Variation of hair cortisol in two herds of migratory caribou (Rangifer tarandus): implications for health monitoring.

Migratory caribou (Rangifer tarandus sspp.) is an ecotype of conservation concern that is experiencing increased cumulative stressors associated with rapid climate change and development in Arctic Canada. Increasingly, hair cortisol concentrations (HCCs) are being used to monitor seasonal hypothalamic-pituitary-adrenal axis activity of ungulate populations; yet, the effect of key covariates for caribou (sex, season, sampling source, body location) are largely unknown. The objectives of this research were 4-fold: first, we assessed the impact of body location (neck, rump) sampling sites on HCC; second, we assessed key covariates (sex, sampling method, season) impacting HCCs of caribou; third, we investigated inter-population (Dolphin and Union (DU), Bluenose-East (BNE)) and inter-annual differences in HCC and fourth, we examined the association between HCCs and indices of biting insect activity on the summer range (oestrid index, mosquito index). We examined hair from 407 DU and BNE caribou sampled by harvesters or during capture-collaring operations from 2012 to 2020. Linear mixed-effect models were used to assess the effect of body location on HCC and generalized least squares regression (GLS) models were used to examine the impacts of key covariates, year and herd and indices of biting insect harassment. HCC varied significantly by body location, year, herd and source of samples (harvester vs capture). HCC was higher in samples taken from the neck and in the DU herd compared with the BNE, decreased linearly over time and was higher in captured versus hunted animals (P < 0.05). There was no difference in HCC between sexes, and indices of biting insect harassment in the previous year were not significantly associated with HCC. This study identifies essential covariates impacting the HCC of caribou that must be accounted for in sampling, monitoring and data interpretation.

Qiviut cortisol reflects hypothalamic-pituitary-adrenal axis activity in muskoxen (Ovibos moschatus).

Muskoxen (Ovibos moschatus) are increasingly exposed to a broad diversity of stressors in their rapidly changing Arctic environment. There is an urgent need to develop validated tools to monitor the impact of these stressors on the hypothalamic-pituitary-adrenal (HPA) axis activity of muskoxen to help inform conservation actions. Here, we evaluated whether muskox qiviut (dense wooly undercoat) cortisol accurately reflects changes in HPA axis activity. Two repeated pharmacological challenges, involving weekly administrations of saline (control group) or adrenocorticotropic hormone (ACTH) during five consecutive weeks, were done on captive muskoxen, in winter (no hair growth) and summer (maximum hair growth). Pre-challenge qiviut cortisol levels were significantly higher in the shoulder than in the neck, but neither differed from rump concentrations. Qiviut cortisol levels significantly increased (p < 0.001) in response to the administration of ACTH during the hair growth phase, but not in the absence of growth (p = 0.84). Cortisol levels in the qiviut segment grown during the summer challenge increased significantly over a six-month period in the ACTH-injected muskoxen with a similar trend occurring in the control animals. Finally, cortisol levels in shed qiviut were significantly higher and not correlated to those of fully grown qiviut shaved three months earlier. Our results show that cortisol is deposited in qiviut during its growth and that qiviut cortisol can thus be used as an integrated measure of HPA axis activity over the period of the hair's growth. Differences in qiviut cortisol across body regions, significant differences in qiviut segments over time, and differences between shed qiviut versus unshed qiviut, highlight the importance of consistent design and methodology for sample collection and analyses in order to account for sources of variation when using qiviut cortisol as a biomarker of HPA axis activity in muskoxen.

Adaptations, life-history traits and ecological mechanisms of parasites to survive extremes and environmental unpredictability in the face of climate change.

Climate change is increasing weather unpredictability, causing more intense, frequent and longer extreme events including droughts, precipitation, and both heat and cold waves. The performance of parasites, and host-parasite interactions, under these unpredictable conditions, are directly influenced by the ability of parasites to cope with extremes and their capacity to adapt to the new conditions. Here, we review some of the structural, behavioural, life history and ecological characteristics of parasitic nematodes that allow them to persist and adapt to extreme and changing environmental conditions. We focus primarily, but not exclusively, on parasitic nematodes in the Arctic, where temperature extremes are pronounced, climate change is happening most rapidly, and changes in host-parasite interactions are already documented. We discuss how life-history traits, phenotypic plasticity, local adaptation and evolutionary history can influence the short and long term response of parasites to new conditions. A detailed understanding of the complex ecological processes involved in the survival of parasites in extreme and changing conditions is a fundamental step to anticipate the impact of climate change in parasite dynamics.

Renal trace elements in barren-ground caribou subpopulations: Temporal trends and differing effects of sex, age and season.

Caribou (Rangifer tarandus) are a culturally significant food resource for communities in northern Canada and Greenland. Many barren-ground caribou subpopulations are currently in decline, some dramatically; understanding the influence of stressors, such as toxic trace metals, is important. These contaminants enter Arctic terrestrial environments via atmospheric transport from industrialized areas and from local sources, accumulating there in the environment. Understanding how trace element concentrations interact and are influenced by caribou sex, age and season of collection is essential to evaluating trends in these elements over time and differences among subpopulations. We used path analysis to model the direct and indirect relationships between these variables in the Porcupine subpopulation and in barren-ground caribou from the Canadian Arctic and Greenland. Renal cadmium (Cd), copper (Cu) and mercury (Hg) varied significantly among subpopulations. Hg was positively correlated with Cd, Cu and selenium (Se) in female Porcupine caribou whereas Cd and Cu were negatively correlated in male Porcupine caribou. Age, season and sex influenced all three element concentrations and should be considered when comparing elements among caribou subpopulations or years. Renal Cd decreased slightly from the Canadian Western Arctic to Greenland and increased slightly over time, possibly reflecting patterns of atmospheric deposition. Renal Hg did not change significantly over time, and differences among subpopulations did not follow specific geographical patterns. Renal Cu declined over time, the changes being markedly different among subpopulations, sexes and seasons. This temporal decline is likely due to changes in diet, which could be driven by various environmental factors. Declining Cu concentrations in caribou is of concern as low levels could negatively affect reproductive success and therefore caribou at a population level. Continuing to monitor element concentrations in caribou is essential to better comprehend potential threats facing the species, and to promote food security in communities harvesting this important resource.

Multi-pathogen serological survey of migratory caribou herds: A snapshot in time.

Pathogens can impact host survival, fecundity, and population dynamics even when no obvious disease is observed. Few baseline data on pathogen prevalence and diversity of caribou are available, which hampers our ability to track changes over time and evaluate impacts on caribou health. Archived blood samples collected from ten migratory caribou herds in Canada and two in Greenland were used to test for exposure to pathogens that have the potential to effect population productivity, are zoonotic or are emerging. Relationships between seroprevalence and individual, population, and other health parameters were also examined. For adult caribou, the highest overall seroprevalence was for alphaherpesvirus (49%, n = 722), pestivirus (49%, n = 572) and Neospora caninum (27%, n = 452). Lower seroprevalence was found for parainfluenza virus type 3 (9%, n = 708), Brucella suis (2%, n = 758), and Toxoplasma gondii (2%, n = 706). No animal tested positive for antibodies against West Nile virus (n = 418) or bovine respiratory syncytial virus (n = 417). This extensive multi-pathogen survey of migratory caribou herds provides evidence that caribou are exposed to pathogens that may have impacts on herd health and revealed potential interactions between pathogens as well as geographical differences in pathogen exposure that could be linked to the bio-geographical history of caribou. Caribou are a keystone species and the socio-economic cornerstone of many indigenous cultures across the North. The results from this study highlight the urgent need for a better understanding of pathogen diversity and the impact of pathogens on caribou health.

Experimental Life-Cycle of Varestrongylus eleguneniensis (Nematoda: Protostrongylidae) in a Captive Reindeer (Rangifer tarandus tarandus) and a Muskox (Ovibos moschatus moschatus).

The life-cycle of a recently described protostrongylid lungworm, Varestrongylus eleguneniensis, which infects caribou, muskoxen, and moose from Arctic and boreal regions of North America, was completed experimentally for the first time. A native North American slug species, Deroceras laeve, was infected with the first-stage larvae (L1) isolated from the feces of wild muskoxen to generate third-stage larvae (L3). These were administered to a captive reindeer calf (250 L3) and an adult captive muskox (380 L3). The prepatent periods for the reindeer and muskox were 56 and 72 days, respectively. Patency lasted for only 19 days in the reindeer, and fecal larval counts were very low (0.09-1.53 larvae per gram of feces). Patency in the muskox was at least 210 days, and likely over 653 days, and the fecal larval counts were higher (0.06-17.8 larvae per gram of feces). This work provides the first experimental completion of the life-cycle of V. eleguneniensis.

Arctic systems in the Quaternary: ecological collision, faunal mosaics and the consequences of a wobbling climate.

Climate oscillations and episodic processes interact with evolution, ecology and biogeography to determine the structure and complex mosaic that is the biosphere. Parasites and parasite-host assemblages are key components in a general explanatory paradigm for global biodiversity. We explore faunal assembly in the context of Quaternary time frames of the past 2.6 million years, a period dominated by episodic shifts in climate. Climate drivers cross a continuum from geological to contemporary timescales and serve to determine the structure and distribution of complex biotas. Cycles within cycles are apparent, with drivers that are layered, multifactorial and complex. These cycles influence the dynamics and duration of shifts in environmental structure on varying temporal and spatial scales. An understanding of the dynamics of high-latitude systems, the history of the Beringian nexus (the intermittent land connection linking Eurasia and North America) and downstream patterns of diversity depend on teasing apart the complexity of biotic assembly and persistence. Although climate oscillations have dominated the Quaternary, contemporary dynamics are driven by tipping points and shifting balances emerging from anthropogenic forces that are disrupting ecological structure. Climate change driven by anthropogenic forcing has supplanted a history of episodic variation and is eliminating ecological barriers and constraints on development and distribution for pathogen transmission. A framework to explore interactions of episodic processes on faunal structure and assembly is the Stockholm Paradigm, which appropriately shifts the focus from cospeciation to complexity and contingency in explanations of diversity.

Parasites, stress and reindeer: infection with abomasal nematodes is not associated with elevated glucocorticoid levels in hair or faeces.

Stress hormones (glucocorticoids), incorporated into hair/fur and faeces, have been proposed as biomarkers of overall health in wildlife. Although such biomarkers may be helpful for wildlife conservation and management, their use has rarely been validated. There is a paucity of studies examining the variation of stress hormones in mammals and how they relate to other health measures, such as parasitism. Parasites are ubiquitous in wildlife and can influence the fitness of individual animals and populations. Through a longitudinal experiment using captive reindeer (Rangifer tarandus tarandus), we tested whether animals infected with Ostertagia gruehneri, a gastrointestinal nematode with negative impacts on fitness of the host, had higher stress levels compared with those that had been treated to remove infection. Faecal samples were collected weekly for 12 weeks (June-September) and hair was collected at the start and end of the study; glucocorticoids were quantified using enzyme immunoassays. Contrary to what was expected, infected reindeer had similar levels of cortisol in hair and slightly lower glucocorticoid metabolites in faeces compared with uninfected reindeer. Faecal corticosterone levels were higher than faecal cortisol levels, and only corticosterone increased significantly after a handling event. These results suggest that reindeer may use a tolerance strategy to cope with gastrointestinal nematodes and raise the question as to whether moderate infection intensities with nematodes are beneficial to the host. By removing nematodes we may have altered the gut microbiota, leading to the observed elevated faecal glucocorticoid metabolite levels in the treated reindeer. These findings demonstrate the importance of considering both cortisol and corticosterone in physiological studies, as there is mounting evidence that they may have different functionalities.

Occurrence of Mycobacterium avium subspecies paratuberculosis and Neospora caninum in Alberta cow-calf operations.

Mycobacterium avium subsp. paratuberculosis (MAP) and Neospora caninum (NC) are two pathogens causing important production limiting diseases in the cattle industry. Significant impacts of MAP and NC have been reported on dairy cattle herds, but little is known about the importance, risk factors and transmission patterns in western Canadian cow-calf herds. In this cross-sectional study, the prevalence of MAP and NC infection in southwest Alberta cow-calf herds was estimated, risk factors for NC were identified, and the reproductive impacts of the two pathogens were assessed. Blood and fecal samples were collected from 840 cows on 28 cow-calf operations. Individual cow and herd management information was collected by self-administered questionnaires and one-on-one interviews. Bayesian estimates of the true prevalence of MAP and NC were computed, and bivariable and multivariable statistical analysis were done to assess the association between the NC serological status and ranch management risk factors, and the clinical effects of the two pathogens. Bayesian estimates of true prevalence indicated that 20% (95% probability interval: 8-38%) of herds had at least one MAP-positive cow, with a within-herd prevalence in positive herds of 22% (8-45%). From the Bayesian posterior distributions of NC prevalence, the median herd-level prevalence was 66% (33-95%) with 10% (4-21%) cow-level prevalence in positive herds. Multivariable analysis indicated that introducing purchased animals in the herd might increase the risk of NC. The negative association of NC with proper carcass disposal and presence of horses on ranch (possibly in relation to herd monitoring and guarding activities), may suggest the importance of wild carnivores in the dynamics of this pathogen in the study area. We also observed an association between MAP and NC serological status and the number of abortions. Additional studies should be done to further examine specific risk factors for MAP and NC, assess the consequences on the reproductive performances in cow-calf herds, and evaluate the overall impact of these pathogens on cow-calf operations.

What attracts elk onto cattle pasture? Implications for inter-species disease transmission.

In Southwest Alberta, beef cattle and wild elk (Cervus elaphus) have similar habitat preferences. Understanding their inter-species contact structure is important for assessing the risk of pathogen transmission between them. These spatio-temporal patterns of interactions are shaped, in part, by range management and environmental factors affecting elk distribution. In this study, resource selection modeling was used to identify factors influencing elk presence on cattle pasture and elk selection of foraging patches; furthermore, consequences for inter-species disease transmission were discussed. Data on pasture management practices and observations of elk were collected from 15 ranchers during interviews. Pasture use by elk was defined based on telemetry data (from GPS collars deployed on 168 elk in 7 herds) and rancher observations. At the patch scale, foraging patches used by elk were identified by spatio-temporal cluster analysis of telemetry data, whereas available patches were randomly generated outside the area delimited by used patches. For pastures and patches, landscape and human-managed features were characterized using remote sensing data and interviews, respectively. Attributes of available and used pastures (or patches) were compared using resource selection functions, on annual and seasonal (or annual and monthly) time scales. Additionally, intensity of pasture use was modeled using negative binomial regression. Cultivated hay land and mineral supplements were associated with elk presence on cattle pastures, whereas pastures with manure fertilization and higher traffic-weighted road densities were less likely to be used by elk. The effects of landscape (elevation, aspect, water access) and vegetation (forest cover, Normalized Difference Vegetation Index) characteristics on patch selection were consistent with typical elk habitat requirements. The presence of cattle and the traffic-weighted road density were negatively associated with patch selection. The apparent avoidance of cattle by elk reduced the risk of direct transmission of pathogens, except during winter months. However, human-managed features attracting elk to cattle pastures (e.g. hay land and mineral supplements) may increase inter-species pathogen transmission through indirect contacts.

Calcium channel antagonist exposures reported to the Poisons Information Center Erfurt.

BACKGROUND: We investigated the toxicity profile of the three main groups of calcium channel antagonists (CCA) and compared mixed CCA exposures (CCA plus another drug) with mono CCA exposures. METHODS: All CCA exposures reported to the PIC Erfurt from 2000 to 2009 were analyzed retrospectively. RESULTS: In total, 727 (230 mono and 497 mixed) CCA exposures were registered. Although CCA exposures increased almost twofold from 56 in 2000 to 108 in 2009 their relative frequency to all exposures remained constant. The five CCAs most frequently involved in exposures were the five most frequently prescribed ones in Germany over the same period. In mono and mixed CCA exposures, none or minor symptoms were most often seen with dihydropyridines (mono: 84.7%; mixed: 68.0%) followed by diltiazem (mono: 71.4%; mixed: 62.5%) and verapamil (mono: 57.1%; mixed: 50.0%). Highest rates of moderate (mono: 8.6%: mixed: 20.2%) and severe symptoms (mono: 18.6%; mixed: 23.7%) were observed after verapamil ingestions. Death most frequently occurred with diltiazem (mono: 28.6%; mixed: 12.5%). Rates of moderate symptoms were higher in mixed (13.3%) than in mono CCA exposures (4.8%). No distinct differences were seen regarding the relative frequency of none or minor symptoms, severe symptoms, and death between mono and mixed CCA exposures. CONCLUSION: Exposures to verapamil more often resulted in moderate and severe symptoms than with dihydropyridines. Death mainly occurred with diltiazem. Moderate symptoms were more frequent in mixed than in mono CCA exposures. The frequency of CCAs involved in exposure was related to their prescription.

Pathogens of domestic and free-ranging ungulates: global climate change in temperate to boreal latitudes across North America.

In North America broad-based research networks explore the interaction of vertebrates, their characteristic arrays of pathogens and emergent disease. A diversity of programmes address the impact of environmental change on animal health, zoonoses, and human health, but as yet no comprehensive framework or strategy has emerged to develop and implement policy and planning. In a regime of climate change and ecological perturbation, the need to document and understand the health, agricultural, societal and economic impact of pathogens and emerging infectious disease is urgent. An integrated and proactive planning process linking national and international resources can lead to informed predictions aboutthe impact of environmental change and can identify pathways for potential management and mitigation. An effective and comprehensive programme will have components for establishing priorities, developing primary data for faunal structure and biodiversity, a capacity for monitoring and surveillance (including scanning and targeted activities), and linkage to historical and contemporary baselines (against which to assess change) established through archival biological collections. Field and laboratory studies are also necessary to determine developmental thresholds, tolerances and tipping points for many pathogens to establish a context for recognising current constraints and future perturbation, and to explore factors that promote emergence for a variety of pathogens, vectors and pest species. Predictive modelling and risk assessment utilising a range of scenarios for climate change is a final step in this multidisciplinary process.

Protostrongylid parasites and pneumonia in captive and wild thinhorn sheep (Ovis dalli).

We describe health significance of protostrongylid parasites (Parelaphostrongylus odocoilei and Protostrongylus stilesi) and other respiratory pathogens in more than 50 naturally infected Dall's sheep (Ovis dalli dalli) from the Mackenzie Mountains, Northwest Territories (1998-2002) as well as in three Stone's sheep (O. d. stonei) experimentally infected with P. odocoilei (2000-2002). Histological lesions in the brain and distribution of P. odocoilei in the muscles of experimentally and naturally infected sheep were consistent with a previously hypothesized "central nervous system to muscle" pattern of migration for P. odocoilei. Dimensions of granulomas associated with eggs of P. odocoilei and density of protostrongylid eggs and larvae in the cranial lung correlated with intensity of larvae in feces, and all varied with season of collection. Prevalence of P. stilesi based on the presence of larvae in feces underestimated true prevalence (based on examination of lungs) in wild Dall's sheep collected in summer and fall. Similarly, counts of both types of protostrongylid larvae in feces were unreliable indicators of parasitic infection in wild Dall's sheep with concomitant bacterial pneumonia associated with Arcanobacterium pyogenes, Pasteurella sp., and Mannheimia sp. Diffuse, interstitial pneumonia due to P. odocoilei led to fatal pulmonary hemorrhage and edema after exertion in one experimentally infected Stone's sheep and one naturally infected Dall's sheep. Bacterial and verminous pneumonia associated with pathogens endemic in wild Dall's sheep in the Mackenzie Mountains caused sporadic mortalities. There was no evidence of respiratory viruses or bacterial strains associated with domestic ruminants, from which this population of wild sheep has been historically isolated.

Bionomics of larvae of Parelaphostrongylus odocoilei (Nematoda: Protostrongylidae) in experimentally infected gastropod intermediate hosts.

Parelaphostrongylus odocoilei is a protostrongylid parasite that has recently been recognized at several locations in sub-Arctic, but not Arctic, North America. We investigated factors that may determine the distribution of P. odocoilei, including suitable gastropod intermediate hosts, temperature requirements for larval development in gastropods, and larval emergence facilitating overwinter transmission. We collected and experimentally infected gastropods from a site in the sub-Arctic where P. odocoilei is at the northern limit of its distribution. Deroceras laeve, Catinella sp., and Euconulus cf fulvus, but not members of the Pupillidae, were suitable intermediate hosts. We describe bionomics of larvae of P. odocoilei in D. laeve and Catinella sp. Infective larvae emerged from all slugs (D. laeve) and 60% of Catinella sp. snails, and emergence from D. laeve was intensity dependent. Emerged infective larvae survived up to 6 mo under conditions approximating that of the subnivean environment. In D. laeve, there was a direct relationship between temperature and development rate of larvae of P. odocoilei. Larvae of P. odocoilei did not develop to infective stage below the theoretical threshold (8.5 C), and required a minimum of 163 degree days to complete development. These developmental parameters can be incorporated into a model to predict larval development in the field. Knowledge of the factors influencing larval bionomics provides the foundation for predicting temporal and spatial patterns of parasite distribution, abundance, and transmission.

Climate change and the epidemiology of protostrongylid nematodes in northern ecosystems: Parelaphostrongylus odocoilei and Protostrongylus stilesi in Dall's sheep ( Ovis d. dalli ).

We describe the epidemiology of the protostrongylid parasites Parelaphostrongylus odocoilei and Protostrongylus stilesi in Dall's sheep (Ovis dalli dalli) from the Mackenzie Mountains, Northwest Territories, Canada (65 degrees N; 128 degrees W). Peak numbers of 1st-stage larvae of both parasites were shed by Dall's sheep on their winter range from March until May. In larval development experiments in the Mackenzie Mountains, peak numbers of infective 3rd-stage larvae of P. odocoilei were available in gastropod intermediate hosts in August-September. For both protostrongylids, the majority of transmission likely occurs on the winter range, with infection of gastropods when they emerge from hibernation in spring, and infection of Dall's sheep upon their return in fall. We validated a degree-day model for temperature-dependent development of larval P. odocoilei in gastropods, and applied degree-day models to describe and predict spatial and temporal patterns in development of P. odocoilei and P. stilesi in northern North America. Temperature-dependent larval development may currently limit northward range expansion of P. odocoilei into naïve populations of Dall's sheep in the Arctic, but climate warming may soon eliminate such constraints. In Subarctic regions where both P. odocoilei and P. stilesi are endemic, the length of the parasite 'growing season' (when temperatures were above the threshold for larval development) and amount of warming available for parasite development has increased over the last 50 years. Further climate warming and extension of the seasonal window for transmission may lead to amplification of parasite populations and disease outbreaks in host populations.

Global warming is changing the dynamics of Arctic host-parasite systems.

Global climate change is altering the ecology of infectious agents and driving the emergence of disease in people, domestic animals, and wildlife. We present a novel, empirically based, predictive model for the impact of climate warming on development rates and availability of an important parasitic nematode of muskoxen in the Canadian Arctic, a region that is particularly vulnerable to climate change. Using this model, we show that warming in the Arctic may have already radically altered the transmission dynamics of this parasite, escalating infection pressure for muskoxen, and that this trend is expected to continue. This work establishes a foundation for understanding responses to climate change of other host-parasite systems, in the Arctic and globally.

Hepatic gene expression patterns in thyroid hormone-treated hypothyroid rats.

Thyroid hormone (T3) is essential for normal development, differentiation and metabolic balance. We have performed DNA microarray experiments using hepatic RNA from hypothyroid and T3-treated hypothyroid rats in order to characterize T3-induced gene expression patterns after various time points (6, 24 and 48 h after the administration of the hormone). Sixty-two of 4608 different genes displayed a reproducible T3-response, and cluster analysis divided these differentially regulated genes into six expression patterns. Thirty-six genes were not significantly regulated within the first 24 h. Transient transfection experiments of eight late-induced gene promoters failed to detect a thyroid hormone response element within their regulatory elements, suggesting an indirect activation mechanism(s). In search for an intermediate factor of T3 action, we examined whether various rather ubiquitous transcription factors, peroxisome proliferator-activated receptors (PPARs) and coactivators of the PPARgamma coactivator 1 family (PGC-1) are regulated by T3. Only PPARgamma and PERC/PGC-1beta exhibit a significant T3-response within the first 6 h after treatment, identifying these factors as candidate components for mediating the late-induced expression pattern. Regulation of early-induced genes within the first 6 h after administration of T3 on transcript levels correlates with altered protein levels after 24 and 48 h in vivo.

TGF-beta1-induced PAI-1 gene expression requires MEK activity and cell-to-substrate adhesion.

The type-1 inhibitor of plasminogen activator (PAI-1) is an important physiological regulator of extracellular matrix (ECM) homeostasis and cell motility. Various growth factors mediate temporal changes in the expression and/or focalization of PAI-1 and its protease target PAs, thereby influencing cell migration by barrier proteolysis and/or ECM adhesion modulation. TGF-beta1, in particular, is an effective inducer of matrix deposition/turnover, cell locomotion and PAI-1 expression. Therefore, the relationship between motility and PAI-1 induction was assessed in TGF-beta1-sensitive T2 renal epithelial cells. PAI-1 synthesis and its matrix deposition in response to TGF-beta1 correlated with a significant increase in cell motility. PAI-1 expression was an important aspect in cellular movement as PAI-1-deficient cells had significantly impaired basal locomotion and were unresponsive to TGF-beta1. However, the induced migratory response to this growth factor was complex. TGF-beta1 concentrations of 1-2 ng/ml were significantly promigratory, whereas lower levels (0.2-0.6 ng/ml) were ineffective and final concentrations > or =5 ng/ml inhibited T2 cell motility. This same growth factor range progressively increased PAI-1 transcript levels in T2 cells consistent with a bifunctional role for PAI-1 in cell migration. TGF-beta1 induced PAI-1 mRNA transcripts in quiescent T2 cells via an immediate-early response mechanism. Full TGF-beta1-stimulated expression required tyrosine kinase activity and involved MAPK/ERK kinase (MEK). MEK appeared to be a major mediator of TGF-beta1-dependent PAI-1 expression and T2 cell motility since PD98059 effectively attenuated both TGF-beta1-induced ERK1/2 activation and PAI-1 transcription as well as basal and growth factor-stimulated planar migration. Since MEK activation in response to growth factors is adhesion-dependent, it was important to determine whether cellular adhesive state influenced TGF-beta1-mediated PAI-1 expression in the T2 cell system. Cells maintained in suspension culture (i.e., over agarose underlays) in growth factor-free medium or treated with TGF-beta1 in suspension expressed relatively low levels of PAI-1 transcripts compared with the significant induction of PAI-1 mRNA evident in T2 cells upon stimulation with TGF-beta1 during adhesion to a fibronectin-coated substrate. Attachment to fibronectin alone (i.e., in the absence of added growth factor) was sufficient to initiate PAI-1 transcription, albeit at levels considerably lower than that induced by the combination of cell adhesion in the presence of TGF-beta1. T2 cells allowed to attach to vitronectin-coated surfaces also expressed PAI-1 transcripts but to a significantly reduced extent relative to cells adherent to fibronectin. Moreover, newly vitronectin-attached cells did not exhibit a PAI-1 inductive response to TGF-beta1, at least during the short 2 hour period of combined treatment. PAI-1 mRNA synthesis in response to substrate attachment, like TGF-beta1-mediated induction in adherent cultures, also required MEK activity as fibronectin-stimulated PAI-1 expression was effectively attenuated by the MEK inhibitor PD98059. These data indicate that cellular adhesive state modulates TGF-beta1 signaling to particular target genes (i.e., PAI-1) and that MEK is a critical mediator of the PAI-1(+)/promigratory phenotype switch induced by TGF-beta1 in T2 cells.

Hormonal regulation of multiple promoters of the rat mitochondrial glycerol-3-phosphate dehydrogenase gene: identification of a complex hormone-response element in the ubiquitous promoter B.

Rat mitochondrial glycerol-3-phosphate dehydrogenase (mGPDH) is regulated by multiple promoters in a tissue-specific manner. Here, we demonstrate that thyroid hormone (3,5,3'-tri-iodo-L-thyronine) and steroid hormone but not the peroxisome proliferator clofibrate and retinoic acid stimulate the activation of the ubiquitous promoter B in a receptor-dependent manner, whereas the more tissue-restricted promoters A and C are not inducible by these hormones. Thyroid hormone action is mediated by a direct repeat +4 (DR+4) hormone-response element as identified by deletion and mutation analyses of promoter B in transient transfection analyses. The DR+4 element was able to bind to an in vitro translated thyroid hormone receptor in band-shift and supershift experiments. The hormone-response element comaps with a recognition site for the transcription factor Sp1, suggesting complex regulation of this sequence element. Mutation of this Sp1-recognition site reduces the basal promoter B activity dramatically in HepG2 and HEK293 cells in transient transfection and abolishes the binding of Sp1 in band-shift experiments. As demonstrated by Western-blot experiments, administration of tri-iodothyronine to euthyroid rats increases hepatic mGPDH protein concentrations in vivo. As it has recently been reported that human mGPDH promoter B is not regulated by tri-iodothyronine, this is the first example of a differentially tri-iodothyronine-regulated orthologous gene promoter in man and rat.

Umingmakstrongylus pallikuukensis (Nematoda: Protostrongylidae) in gastropods: larval morphology, morphometrics, and development rates.

Morphological and morphometric aspects of larval development of Umingmakstrongylus pallikuukensis in Deroceras laeve and the effects of temperature on development rates in D. laeve and Deroceras reticulatum were investigated in the laboratory. Larval stages were best differentiated by separation of cuticular sheaths, tail structure, and viability following digestion. Growth in body and esophagus width was observed during the first-stage within the intermediate host, but the major increases in body length and width occurred immediately following the second molt. Larval development in D. laeve and D. reticulatum occurred more rapidly at warmer temperatures. The calculated threshold temperatures were 8.5 and 9.5 C in D. laeve and D. reticulatum, respectively, and 167 degree-days were required for development to third-stage larvae (L3) in both hosts. These thresholds are higher than those calculated from published data for the closely related Muellerius capillaris (4.2 C) but are similar to those for the more distantly related northern protostrongylid, Elaphostrongylus rangiferi (8.3-10.3 C). Conversely, degree-days required for development to infective L3 were more similar among the Muelleriinae than between this group and the Elaphostrongylinae. Developmental parameters for protostrongylid larvae may be influenced both by the environment and by features of the parasites and the intermediate hosts, including phylogeny.