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Biomedical subjects

J P Morin

Publications and source records attributed to J P Morin.

At least 19 recordsLinked to original sources

Comparative analysis of morphological traits among Drosophila melanogaster and D. simulans: genetic variability, clines and phenotypic plasticity.

The two sibling cosmopolitan species, Drosophila melanogaster and D. simulans, are able to proliferate under very different climatic conditions. This has resulted in local adaptations, which are often arranged in latitudinal clines. Such clines are documented for body weight, wing and thorax length, sternopleural and abdominal bristle number, ovariole number and thoracic pigmentation. The overall magnitude of geographical differentiation is, however, much less in D. simulans than in D. melanogaster, and latitudinal clines are less pronounced. The fact that natural populations live under different climates raises the problem of interaction between temperature and phenotype. The reaction norms of morphometrical traits have been investigated as a function of growth temperature. The shapes of the response curves vary according to the investigated trait. They are generally curvilinear and can be described by calculating characteristic values after polynomial adjustments. For a given trait, the reaction norms of the two species are similar in their shape, although some significant differences may be observed. Within each species, significant differences are also observed between geographic populations: reaction norms are not parallel and the divergence is better marked when more distant populations (e.g., temperate and tropical) are compared. It thus appears that besides mean trait value, phenotypic plasticity is also a target of natural selection. A specific analysis of wing shape variation according to growth temperature was also undertaken. Reaction norms with different shapes may be observed in various parts of the wing: the major effect is found between the basis and the tip of the wing, but in a similar way in the two species. By contrast, some ratios, called wing indices by taxonomists, may exhibit completely different reaction norms in the two species. For a single developmental temperature (25 degrees C) the phenotypic variability of morphometrical traits is generally similar in the two species, and also the genetic variability, estimated by the intraclass correlation. A difference exists, however, for the ovariole number which is less variable in D. simulans. Variance parameters may vary according to growth temperature, and a detailed analysis was made on wing dimensions. An increase of environmental variability at extreme, heat or cold temperatures, has been found in both species. Opposite trends were, however, observed for the genetic variability: a maximum heritability in D. simulans at middle temperatures, corresponding to a minimum heritability in D. melanogaster. Whether such a difference exists for other traits and in other populations deserves further investigations. In conclusion, morphometrical analyses reveal a large amount of significant differences which may be related to speciation and to the divergence of ecological niches. Within each species, numerous geographic variations are also observed which, in most cases, reflect some kinds of climatic adaptation.

Animals↗

Rotating wall vessel as a new in vitro shear stress generation system: application to rat coronary endothelial cell cultures.

In this paper, we describe a simple new design for the application of controlled, top-hat profiled wall shear stress forces in a way that is independent of hydrostatic pressure and oxygen tension, based on a rotating wall vessel system. This system has been applied to the culture of rat coronary endothelial cells obtained with a Langendorff-derived procedure isolation. Endothelial cells are immunopurified on the basis of RECA expression, and conservation of endothelial phenotype has been assessed on the basis of morphology, RECA and von Willebrand factor expressions and diI-Ac-LDL uptake. Shear stress induced by the rotating wall vessel was measured using a mathematical formula specifically designed for this type of model, and its impact on coronary endothelial cells was evaluated. Shear stress produced cell orientation parallel to the flux direction, elevated NO production and decreased monocyte adhesion. Cells were kept viable and functional for at least 4 days under shear. This simple design allows the handling and management of numerous vials in parallel and appears to be suitable for large-scale studies of both the acute and chronic impact of modulation of the physico-chemical environment on endothelial cell physiology and function.

Animals↗

Biphasic culture of rat lung slices for pharmacotoxicological evaluation of complex atmospheres.

The relevance to the in vivo situation of in vitro toxicity studies of complex atmospheres has frequently been limited by the procedures used for the exposure of the biological samples. We have evaluated from on-road measurements the size distribution pattern and the subsequent respiratory tract deposition rates of particulate matter from urban atmospheric aerosols, which are in the range of 110 and 3 pg/cm2 per min for tracheobronchial and alveolar areas, respectively. Continuous flow-through rotating chambers and a specific design for exhaust sampling and dilution with controlled adjustment of pO2 and pCO2 to 20% and 5%, respectively, have been developed to expose biphasic air/liquid organotypic cultures of rat lung slices to continuous flows of diluted exhausts from diesel engines with preservation of the physicochemical properties of the exhaust. The size distribution of the particulate matter and the bioavailability of pollutants were preserved, thus allowing us to closely mimic in vitro the in vivo atmosphere/tissue interactions that occur mainly through diffusion mechanisms. The toxicity response profile has been assessed in terms of tissue viability, oxidative stress, DNA injury, and the early phase of inflammatory reaction. Exhaust filtration, addition to fuel of rapeseed methyl ester, and preincubation of lung tissue with soy isoflavones modulated the toxicity response profile of exhausts. The importance of preserving both particulate matter size distribution and adsorbed pollutant bioavailability, which could not be ascertained using more classical in vitro approaches, is discussed and should be considered a prerequisite for further developments of in vitro studies to modelize in vivo inhalation of complex atmospheres.

Aerosols↗

Seasonality and genetic architecture of development time and body size of the birch feeding sawfly Priophorus pallipes.

We tested, using the sawfly Priophorus pallipes feeding on leaves of mountain birch, whether the expression of genetic (co)variation of larval development time and body size can be altered by exposing larvae to diets with differential seasonal changes in quality. In nature, larvae feed mainly on mature leaves, but occasionally they are forced to consume senescing leaves. Sixty families were assayed on three experimentally simulated diets: mature leaves of high quality, senescing leaves of rapidly declining quality, and senesced leaves of low quality. The intuitively obvious positive phenotypic and genetic correlations between development time and final mass were observed when the larvae consumed leaves of stable high quality, but low and declining food quality prevented long-growing individuals and families from achieving high final mass, switching the correlations to close to zero or negative in these treatments. The amount of genetic variation for body size showed a non-linear change across the diet quality gradient, whereas genetic variation for development time increased with decreasing diet quality. The among-trait difference in the degree reaction norms crossed along the diet gradient caused the changes in the expression of genetic (co)variation within the environments. Our results show that seasonally varying diet quality induces dramatic changes in the genetic (co)variation of development time and body size, and that simultaneous analysis of reaction norms and environment-specific expression of genetic (co)variation is necessary for the understanding of the genetic characteristics underlying the construction of phenotypes in heterogeneous environments.

Analysis of Variance↗

Seasonally varying diet quality and the quantitative genetics of development time and body size in birch feeding insects.

Genetic variance-covariance structures (G), describing genetic constraints on microevolutionary changes of populations, have a central role in the current theories of life-history evolution. However, the evolution of Gs in natural environments has been poorly documented. Resource quality and quantity for many animals and plants vary seasonally, which may shape genetic architectures of their life histories. In the mountain birch-insect herbivore community, leaf quality of birch for insect herbivores declines profoundly during both leaf growth and senescence, but remains stable during midsummer. Using six sawfly species specialized on the mountain birch foliage, we tested the ways in which the seasonal variation in foliage quality of birch is related to the genetic architectures of larval development time and body size. In the species consuming mature birch leaves of stable quality, that is, without diet-imposed time constraints for development time, long development led to high body mass. This was revealed by the strongly positive phenotypic and genetic correlations between the traits. In the species consuming growing or senescing leaves, on the other hand, the rapidly deteriorating leaf quality prevented the larvae from gaining high body mass after long development. In these species, the phenotypic and genetic correlations between development time and final mass were negative or zero. In the early-summer species with strong selection for rapid development, genetic variation in development time was low. These results show that the intuitively obvious positive genetic relationship between development time and final body mass is a probable outcome only when the constraints for long development are relaxed. Our study provides the first example of a modification in guild-wide patterns in the genetic architectures brought about by seasonal variation in resource quality.

Animal Feed↗

Role of solid-phase microextraction in the identification of highly volatile pheromones of two Rhinoceros beetles Scapanes australis and Strategus aloeus (Coleoptera, Scarabaeidae, Dynastinae).

Solid-phase microextraction (SPME) samplings from live insects or natural secretion allowed one to identify the aggregation pheromones of the pest beetles Scapanes australis and Strategus aloeus by efficient and rapid isolation of their highly volatile (72 < M(r) < 116) components. S. australis male pheromone was identified as a 84:12:4 (w/w) mixture of 2-butanol [67:33 (R)-(-):(S)-(+) ratio], 3-hydroxy-2-butanone and 2,3-butanediol [43:17:40 (R,R)-(-):(S,S)-(+):meso ratio], and S. aloeus pheromone as a 95.5:4.0:0.5 (w/w) mixture of 2-butanone, 3-pentanone and sec.-butyl acetate by GC-MS using conventional and chiral capillary columns. This is the first report of Scarabaeidae pheromones based on such small and common molecules.

Animals↗

Toxicity of diesel engine exhausts in an in vitro model of lung slices in biphasic organotypic culture: induction of a proinflammatory and apoptotic response.

Precision-cut rat lung slices in organotypic culture placed in a biphasic air/liquid system were used for this study. This model allowed pathological as well as cellular and molecular biology investigations to be carried out. Slices were exposed to a continuous flow of diluted diesel exhaust, with a pO2 adjusted to 20% to avoid hypoxia-induced effects. The exposure system allowed five exhaust concentrations from the same diesel engine to be studied concomitantly, and also allowed the impact of removing the particulate matter using a filter cap on the exposure vials to be evaluated. Lung slices were exposed for 3 or 6 h to whole or filtered diesel exhaust. DNA integrity was characterized by two different techniques: (1) an ELISA for the determination of nucleosomes, and (2) the histochemical TUNEL method. By the TUNEL method, apoptotic cells were detected after a 6-h exposure followed by an incubation period of 18 h in a controlled atmosphere comprising 5% CO2/95% O2. Under these conditions, apoptotic nuclei were more frequent in slices exposed to diesel exhaust than in control slices. Cytokine production (tumor necrosis factor alpha, interleukin-1beta) in the culture medium was measured using an ELISA technique. After a 3-h exposure only TNF-alpha was detected and increased in the culture medium of lung slices exposed to diesel exhaust. Under the same conditions, nucleosome levels in the slices increases in a dose-dependent way. In conclusion, whole diesel exhaust induced an inflammatory response and DNA alterations which were reduced by filtration, thus indicating the important role of the particulate matter in diesel exhaust.

Animals↗

The genetics of phenotypic plasticity. IX. Genetic architecture, temperature, and sex differences in Drosophila melanogaster.

We examined the genetic architecture of plasticity of thorax and wing length in response to temperature in Drosophila melanogaster. Reaction norms as a function of growth temperature were analyzed in 20 isofemale lines in a natural population collected from Grande Ferrade near Bordeaux (southern France) in two different years. We found evidence for a complex genetic architecture underlying the reaction norms and differences between males and females. Reaction norms were negative quadratics. Genetic correlations were moderately high between traits within environments. Among characteristic values, the magnitudes of genetic correlations varied among traits and sexes. We hypothesized that genetic correlations among environments would decrease as temperatures became more different. This expectation was upheld for only one trait, female thorax length. For males for both traits, the correlations were large for both very similar and very different temperatures. These correlations may constrain the evolution of the shape of the reaction norms. Whether the extent of independence implies specific regulatory genes or only a specific allelic regulation of trait genes can not be decided from our results.

Animals↗

Acrolein toxicity: comparative in vitro study with lung slices and pneumocytes type II cell line from rats.

Toxicological effects of acrolein have been studied in precision-cut rat lung slices and in L2 cells, a rat pneumocyte II cell line. These two models were cultured for 24 h with or without acrolein (0-100 microM in L2 cells; 0-200 microM in lung slices). Treatment with this pneumotoxicant produced a concentration dependent decrease in intracellular ATP levels. Acrolein concentrations higher than 50 microM induced ATP decrease in slices, while this decrease occurred from 10 microM acrolein in L2 cells. Detoxification marker evaluations showed that mostly the glutathione pathway was altered after acrolein treatment in both models. Intracellular glutathione (GSH) levels were drastically increased with an acrolein concentration of 10 microM. This increase was concomitant with glutathione-S-transferase (GST) and glutathione reductase (GRED) activities in L2 cells. After this strong increase, these enzymatic activities as well as GSH levels were quickly decreased. In precision-cut rat lung slices, the induction of the glutathione pathway was less clear-cut. A bell-shaped dose response curve was observed with a maximum for 5 microM acrolein for GST and GRED activities. These differences between acrolein toxic ranges could be explained by the presence of an active detoxification pathway in slices compared to its relative lack in L2 cells.

Acrolein↗

Development of a new in vitro system for continuous in vitro exposure of lung tissue to complex atmospheres: application to diesel exhaust toxicology.

The purpose of this study was the development of a new incubation system that can allow continuous exposure of lung tissue to complex atmospheres as a tool for the assessment of aerial environmental lung toxicology. To assess the pertinence of this new exposure system, we studied the impact of diesel engine exhausts as a complex atmosphere containing both gaseous and particulate fractions and have been able to discriminate between the toxicological impacts of the gaseous phase and particulate matter from diesel exhausts. Continuous flow-through rotating chambers with controlled PO2, pCO2, and hygrometry have been designed in which lung slices are positioned in rolling inserts that allow free access of atmosphere to the exposed lung tissue. Under control conditions, cell viability was preserved for at least 48 h as assessed by intracellular ATP, GSH, and K+ levels and slice O2 consumption levels. Short-term exposure (1 h) to diesel whole exhausts did not affect intracellular potassium or slice O2 consumption, while intracellular ATP and GSH levels were markedly decreased. Exposure to filtered exhausts showed less marked effects on both ATP and GSH levels. Superoxide dismutase activity was decreased in a similar way by both total and filtered exhausts while Se(+)-dependent glutathione peroxidase activity was induced by filtered exhausts to a larger extent than after total exhaust exposure, showing different response patterns of lung tissue after exposure to whole or filtered exhausts. In conclusion, this newly designed model opens a promising area in in vitro environmental lung toxicology testing.

Adenosine Triphosphate↗

Morphological and biochemical characterization of primary culture of rabbit proximal kidney tubule cells grown on collagen-IV coated Millicell-CM.

The aim of this study was to better characterize rabbit proximal kidney tubule cells cultured on collagen IV-coated porous inserts, as compared to the same cells seeded in standard plastic wells. Total protein contents in confluent monolayers on permeable membranes were about twofold higher than those measured in confluent cultures in plastic wells. Microscopy examinations suggested that such a difference was probably due to a higher cell density and to an impressive development of the apical brush-border membrane. Moreover, measurement of unidirectional transport of p-aminohippuric acid and tetraethylammonium bromide confirmed the high polarization level of cultures on porous inserts. Results of methyl(alpha-D-[U-14C]glyco)pyranoside uptake suggested that cell phenotype was probably influenced by culture conditions. Analysis of different markers as a function of time in culture showed decreases of alkaline phosphatase (AP), gamma-glutamyltranspeptidase (GGT), and Na(+)-K(+)-ATPase activities as well as increases in LDH, ATP, and glutathione levels, similar to those formerly reported for cells cultured in standard plastic plates. However, comparative data from 6-d-old monolayers have shown that AP, GGT, Na(+)-K(+)-ATPase, glutathione reductase (GRED), and selenium-dependent glutathione peroxidase (Se-GPX) activities were 2.8-, 2.6-, 1.6-, 1.2-, and 2.1-fold, respectively, better preserved on precoated permeable membranes. On the other hand, this paper reports for the first time in the literature that GRED and SE-GPX, two phase II detoxification enzymes, were well maintained in cultures of rabbit proximal kidney tubule cells. Our results show that culturing rabbit proximal kidney tubule cells on collagen IV-coated porous membranes was accompanied by an improvement of both morphological and biochemical properties of the cells.

Adenosine Triphosphate↗

Cell lines with extended in vitro growth potential from human renal proximal tubule: characterization, response to inducers, and comparison with established cell lines.

Few model systems exist for the study of injury to human renal proximal tubule epithelium. Optimized differentiated human renal epithelial cell lines with extended in vitro growth potential would provide an alternative model system to primary culture or other available non-human mammalian kidney cell lines. For this purpose, human renal tubule epithelial cells were isolated from normal kidney cortex and exposed in culture to a hybrid immortalizing virus, adenovirus 12-SV40. Cell lines were developed by limiting dilution, and three selected cell lines were screened for growth pattern, production of immortalizing virus, tumorigenicity, and ploidy. Cell lines were also monitored for response to inducer agents and matrix factors and were screened for expression of biochemical properties and differentiation markers of renal epithelium. All three are nonproducers of the immortalizing virus and are nontumorigenic. They grow in monolayer, have intermediate growth kinetics, and express markers of renal proximal tubular epithelium by immunohistochemistry. They also express biochemical properties comparable to other widely used proximal tubular cell lines including LLC-RK1, OK, and HK-2 and comparable to human tubular cells in stable culture. Growth medium containing low levels of fetal calf serum, or epidermal growth factor combined with parathyroid hormone, produced optimal growth characteristics, brush border enzyme expression, biochemical properties, and glucose transport in a selected cell line. The addition of dimethyl sulfoxide allows maintenance in morphologically intact monolayers for prolonged periods. These cell lines should be useful model systems for the study of human renal proximal tubular injury or disease.

Adenosine Triphosphate↗

Evolutionary changes of nonlinear reaction norms according to thermal adaptation: a comparison of two Drosophila species.

While the adaptive significance of discontinuous reaction norms is generally accepted, the evolutionary interpretation of continuous response curves remains speculative, and the occurrence of internal constraints is often suggested as an explanation of experimental observations. In Drosophila melanogaster, various morphometrical traits exhibit convex reaction norms to growth temperature, with a maximum value within the developmental thermal range. We compared a cold-adapted species (D. subobscura) with a mid thermal range at 16 degrees C, to the warm-adapted D. melanogaster (mid thermal range at 22 degrees C) for three different morphometrical traits: wing and thorax length in both sexes and ovariole number in females. Maximum value temperatures were ordered in the same way for the three traits in both species: ovariole number > thorax length > wing length. Significant differences were also observed between the two species for the curvature parameter of the quadratic adjustment. The major observation was a significant lateral shift in the reaction norms: maximum values were observed at much lower temperatures in the cold-adapted species than in the warm-adapted one. The parallelism between mid thermal range variation and the position of the maximum value strongly suggests an adaptive displacement of the response curves. Natural selection may thus act not only on trait mean values but also on phenotypic plasticity and on the shape of reaction norms.

Animals↗

Comparative impact of cephaloridine on glutathione and related enzymes in LLC-PK1, LLC-RK1, and primary cultures of rat and rabbit proximal tubule cells.

Among kidney tubular epithelial cell types, proximal tubule cells are one of the major renal targets for xenobiotics. Several in vitro culture models have been proposed for use of proximal tubule cells for in vitro pharmacotoxicology studies. This paper reports a comparative study of the response to cephaloridine exposure of two established cell lines from pig (LLC-PK1) and rabbit (LLC-RK1) kidneys and primary cultures of rat and rabbit proximal tubule cells. These cultured cells were first compared for their levels of activity of alpha-methylglucopyranoside transport, alkaline phosphatase, succinate dehydrogenase, and NADPH cytochrome c reductase, their glutathione-dependent activity levels, and their adenylate cyclase response pattern to stimulation by PTH and AVP. The results presented show major phenotypic differences between these four cellular models. The differences observed in glutathione-dependent mechanism activities and regulation may in part be responsible for the variability of the responses of these four cellular models when exposed to cephaloridine.

Animals↗