for life in cold environments: 1. Identify the amino acid substitutions in the fatty acid-binding pocket of fatty acyl CoA synthetase (FACS) that explain its substrate specificity. Fatty acids are a major fuel of energy metabolism in Antarctic fishes. FACS catalyzes the condensation of CoASH and fatty acids to fatty acyl CoA esters, a step required for subsequent metabolism of these important compounds. This research may permit us to resolve the specific amino acid substitutions that explain both substrate specificity and preservation of catalytic rate of notothenioid FACS at cold physiological temperatures.2. Produce a rigorous biochemical and biophysical characterization of the intracellular calcium-binding protein, parvalbumin, from white axial musculature of Antarctic fishes. Parvalbumin plays a pivotal role in facilitating the relaxation phase of fast-contracting muscles and is a likely site of strong selective pressure. Preliminary data strongly indicate that the protein from Antarctic fishes has been modified to ensure function at cold temperature. A suite of physical techniques will be used to determine dissociation constants of Antarctic fish parvalbumins for calcium and magnesium and unidirectional rate constants of ion-dissociation from the protein. Full-length cDNA clones for Antarctic fish parvalbumin(s) will permit deduction of primary amino acid sequence These data will yield insight into structural elements that permit the protein from notothenioid fishes to function at very cold body temperature.3. Conduct a broad survey of the pattern of cardiac myoglobin expression in the Suborder Notothenoidei. Previous work has indicated a variable pattern of presence or absence of the intracellular oxygen-binding protein, myoglobin (Mb), in hearts of one family of Antarctic notothenioid fishes (Channichthyidae; icefishes). Because Mb is of physiological value in species that express the protein, the observed pattern of interspecific expression has been attributed to unusually low niche competition in the Southern Ocean. This leads to the prediction that similar loss of cardiac Mb should be observed in other notothenioid taxa. This part of the project will survey for the presence and absence of cardiac Mb in as many notothenioid species as possible and, if Mb-lacking species are detected, will extend analyses to determine the mechanism(s) responsible for loss of its expression using molecular biological techniques

Collaborative Research: Differential Expression of Oxygen-binding Proteins in Antarctic Fishes Affects Nitric Oxide-mediated Pathways of Angiogenesis and Mitochondrial Biogenesis

The polar ocean presently surrounding Antarctica is the coldest, most thermally stable marine environment on earth. Because oxygen solubility in seawater is inversely proportional to temperature, the cold Antarctic seas are an exceptionally oxygen-rich aquatic habitat. Eight families of a single perciform suborder, the Notothenioidei, dominate the present fish fauna surrounding Antarctica. Notothenioids account for approximately 35% of fish species and 90% of fish biomass south of the Antarctic Polar Front. Radiation of closely related notothenioid species thus has occurred rapidly and under a very unusual set of conditions: relative oceanographic isolation from other faunas due to circumpolar currents and deep ocean trenches surrounding the continent, chronically, severely cold water temperatures, very high oxygen availability, very low levels of niche competition in a Southern Ocean depauperate of species subsequent to a dramatic crash in species diversity of fishes that occurred sometime between the mid-Tertiary and present. These features make Antarctic notothenioid fishes an uniquely attractive group for the study of physiological and biochemical adaptations to cold body temperature. Few distinctive features of Antarctic fishes are as unique as the pattern of expression of oxygen-binding proteins in one notothenioid family, the Channichthyidae (Antarctic icefishes). All channichthyid icefishes lack the circulating oxygen-binding protein, hemoglobin (Hb); the intracellular oxygen-binding protein, myoglobin (Mb) is not uniformly expressed in species of this family. Both proteins are normally considered essential for adequate delivery of oxygen to aerobically poised tissues of animals. To compensate for the absence of Hb, icefishes have developed large hearts, rapidly circulate a large blood volume and possess elaborate vasculature of larger lumenal diameter than is seen in red-blooded fishes. Loss of Mb expression in oxidative muscles correlates with dramatic elevation in density of mitochondria within the cell, although each individual organelle is less densely packed with respiratory proteins. Within the framework of oxygen movement, the adaptive significance of greater vascular density and mitochondrial populations is understandable but mechanisms underlying development of these characteristics remain unknown. The answer may lie in another major function of both Hb and Mb, degradation of the ubiquitous bioactive compound, nitric oxide (NO). The research will test the hypothesis that loss of hemoprotein expression in icefishes has resulted in an increase in levels of NO that mediate modification of vascular systems and expansion of mitochondrial populations in oxidative tissues. The objectives of the proposal are to quantify the vascular density of retinas in +Hb and -Hb notothenioid species, to characterize NOS isoforms and catalytic activity in retina and cardiac muscle of Antarctic notothenioid fishes, to evaluate level of expression of downstream factors implicated in angiogenesis (in retinal tissue) and mitochondrial biogenesis (in cardiac muscle), and to determine whether inhibition of NOS in vivo results in regression of angiogenic and mitochondrial biogenic responses in icefishes. Broader impacts range from basic biology, through training of young scientists, to enhanced understanding of clinically relevant biomedical processes

Holocene sea level change and archaeology in the inner Thames estuary, London, UK

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Collaborative Research: Linkages among Mitochondrial Form, Function and Thermal Tolerance of Antarctic Notothenioid Fishes

Antarctic notothenioid fishes have evolved in the Southern Ocean for 10-14 MY under an unusual set of circumstances. Their characteristics include the complete absence of the circulating oxygen-binding protein, hemoglobin (Hb) within the Channichthyid (Icefish) family of notothenioids. Moreover, some species within the 16 members of this family have also lost the ability to express the oxygen-binding and storage protein, myoglobin (Mb) in cardiac muscle. Our previous work has determined that the loss of Hb and/or Mb is correlated with significant increases in densities of mitochondria within oxidative tissues, and extensive remodeling of these vital organelles. To date, nothing is known about how modifications in mitochondrial architecture of icefishes affect organelle function, or more importantly, how they affect organismal-level physiology. Most critical for Antarctic fishes is that mitochondrial characteristics have been linked to how well ectotherms can withstand increases in temperature. This collaborative research project will address the hypothesis that the unusual mitochondrial architecture of Antarctic Channichthyids has led to changes in function that impact their ability to withstand elevations in temperature. Specifically, the research will (1) determine if the unusual mitochondrial architecture of icefishes affects function and contributes to organismal thermal sensitivity, (2) identify differences in organismal thermal tolerance between red- and white- blooded notothenioids, (3) identify molecular mechanisms regulating changes in mitochondrial structure in icefishes. The results may establish channichthyid icefishes as a sentinel taxon for signaling the impact of global warming on the Southern Ocean. Broad impacts of this project will be realized by participation of high school biology teachers in field work through cooperation with the ARMADA project at the University of Rhode Island, as well as graduate education

Evolution of an Oxygen-Binding Hemoprotein in a Unique Environment: Myoglobin in the Hemoglobinless Antarctic Icefishes

For approximately the last 40 million years, a unique fish fauna has been evolving in the frigid seas surrounding Antarctica. Physiological function of these water-breathing animals has demanded many adaptations to ensure proper metabolism and regulation of biochemical processes at cell temperatures of about OoC. Among the polar fishes, one family is particularly unusual, the Channichthyid icefishes. Species in this family lack hemoglobin in their circulating blood and, at least the majority, do not possess the intracellular respiratory protein, myoglobin, that is normally responsible for enhancing movement of oxygen through aerobic muscle tissues. The PI\u27s have recently found two species of icefish, Pseudochaenichthys georgianus and Chionodraco rastrospinosus, however, that express myoglobin, but only in one tissue - heart ventricle. The metabolism of all icefish species is highly aerobic and is based largely on the combustion of fatty fuels as energy sources. The field work will consist of a combination of trawling from R/V Polar Duke to capture icefish species and laboratory work at Palmer Station using tissues from these animals. At Palmer Station they will conduct experiments and prepare purified, fixed and or frozen material for shipment back to our CONUS laboratory for further analyses. Our overall goals are: 1) to test whether to determine myoglobin protein remains physiologically significant at the cold body temperatures of Antarctic fishes, 2) to reconstruct the evolution of the myoglobin gene within the icefish family and, 3) to identify the mechanisms controlling expression of myoglobin within icefish and related species

Ski area effects on headwater streamflow

2022 Summer.Includes bibliographical references.Colorado headwater streams produce water supply for the West. The effects of singular land use changes on headwater watersheds have been studied at length, but much less is known about the combined interactions of multiple land use changes on headwater streamflow generation. We examined how the interactions of three land use changes associated with ski area developments (tree clearing, trail and road building, and artificial snow application) affected streamflow at a ski area in northern Colorado. Our study area included three watersheds with stratified levels of development, within a United States Forest Service ski area permit boundary. Three main creeks and their tributaries were equipped with twelve pressure transducers scheduled for data collection at continuous 15 minute intervals over two water years beginning in late summer 2019. Burgess Creek (5.91 km2), which had the greatest degree of development and creek accessibility, was equipped with 9 data loggers; Priest Creek (2.35 km2) had two monitoring sites, and Beaver Creek (2.28 km2) had one. We initially performed an ANOVA comparison of our ski area stream data to two reference watersheds, Hot Spring Creek (14.87 km2) and Spring Creek (2.65 km2) and detected no significant differences in streamflow generation or timing. We then examined how streamflow generation and timing related to the degree of development and watershed characteristics using both univariate correlation analysis and multivariate models. Mean basin elevation was the most significant driver of the timing of flow delivery; development also plays an obvious role in both streamflow generation and timing. Total seasonal and annual streamflow generation increase significantly with development, and the timing of streamflow is earlier in the season in developed watersheds. Overall, this study shows that development affects how and when streamflow is generated from forested headwater stream systems, but our conclusions apply to just one ski area in northern Colorado. Long-term stream monitoring across watersheds with multiple disturbances, like those seen on ski resorts, should be a priority to understand how water delivery is affected by development

Retinoic acid induces the differentiation of B cell hybridomas from patients with common variable immunodeficiency.

Human-human B cell hybridomas constructed from B lymphocytes of common variable immunodeficiency (CVI) patients and the nonsecreting cell line WIL2/729 HF consistently secrete low levels of Ig and appear to retain a defect characteristic of the CVI patient's B cells. We assessed the differentiative capacity of retinoic acid (RA) on these hybridomas, as well as on hybridomas constructed from normal B cells and from patients with selective IgA deficiency. RA at concentrations varying between 10(-5) and 10(-9) M augmented IgM secretion 4-20-fold from four of four CVI hybridomas tested, but did not affect Ig secretion from normal or IgA-deficiency hybridomas. In support of this elevated Ig secretion, RA enhanced the de novo synthesis of biosynthetically labeled light (kappa) and heavy (mu) Ig (up to 4- and 15-fold, respectively) in the CVI hybridoma line JK32.1. The increase in IgM synthesis/secretion could not be accounted for by RA-induced alteration in the cell cycle. In inducing this increase in IgM production, RA was found to affect two aspects of Ig gene expression: (a) the steady-state levels of heavy and light chain mRNAs were enhanced, and (b) the processing of mu heavy chain transcripts to the secreted mRNA form became favored over the membrane mRNA form. We also show that expression of Leu-17 (CD38), a surface marker that is re-expressed in the late pre-plasma stage of B cell development, was increased by RA from less than 20% to greater than 90% of the total cell population, with a concomitant 4-10-fold augmentation in the mean fluorescence intensity. Changes in both Leu-17 expression and de novo Ig synthesis were prominent by 24 h, but could be observed as early as 8 h after induction. Taken together, our study demonstrates that RA affects a marked alteration in the differentiated state of the CVI hybridoma clones. This finding suggests that retinoids can enhance the functional capabilities of B cells with defects in maturation and support further studies to evaluate their clinical potential in CVI