Can associations between free fatty acid levels and metabolic parameters determine insulin resistance development in obese Zucker rats?

Elevated levels of serum free fatty acids (FFA) may be the metabolic alteration in obesity that leads to insulin resistance (IR) and type 2 diabetes mellitus (DM). The obese Zucker rat (ZR) is a genetic model of juvenile-onset obesity and type 2 DM. Compared with its lean sibling, the obese ZR is hyperinsulinemic, hypertriglyceridemic, and, beginning at about 6 months, hyperglycemic. The obese ZR demonstrates also IR, hyperphagia, increased lipogenesis, adipocyte hypertrophy and hyperplasia, and increased serum FFA levels. This study was designed to determine if serum FFA levels in lean and obese ZRs correlate with metabolic parameters associated with altered energy metabolism and IR. We hypothesized that serum FFA levels correlate with such serum parameters such as insulin, glucose, triglyceride, and total cholesterol, as well as such tissue parameters as retroperitoneal, perirenal, and epididymal fat pad weights and liver total lipid content. Twenty lean and 20 obese ZR were age/weight matched. For 14 days each rat had ad libitum access to a single bowl diet that was 50% fat, 30% carbohydrate, and 20% protein. Body weights and caloric intakes were measured daily. After 14 days, all animals were fasted overnight and euthanized. Serum and tissue measurements were made and various parameters were correlated with FFA levels. Serum FFA levels were almost 2 times higher in the obese ZR (approximately 1 mmol/L) compared to the lean (approximately 0.6 mmol/L). Each variable measured was significantly (p < or = 0.05) greater in the obese ZR compared to the lean. There were significant correlations between serum FFA levels and certain variables when data from all ZR were plotted against serum and tissue parameters. However, within phenotypes, there were no significant correlations. Serum FFA levels predict serum and tissue parameters that accompany obesity and IR when comparing lean and obese rats. However, FFA do not predict such parameters within one phenotype.     

α-, β-, and γ-Tubulin Polymerization in Response to DNA Damage

Microtubules provide structural support for a cell and play key roles in cell motility, mitosis, and meiosis. They are also the targets of several anticancer agents, indicating their importance in maintaining cell viability. We have investigated the possibility that alterations in microtubule structure and tubulin polymerization may be part of the cellular response to DNA damage. In this report, we find that γ-radiation stimulates the production and polymerization of α-, β-, and γ- tubulin in hematopoeitic cell lines (Ramos, DP16), leading to visible changes in microtubule structures. We have found that this microtubule reorganization can be prevented by caffeine, a drug that concomitantly inhibits DNA damage-induced cell cycle arrest and apoptosis. Our results support the idea that microtubule polymerization is an important facet of the mammalian response to DNA damage.     

Qualification of translational safety biomarkers

Safety biomarkers are important drug development tools, both preclinically and clinically. It is a straightforward process to correlate the performance of nonclinical safety biomarkers with histopathology, and ideally, the biomarker is useful in all species commonly used in safety assessment. In clinical validation studies, where histopathology is not feasible, safety biomarkers are compared to the response of standard biomarkers and/or to clinical adjudication. Worldwide, regulatory agencies have put in place processes to qualify biomarkers to provide confidence in the manner of use and interpretation of biomarker data in drug development studies. This paper describes currently qualified safety biomarkers which can be utilized to monitor for nephrotoxicity and cardiotoxicity and ongoing projects to qualify safety biomarkers for liver, skeletal muscle, and vascular injury. In many cases, the development and use of these critical drug development tools is dependent upon partnerships and the precompetitive sharing of data to support qualification efforts.     

Immunoreactive adrenocorticotrophin is present in the ovary and in particular the oocyte of several mammalian species.

Proopiomelanocorticotrophin (POMC)-derived peptides have been identified in both male and female reproductive systems. However, there have been few reports of ACTH, the major biologically active POMC product, in the mammalian ovary. We sought evidence for the presence and localization of immunoreactive (ir)-ACTH in ovaries from sheep, humans, cows, pigs, rats and cats using immunohistochemical techniques. Tissue sections were stained with diaminobenzidine following incubation with a primary antibody raised against ACTH1-24. There was positive staining for ACTH in cells scattered throughout the interstitium of ovaries from all species examined. Immunoreactive ACTH was observed in the oocytes of ovaries from humans, cows, pigs, pregnant and non-pregnant sheep, but not from cats or rats. Positive staining of oocytes was associated with all tertiary and secondary follicles, and some primary follicles. There was no apparent difference in the pattern of staining between pregnant and non-pregnant sheep. Staining for ir-ACTH was absent in ovaries from fetal sheep. We conclude that ir-ACTH is present in ovarian tissue, and in particular the oocyte, from several species of mammal. The presence of ir-ACTH within the oocyte is dependent on species and stage of follicular maturation.     

Rescue of retroviral envelope fusion deficiencies by cationic liposomes

Background

Retroviral particles that are inappropriately enveloped can transduce target cells if pre‐associated with cationic liposomes. This study optimises and addresses the mechanism of liposome‐enhanced gene delivery, and explores the potential for such agents to compensate for fusion deficiency associated with chimaeric envelope proteins.

Methods

Particles bearing wild‐type, chimaeric or no envelope proteins were complexed with DOTAP or DC‐Chol/DOPE cationic liposomes and added to target cells for various times. Particle binding was determined by detection of cell‐associated capsid protein and infectivity was measured histochemically.

Results

Stable association of cationic liposomes with retrovirus particles significantly enhanced their binding rate to target cells in proportion to the increase of transduction kinetics for infectious virus. Binding of virus was equivalent with or without envelope protein and/or virus receptor, indicating that a non‐specific interaction precedes receptor recognition. Non‐infectious combinations were rescued by the intrinsic fusogenicity of the cationic liposomes, which enabled entry of the viral core, but left subsequent events unaltered. The optimised transduction rate with non‐enveloped particles and DOTAP approached that of amphotropic‐enveloped virus in some cases, although the effect was target‐cell‐dependent. DC‐Chol/DOPE was less potent at direct fusion but was able to enhance 600‐fold the receptor‐dependent action of chimaeric envelopes that were deficient in fusion by virtue of the addition of targeting domains.

Conclusions

These data have implications for the development of retroviral vector targeting strategies from the perspectives of the specificity of target cell interaction and compensating for chimaeric envelope fusion deficiency. Copyright © 2002 John Wiley & Sons, Ltd.

     

PLUNC: a multifunctional surfactant of the airways

PLUNC (palate, lung and nasal epithelium clone) protein is an abundant secretory product of epithelia throughout the mammalian conducting airways. Despite its homology with the innate immune defence molecules BPI (bactericidal/permeability-increasing protein) and LBP (lipopolysaccharide-binding protein), it has been difficult to define the functions of PLUNC. Based on its marked hydrophobicity and expression pattern, we hypothesized that PLUNC is an airway surfactant. We found that purified recombinant human PLUNC exhibited potent surfactant activity by several different measures, and experiments with airway epithelial cell lines and primary cultures indicate that native PLUNC makes a significant contribution to the overall surface tension in airway epithelial secretions. Interestingly, we also found that physiologically relevant concentrations of PLUNC-inhibited Pseudomonas aeruginosa biofilm formation in vitro without acting directly as a bactericide. This finding suggests that PLUNC protein may inhibit biofilm formation by airway pathogens, perhaps through its dispersant properties. Our data, along with reports from other groups on activity against some airway pathogens, expand on an emerging picture of PLUNC as a multifunctional protein, which plays a novel role in airway defences at the air/liquid interface.     

Frequency of alleles conferring resistance to Bt maize in French and US corn belt populations of the European corn borer,Ostrinia nubilalis

Farmers, industry, governments and environmental groups agree that it would be useful to manage transgenic crops producing insecticidal proteins to delay the evolution of resistance in target pests. The main strategy proposed for delaying resistance to Bacillus thuringiensis ( Bt) toxins in transgenic crops is the high-dose/refuge strategy. This strategy is based on the unverified assumption that resistance alleles are initially rare (<10(-3)). We used an F(2) screen on >1,200 isofemale lines of Ostrinia nubilalis Hübner (Lepidoptera: Crambidae) collected in France and the US corn belt during 1999-2001. In none of the isofemale lines did we detect alleles conferring resistance to Bt maize producing the Cry1Ab toxin. A Bayesian analysis of the data indicates that the frequency of resistance alleles in France was <9.20 x 10(-4) with 95% probability, and a detection probability of >80%. In the northern US corn belt, the frequency of resistance to Bt maize was <4.23 x 10(-4) with 95% probability, and a detection probability of >90%. Only 95 lines have been screened from the southern US corn belt, so these data are still inconclusive. These results suggest that resistance is probably rare enough in France and the northern US corn belt for the high-dose plus refuge strategy to delay resistance to Bt maize.     

Mechanistic niche modelling: combining physiological and spatial data to predict species' ranges

Species distribution models (SDMs) use spatial environmental data to make inferences on species' range limits and habitat suitability. Conceptually, these models aim to determine and map components of a species' ecological niche through space and time, and they have become important tools in pure and applied ecology and evolutionary biology. Most approaches are correlative in that they statistically link spatial data to species distribution records. An alternative strategy is to explicitly incorporate the mechanistic links between the functional traits of organisms and their environments into SDMs. Here, we review how the principles of biophysical ecology can be used to link spatial data to the physiological responses and constraints of organisms. This provides a mechanistic view of the fundamental niche which can then be mapped to the landscape to infer range constraints. We show how physiologically based SDMs can be developed for different organisms in different environmental contexts. Mechanistic SDMs have different strengths and weaknesses to correlative approaches, and there are many exciting and unexplored prospects for integrating the two approaches. As physiological knowledge becomes better integrated into SDMs, we will make more robust predictions of range shifts in novel or non-equilibrium contexts such as invasions, translocations, climate change and evolutionary shifts.