Resilin matrix distribution, variability and function in Drosophila.

BACKGROUND: Elasticity prevents fatigue of tissues that are extensively and repeatedly deformed. Resilin is a resilient and elastic extracellular protein matrix in joints and hinges of insects. For its mechanical properties, Resilin is extensively analysed and applied in biomaterial and biomedical sciences. However, there is only indirect evidence for Resilin distribution and function in an insect. Commonly, the presence of dityrosines that covalently link Resilin protein monomers (Pro-Resilin), which are responsible for its mechanical properties and fluoresce upon UV excitation, has been considered to reflect Resilin incidence. RESULTS: Using a GFP-tagged Resilin version, we directly identify Resilin in pliable regions of the Drosophila body, some of which were not described before. Interestingly, the amounts of dityrosines are not proportional to the amounts of Resilin in different areas of the fly body, arguing that the mechanical properties of Resilin matrices vary according to their need. For a functional analysis of Resilin matrices, applying the RNA interference and Crispr/Cas9 techniques, we generated flies with reduced or eliminated Resilin function, respectively. We find that these flies are flightless but capable of locomotion and viable suggesting that other proteins may partially compensate for Resilin function. Indeed, localizations of the potentially elastic protein Cpr56F and Resilin occasionally coincide. CONCLUSIONS: Thus, Resilin-matrices are composite in the way that varying amounts of different elastic proteins and dityrosinylation define material properties. Understanding the biology of Resilin will have an impact on Resilin-based biomaterial and biomedical sciences.

Effects of isoflurane-induced anaesthesia on cognitive performance in a mouse model of Alzheimer's disease: A randomised trial in transgenic APP23 mice.

BACKGROUND: Results from in-vitro experiments suggest that inhalational anaesthetics may have a detrimental effect on the course and incidence of Alzheimer's disease. However, case-control studies in humans show no negative impact of anaesthetics on the course of Alzheimer's disease. OBJECTIVE: To test the hypothesis that 2 h of general anaesthesia with 1 MAC isoflurane changes learning abilities of young and old transgenic Alzheimer's mice (APP23 mice). DESIGN: Randomised controlled double-blinded study in mice. SETTING: Animal laboratory and operating theatre in the Klinik für Anästhesiologie, Technische Universität München, Germany ANIMALS: Ninety-six male mice divided in four groups: young (4 months) APP23 mice and corresponding wild-type mice; old (14 to 16 months) APP23 and corresponding wild-type mice. INTERVENTION: Mice were either anaesthetised for 2 h with 1 MAC isoflurane or sham-anaesthetised ('isoflurane' or 'control'). MAIN OUTCOME MEASURES: Learning and locomotor activity during the following 8 days using the modified Hole Board Test for mice. Results are median (interquartile range) and median difference (95% confidence interval). RESULTS: Young mice, [1.0 (1.3)] as assessed by the number of omission errors, learned better than old [1.8 (1.8); age: P = 0.004, median difference 0.5 (0.2 to 1.0)]. Anaesthetised animals [0.8 (1.5)] learned better than controls [1.6 (1.7); anaesthesia: P = 0.010, median difference 0.5 (0.1 to 0.9)]. This was accompanied by higher locomotor activity in young compared to old mice as assessed by number of line crossings per minute [10 (5) min(-1) vs. 7 (3) min(-1), P < 0.001, median difference 3 (2 to 4) min(-1)]. Anaesthesia and genotype Alzheimer's disease had no impact on locomotor activity. CONCLUSION: Isoflurane may have protective, rather than detrimental, effects on cognition in Alzheimer's disease.

Cognitive deficits after systemic induction of inducible nitric oxide synthase: a randomised trial in rats.

BACKGROUND: Nitric oxide acts as an important neurotransmitter as well as a sepsis mediator. During sepsis, high levels of nitric oxide, produced by the inducible form of the nitric oxide synthase (iNOS), may lead to disturbances concerning these conflicting roles and cause septic encephalopathy. To evaluate this theory, we aimed at first, to demonstrate cognitive dysfunction in a rat model based on systemic iNOS induction; second, to elucidate molecular mechanisms; and third, to prevent cognitive deficits in our sepsis model. METHODS: We used a rat systemic inflammation model that is based on the induction of iNOS by heat-killed Corynebacterium parvum in different doses (30 or 60 mg kg). NO2/NO3 plasma levels were measured to prove iNOS induction. Cognitive performance was investigated. In brain tissue, NOS protein and NOS activity were determined. To prevent cognitive deficits, two groups of rats received L-N-(1-Iminoethyl)-lysine (L-NIL), a specific iNOS inhibitor in the drinking water. RESULTS: The rats[Combining Acute Accent] cognitive performance, that is, short-term memory as well as long-term memory was impaired in C. parvum rats with a peak at the third day after injection in the 60 mg kg group. At the same day, neuronal NOS (nNOS)-protein content in the prefrontal cortex was reduced in C. parvum rats. nNOS activity was also reduced in C. parvum rats. The cognitive deficit in short-term memory could be prevented by L-NIL. CONCLUSION: We demonstrate early, reversible cognitive deficits in a rat model of systemic inflammation with increased systemic iNOS activity. As systemic inhibition of iNOS activity prevented rats from the deficit in short-term memory, an involvement of systemic iNOS induction in this deficit is likely. Whether the reduced nNOS-protein expression and nNOS activity are connected to systemic iNOS induction, however, remains unclear.

Transgenic Alzheimer mice have a larger minimum alveolar anesthetic concentration of isoflurane than their nontransgenic littermates.

BACKGROUND: More than 12% of all people older than 65 yr have Alzheimer's disease. Because nothing is known about changes in demand of volatile anesthetics in this disease, we determined minimum alveolar anesthetic concentration (MAC) values of isoflurane in young and aged transgenic mice at risk of developing Alzheimer's disease (heterozygote APP23 mice with the "Swedish double mutation"). To differentiate between unspecific effects of the transgenic model and specific Alzheimer effects, we additionally evaluated MAC values in mice with the same genetic construct but without the Alzheimer's disease-causing Swedish double mutation (heterozygote APP51/16 mice). METHODS: MAC was determined in 60 mice (10 per group): heterozygote APP23 mice and their wild type littermates at the age of 4 and 18 mo, respectively, and heterozygote APP51/16 mice and their wild type littermates at the age of 18 mo. Anesthesia was induced with isoflurane in oxygen/air. The concentration of inhaled isoflurane varied between 1.0 and 2.0 Vol%, and the motor reaction to toeclamping was recorded. Means of the MAC values were compared with an unpaired t-test. RESULTS: The MAC of 18-mo-old heterozygote APP23 mice was 1.67 +/- 0.09, i.e., 9% larger than the MAC of their wild type littermates (1.53 +/- 0.14; P = 0.020). Heterozygote APP51/16 mice had a lower MAC than their wild type littermates (1.32 +/- 0.14 vs 1.48 +/- 0.13; P = 0.037). All wild type groups and young heterozygote APP23 mice had comparable MAC values. CONCLUSIONS: The increased MAC value in aged heterozygote APP23 mice seems to be attributable to changes related to Alzheimer's disease.

The impact of cardiopulmonary bypass on systemic interleukin-6 release, cerebral nuclear factor-kappa B expression, and neurocognitive outcome in rats.

BACKGROUND: Neurocognitive deficits after cardiac surgery with cardiopulmonary bypass (CPB) continue to affect patients' quality of life, and an inflammatory reaction may be one of the contributors. We designed this experiment to study perioperative systemic interleukin-6 (IL-6) concentrations, cerebral expression of nuclear factor-kappa B (NF-kappaB), and neurocognitive outcome after CPB in young rats. The impact of oxygenator size on these outcomes was also assessed. METHODS: Rats were randomly assigned to 1 of 4 groups: control (n = 7, nonanesthetized), sham-operated rats (n = 10, anesthetized, cannulated, and not connected to CPB), and 2 CPB groups, anesthetized, cannulated, and subjected to 90 min of CPB, using either a small-volume rat oxygenator (CPB/rat oxygenator, n = 10) or a neonate oxygenator (CPB/neonate oxygenator, n = 10). Systemic IL-6 was determined before, at the end of, and 2 h after CPB or at equivalent times. Hippocampal NF-kappaB expression was assessed on postoperative day 21 using immunohistochemistry. Neurocognitive performance was assessed with the modified hole-board test at baseline and for 21 postoperative days. RESULTS: Both CPB groups had increased systemic IL-6 levels compared with sham, with the neonate oxygenator causing a substantially larger increase at 2 h after CPB compared with the rat oxygenator group (CPB/rat oxygenator: 220 pg/mL [16-415]; CPB/neonate oxygenator: 1400 pg/mL [592-5812]) (P < 0.05). Hippocampal NF-kappaB was increased in experimental groups compared with controls (10 +/- 4). CPB resulted in more NF-kappaB-positive neurons (271 +/- 57 CPB/neonate oxygenator and 269 +/- 72 CPB/rat oxygenator) compared with sham operation (173 +/- 24). Neurocognitive and behavioral performances were unaltered and comparable among all groups. CONCLUSIONS: Pronounced systemic inflammatory responses to experimental CPB associated with increased hippocampal expression of NF-kappaB were not accompanied by neurocognitive impairment. This suggests that other factors beyond CPB and inflammatory responses might contribute to adverse neurocognitive outcomes after cardiac surgery.

The long-term effect of four hours of hyperventilation on neurocognitive performance and lesion size after controlled cortical impact in rats.

BACKGROUND: We investigated the effects of 4 h of posttraumatic hyperventilation on neurocognitive performance, motor function, and coordination as well as lesion volume in rats subjected to focal traumatic brain injury. METHODS: After a 14-day training period with various neurocognitive tests including hole-board, beam walk, and beam balance, 21 male Sprague-Dawley rats (369 +/- 15 g) were anesthetized with halothane, tracheally intubated, their lungs mechanically ventilated, and subjected to controlled cortical impact (1.75 mm depth, diameter 5 mm, 4 m/s). They were then randomized to either normoventilation (n = 10; PaCO(2) = 38-42 mm Hg) or hyperventilation (n = 11; PaCO(2) = 28-32 mm Hg) and ventilated for 4 h, respectively. Posttraumatic performance in the behavioral and motor tests was evaluated for 20 days. Rats were then decapitated under deep anesthesia, and their brains frozen and sliced to evaluate lesion volume. RESULTS: Hyperventilated animals performed significantly worse in explicit memory tests compared with normoventilated rats over time. Both groups showed deficits in advanced motor function and coordination (evaluated by beam walk and beam balance) initially, with a significantly worse performance of hyperventilated compared with normoventilated animals. However, there was no difference between groups by the end of the study. On Day 20 after injury, lesion volume was significantly larger with hyperventilated (69.7 +/- 13.0 mm(3)) compared with normoventilated animals (48.3 +/- 15.6 mm(3)). CONCLUSIONS: Although hyperventilation enhanced histologic damage, there was no long-term adverse neurocognitive effect from 4 h of posttraumatic hyperventilation (PaCO(2) = 28-32 mm Hg) in rats.

The motility-based swim-up technique separates bull sperm based on differences in metabolic rates and tail length.

Swim-up is a sperm purification method that is being used daily in andrology labs around the world as a simple step for in vitro sperm selection. This method accumulates the most motile sperm in the upper fraction and leaves sperm with low or no motility in the lower fraction. However, the underlying reasons are not fully understood. In this article, we compare metabolic rate, motility and sperm tail length of bovine sperm cells of the upper and lower fraction. The metabolic assay platform reveals oxygen consumption rates and extracellular acidification rates simultaneously and thereby delivers the metabolic rates in real time. Our study confirms that the upper fraction of bull sperm has not only improved motility compared to the cells in the lower fraction but also shows higher metabolic rates and longer flagella. This pattern was consistent across media of two different levels of viscosity. We conclude that the motility-based separation of the swim-up technique is also reflected in underlying metabolic differences. Metabolic assays could serve as additional or alternative, label-free method to evaluate sperm quality.

Chronic Escherichia coli infection induces muscle wasting without changing acetylcholine receptor numbers.

OBJECTIVE: Muscle weakness in septic patients is a serious problem as it complicates and lengthens hospital stays, prolongs rehabilitation and increases costs. We examined the effects of a chronic infection with Escherichia coli on muscle function, muscle mass, and the expression of nicotinic acetylcholine receptors (AChRs). DESIGN AND SETTING: Prospective, randomized animal study in an animal laboratory, university hospital. SUBJECTS: Sprague-Dawley rats. INTERVENTIONS: A catheter was implanted into the external jugular vein of anesthetized rats, and a dose of 3.2x10(8) CFU of E. coli bacteria was injected; the catheter was then sealed and tunneled subcutaneously. MEASUREMENTS AND RESULTS: Animals injected with E. coli bacteria showed a significant decrease in body and muscle mass over the 14-day experimental period. Neuromuscular function was tested by mechanomyography on days 3, 7, and 14 following injection. Tetanic tension decreased over the time course of sepsis, without effecting tetanic fade. Serum levels of acute-phase protein, alpha1-acid glycoprotein, were increased by day 3, and remained significantly higher until day 14. AChRs were quantitated using 125I-labeled bungarotoxin and revealed no differences between groups. CONCLUSIONS: Central venous injection of E. coli bacteria induces systemic inflammation evidenced as loss of body weight, muscle mass and increased alpha1-acid glycoprotein levels. The inflammation-induced muscle weakness is due to muscle atrophy and not to upregulated AChRs. This model may prove useful for studying maneuvers to prevent muscle wasting with inflammation.

Anesthetics promoting in vitro AßPP metabolism and amyloid-ß toxicity.

At present, more than 35 million people worldwide have Alzheimer's disease (AD). With increasing incidence and a growing number of aged patients undergoing surgery, the relevance of a possible interaction between anesthetics and AD is growing as well. Below, we review in vitro studies investigating the effects of anesthetics on the metabolism of amyloid precursor protein and its metabolite amyloid-ß.