Modulation of glycolysis induction in primary cultures of rabbit kidney proximal tubule cells: the role of shaking,glucose and insulin.

We have assessed the impact of increasing oxygen availability on cellular phenotype expression of rabbit proximal tubule cells in primary culture developed with variable glucose and/or insulin contents. To mitigate hypoxia at the cell/medium interface, cells were shaken for the whole culture duration and their expressed phenotype was compared with those expressed by static cultures. O₂ and CO₂ tensions were kept constant in the incubator atmosphere. Glycolysis and gluconeogenesis pathways, detoxication system, and mitochondrial, apical and basolateral membrane marker enzyme activities were assessed. This study showed that the induction of glycolysis which appear in primary cultures of proximal tubule cells may be partially prevented by continuously shaking the cultures. This effect was more marked in the presence of glucose, suggesting better substrate oxidation in shaken cultures.     

A comparison of modeling techniques to predict juvenile 0+ fish species occurrences in a large river system

Even if European river management and restoration are largely supported by the use of reliable tools, these tools are most often “generalist” and provide only initial leads of alteration sources. Acknowledging that young-of-the-year (YOY) fish assemblages are highly dependent on riverine habitat conditions, the development of a YOY-based tool might be very useful or even essential in the design and implementation of conservation or restoration plan of large rivers, in measuring more straight-forward the losses and gains of hydro-ecological functionalities. In the past 20 years, new modeling techniques have emerged from a growing sophistication of statistical model applied to ecology. “Machine learning methods” (ML) are now recognized as holding great promise for the advancement of understanding and prediction of ecological phenomena. The aim of this work was to select the appropriate statistical technique to model YOY assemblages according to different meso-scale habitat variables that are meaningful to planners. To do this, two “Machine Learning” methods, Classification and Regression Trees (CART) and Boosted Regression Trees (BRT), were compared to Generalized Linear Models (GLM). We modeled the occurrence of 9 species from the Seine River basin (France) in order to compare models abilities to accurately predict the presence and absence of each species. BRT appeared to be the best technique for modeling 0+ fish occurrences in our dataset.     

Diversity of Superoxide-Dismutases Among Clinical and Soil Isolates of Streptomyces Species

Comparison of the nature, activity, and cellular localization of superoxide-dismutases (SOD) from soil and clinical isolates of Streptomyces species was investigated to identify possible factors that could account for the pathological role of the strains isolated from human lesions. Results showed that all of the studied strains possessed a cytoplasmic Ni-SOD. This particular SOD, found in isolates from patients, could be a new taxonomic criterion to identify Streptomyces species with greater precision. A second minor SOD, assimilated to an Fe/Zn-SOD, was detected in some strains, but no relationship was established between the presence of this enzyme and the clinical origin of the strains. Received: 9 April 1999 / Accepted: 9 June 1999     

Neurotrophin-independent attraction of growing sensory and motor axons towards developing Xenopus limb buds in vitro

The mechanisms for directing axons to their targets in developing limbs remain largely unknown though recent studies in mice have demonstrated the importance of neurotrophins in this process. We now report that in co-cultures of larval Xenopus laevis limb buds with spinal cords and dorsal root ganglia of Xenopus and axolotl (Ambystoma mexicanum) axons grow directly to the limb buds over distances of up to 800 microm and in particular to sheets of epidermal cells which migrate away from the limb buds and also tail segments in culture. This directed axonal growth persists in the presence of trk-IgG chimeras, which sequester neurotrophins, and k252a, which blocks their actions mediated via trk receptors. These findings indicate that developing limb buds in Xenopus release diffusible factors other than neurotrophins, able to attract growth of sensory and motor axons over long distances.     

Rat model of pulmonary arteriovenous malformations after right superior cavopulmonary anastomosis

We developed a rat model of pulmonary arteriovenous malformations after cavopulmonary anastomosis. We sought to determine whether this model reproduces the angiographic and histologic features seen in the human condition. Eight Sprague-Dawley rats underwent a right superior cavopulmonary anastomosis with the use of microsurgical techniques. Between 2 and 13 mo, pulmonary angiography was performed, the animals were euthanized, and the lungs were removed. Microscopic sections of the lung were stained with an endothelial-specific antibody (von Willebrand factor). Microvessel density was determined by counting vessels staining positively for von Willebrand factor, and the shunted and nonshunted (control) lungs were compared for each animal. Pulmonary angiography revealed time-dependent development of arteriovenous malformations. Microvessel density demonstrated a time-dependent increase in the shunted lung compared with the control lung (simple linear regression of the ratio of the microvessel density of the shunted lung divided by the microvessel density of the control lung on time; R(2) = 0.79, P = 0.003). This animal model reproduces the same angiographic and microscopic features of pulmonary arteriovenous malformations that develop in humans after cavopulmonary anastomosis. This appears to be a valid model that may be used to further study etiologic mechanisms for this phenomenon.     

New insights into early sequential PrPsc accumulation in scrapie infected mouse brain evidenced by the use of streptomycin sulfate

To investigate the amplifying potentialities of streptomycin sulfate in the immunohistochemical (IHC) detection of the abnormal prion protein (PrPsc), we used a sequential brain sampling from C506M3 scrapie strain inoculated C57Bl/6 mice. The weekly removed brains, from 7 to 63 days post intra-cranial inoculation were analysed using PrPsc IHC. The introduction of streptomycin sulfate, a technique developed for accurate cellular and regional mapping of PrPsc deposition in several animal TSEs, revealed a substantial amplifying effect and a clear specific PrPsc detection as early as 28 days post inoculation. The location of the first detected PrPsc deposits suggests a possible involvement of the cerebrospinal fluid in the early dissemination of the infectious agent. The meaning of these newly accessible PrPsc deposits is discussed in relation to a possible nascent form of PrPsc molecules detected in situ for the first time. Altogether, these findings argue that this method can be highly useful to study the early stages after infection with prion agents.     

Differential effects of hypoxia on untreated and NGF treated PC12 cells

Perinatal hypoxia is known to induce long-lasting changes in the central dopaminergic system. In order to understand the cellular mechanism of these changes, we studied the effects of hypoxia on the levels of dopamine (DA) and tyrosine hydroxylase (TH) mRNA in untreated and NGF treated PC12 cells. On the second day after plating (DAP), cells were exposed to a hypoxic episode (pO2 = 10-20 mm Hg, 24 h), and the levels of DA and TH mRNA were examined on DAP 4 and DAP 8. In untreated cells, hypoxia induced a two fold increase both in DA and TH mRNA levels on DAP 4 which normalized up to DAP 8. This increase correlated with an activation of the hypoxia inducible factor (HIF-1alpha), measured with a reporter gene. In contrast, NGF treated cells responded to hypoxia with an increase of DA level on DAP 8. In these cells neither an increase of the HIF-1alpha activity measured immediately after hypoxia nor a significant increase of the TH mRNA level on DAP 8 were found. The findings indicate that NGF shifts the hypoxia induced changes of DA levels from a short-term to a long-term mode. The long-term increase of dopamine levels is the most likely result of changes connected with cell growth and differentiation and not the result of a long-term TH mRNA level increase.     

Temperature Threshold and Preservation of Signaling for Mitochondrial Membrane Proteins during Ischemia in Rabbit Heart

Temperature modulates both myocardial energy requirements and production. We have previously demonstrated that myocardial protection induced by hypothermic adaptation preserves expression of genes regulating heat shock protein and the nuclear-encoded mitochondrial proteins, the adenine nucleotide translocator isoform 1 (ANT₁), and the β subunit of F₁-ATPase (βF₁-ATPase). This preservation is associated with a reduction in ATP depletion similar to that noted in cardioplegic arrested hearts preserved at a critical temperature (30°C) or below. We tested the hypothesis that expression of these genes may also be subject to this temperature threshold phenomenon. Isolated perfused rabbit hearts were subjected to ischemic cardioplegic arrest at 4, 30, or 34°C for 120 min. Cardiac function indices and steady-state mRNA levels for ANT₁, βF₁-ATPase, and HSP70-1 were measured prior to ischemia (B) and after 45 min of reperfusion. Cardiac function was significantly depressed in the 34°C group. Ischemia at 34°C reduced steady-state mRNA levels for ANT₁and βF₁-ATPase from B, but these levels were similarly preserved at 4 and 30°C. HSP70-1 levels were mildly elevated (fourfold) above B to similar levels at all three temperatures. These results indicate that mRNA expression for ANT₁and βF₁-ATPase is specifically preserved in a pattern consistent with the temperature threshold phenomenon. HSP70-1 expression is not influenced by ischemic temperature. Preservation of gene expression for these mitochondrial proteins implies that signaling for mitochondrial biogenesis or resynthesis is maintained after ischemic insult.