A novel, ataxic mouse model of ataxia telangiectasia caused by a clinically relevant nonsense mutation.

Ataxia Telangiectasia (A-T) and Ataxia with Ocular Apraxia Type 1 (AOA1) are devastating neurological disorders caused by null mutations in the genome stability genes, A-T mutated (ATM) and Aprataxin (APTX), respectively. Our mechanistic understanding and therapeutic repertoire for treating these disorders are severely lacking, in large part due to the failure of prior animal models with similar null mutations to recapitulate the characteristic loss of motor coordination (i.e., ataxia) and associated cerebellar defects. By increasing genotoxic stress through the insertion of null mutations in both the Atm (nonsense) and Aptx (knockout) genes in the same animal, we have generated a novel mouse model that for the first time develops a progressively severe ataxic phenotype associated with atrophy of the cerebellar molecular layer. We find biophysical properties of cerebellar Purkinje neurons (PNs) are significantly perturbed (e.g., reduced membrane capacitance, lower action potential [AP] thresholds, etc.), while properties of synaptic inputs remain largely unchanged. These perturbations significantly alter PN neural activity, including a progressive reduction in spontaneous AP firing frequency that correlates with both cerebellar atrophy and ataxia over the animal's first year of life. Double mutant mice also exhibit a high predisposition to developing cancer (thymomas) and immune abnormalities (impaired early thymocyte development and T-cell maturation), symptoms characteristic of A-T. Finally, by inserting a clinically relevant nonsense-type null mutation in Atm, we demonstrate that Small Molecule Read-Through (SMRT) compounds can restore ATM production, indicating their potential as a future A-T therapeutic.

Limitations to vasodilatory capacity and .VO2 max in trained human skeletal muscle.

To further explore the limitations to maximal O(2) consumption (.VO(2 max)) in exercise-trained skeletal muscle, six cyclists performed graded knee-extensor exercise to maximum work rate (WR(max)) in hypoxia (12% O(2)), hyperoxia (100% O(2)), and hyperoxia + femoral arterial infusion of adenosine (ADO) at 80% WR(max). Arterial and venous blood sampling and thermodilution blood flow measurements allowed the determination of muscle O(2) delivery and O(2) consumption. At WR(max), O(2) delivery rose progressively from hypoxia (1.0 +/- 0.04 l/min) to hyperoxia (1.20 +/- 0.09 l/min) and hyperoxia + ADO (1.33 +/- 0.05 l/min). Leg .VO(2 max) varied with O(2) availability (0.81 +/- 0.05 and 0.97 +/- 0.07 l/min in hypoxia and hyperoxia, respectively) but did not improve with ADO-mediated vasodilation (0.80 +/- 0.09 l/min in hyperoxia + ADO). Although a vasodilatory reserve in the maximally working quadriceps muscle group may have been evidenced by increased leg vascular conductance after ADO infusion beyond that observed in hyperoxia (increased blood flow but no change in blood pressure), we recognize the possibility that the ADO infusion may have provoked vasodilation in nonexercising tissue of this limb. Together, these findings imply that maximally exercising skeletal muscle may maintain some vasodilatory capacity, but the lack of improvement in leg .VO(2 max) with significantly increased O(2) delivery (hyperoxia + ADO), with a degree of uncertainty as to the site of this dilation, suggests an ADO-induced mismatch between O(2) consumption and blood flow in the exercising limb.

Smoking in contemporary American cinema.

OBJECTIVE: The true prevalence of smoking among characters portrayed in the movies is unknown. This study examines this prevalence objectively. DESIGN: The top 10 movies on the weekly box office charts were reviewed. Whether or not the top five characters in these movies smoked, was documented. It was determined prior to the start of the study that 300 male characters and 300 female characters were needed to detect any significant difference. A total of 447 movies, composed of 193 movies rated restricted (R) [children < 17 years of age must be accompanied by an adult], 131 movies rated PG13 for parental guidance suggested for children < 13 years of age (PG) and 123 movies rated PG for parental guidance suggested, were examined until the sample size was reached. RESULTS: Smoking prevalence is the same in contemporary American movies and in the general US population (23.3% vs 24.8%, respectively). However, there was more smoking in these movies among men than among women (25.5% vs 20.5%, respectively; p < 0.006), among antagonists than among protagonists (35.7% vs 20.6%, respectively; p < 0.001), lower vs middle vs upper socioeconomic class (SEC) [48.2%, 22.9%, and 10.5%, respectively; p < 0.001], among independent vs studio movies (46.2% vs 18.2%, respectively; p < 0.001); and among R-rated vs PG13-rated vs PG-rated movies (37.3%, 16.2%, and 8.1%, respectively; p < 0.001). In R-rated movies, and in both subcategories of R-rated studio movies and R-rated independent movies, smoking prevalence is higher than in the US population (37.3%, 30.5%, and 50.6% vs 24.8%, respectively; p < 0.001 for all). Additionally, compared to the US population, men, women and lower SEC members smoke more in R-rated movies, R-rated studio movies, and R-rated independent movies. In R-rated movies, antagonists smoke more than protagonists (43.9% vs 35.8%, respectively; p < 0.001), and whites smoke more than nonwhites (38.3% vs 26.4%, respectively; p < 0.001). In R-rated studio movies, antagonists smoke more than protagonists (42.6% vs 26.6%, respectively; p < 0.001), and men smoke more than women (32.0% vs 27.9%, respectively; p = 0.03). In R-rated independent movies, whites smoke more than nonwhites (51.8% vs 40.5%, respectively; p < 0.001). Smoking prevalence is higher in R-rated independent movies than in R-rated studio movies (50.6% vs 30.5%, respectively; p < 0.001). Smoking prevalence is also higher in R-rated independent movies than in R-rated studio movies in subcategories of men (32.0% vs 49.8%, respectively; p < 0.001), women (21.8 vs 51.8%, respectively; p < 0.001), protagonists (26.6% vs 51.6%, respectively; p < 0.001), whites (31.5% vs 51.8%, respectively; p < 0.001), nonwhites (24.7% vs 40.5%, respectively; p < 0.001), and all three SECs. CONCLUSIONS: In contemporary American cinema, the smoking prevalence is higher for men, antagonistic characters, lower SEC, independent movies, and R-rated movies. Smoking prevalence is higher than in the general US population in R-rated movies, and in both its subcategories of R-rated studio movies and R-rated independent movies. There is more smoking in R-rated independent movies than in R-rated studio movies. Smoking in contemporary American cinema is associated with male sex, lower SEC, and antagonistic (ie, bad) characters.

Continuous transforming growth factor beta1 secretion by cell-mediated gene therapy maintains chondrocyte redifferentiation.

One of the most important factors in the production of cartilage is transforming growth factor beta1 (TGF-beta1). To obtain sustained release of TGF-beta1, a cell-mediated gene therapy technique was introduced. We infected chondrocytes with a retroviral vector carrying the TGF-beta1 gene. The single clone derivative showed sustained TGF-beta1 secretion. It also showed constitutive type II collagen expression. Whereas the TGF-beta1 protein itself is unable to induce formation of cartilage in vivo, human chondrocytes engineered to express a retroviral vector encoding TGF-beta1 showed cartilage formation in vivo when cells were injected into nude mice intradermally. These data suggest that cell-mediated gene therapy using TGF-beta1 as a transgene would be a promising treatment for osteoarthritis.

Anaphylactoid response to Optison(R) and its effects on pulmonary function in two dogs.

Two of 26 anesthetized dogs given the cardiac echo-enhancing agent Optison showed anaphylactoid responses (AR) related to the human albumin component of this agent. The episodes of AR were self-limited, and could be reproduced by human albumin injection alone. Gas exchange was maintained by mechanical ventilation and 5 cm H(2)O PEEP, and dispersion of ventilation remained normal during AR despite severe hypotension. We suggest that: (1) pre-screening by measuring blood pressure response to intravenous injection of small doses of Optison, and (2) availability of access to the airway in addition to emergency agents may be prudent preventive measures when Optison is used in animals to enhance echocardiographic imaging.

Regulation of free radical outflow from an isolated muscle bed in exercising humans.

Incremental knee extensor (KE) exercise performed at 25, 70, and 100% of single-leg maximal work rate (WR(MAX)) was combined with ex vivo electron paramagnetic resonance (EPR) spectroscopic detection of alpha-phenyl-tert-butylnitrone (PBN) adducts, lipid hydroperoxides (LH), and associated parameters in five males. Blood samples were taken from the femoral arterial and venous circulation that, when combined with measured changes in femoral venous blood flow, permitted a direct examination of oxidant exchange across a functionally isolated contracting muscle bed. KE exercise progressively increased the net outflow of LH and PBN adducts (100% > 70% > 25% WR(MAX), P < 0.05) consistent with the generation of secondary, lipid-derived oxygen (O(2))-centered alkoxyl and carbon-centered alkyl radicals. Radical outflow appeared to be more intimately associated with predicted decreases in intracellular Po(2) (iPo(2)) as opposed to measured increases in leg O(2) uptake, with greater outflow recorded between 25 and 70% WR(MAX) (P < 0.05 vs. 70-100% WR(MAX)). This bias was confirmed when radical venoarterial concentration differences were expressed relative to changes in the convective components of O(2) extraction and flow (25-70% WR(MAX) P < 0.05 vs. 70-100% WR(MAX), P > 0.05). Exercise also resulted in a net outflow of other potentially related redox-reactive parameters, including hydrogen ions, norepinephrine, myoglobin, lactate dehydrogenase, and uric acid, whereas exchange of lipid/lipoproteins, ascorbic acid, and selected lipid-soluble anti-oxidants was unremarkable. These findings provide direct evidence for an exercise intensity-dependent increase in free radical outflow across an active muscle bed that was associated with an increase in sarcolemmal membrane permeability. In addition to increased mitochondrial electron flux subsequent to an increase in O(2) extraction and flow, exercise-induced free radical generation may also be regulated by changes in iPo(2), hydrogen ion generation, norepinephrine autoxidation, peroxidation of damaged tissue, and xanthine oxidase activation.

Effects of Optison on pulmonary gas exchange and hemodynamics.

Optison is a contrast-enhancing agent used in myocardial contrast echocardiography. It consists of small albumin spherules (approximately 4-microm diameter) containing a fluorocarbon gas, octafluoropropane. It is injected IV and, thus, may cause pulmonary manifestations of microembolism. To determine if any such effects do occur, we injected sequential doses of 1, then 3 and then 5 mL Optison IV into 25 kg anesthetized dogs, and measured pulmonary hemodynamic and gas exchange variables frequently for 30 min after each dose. This was done in both 6 healthy and 6 pulmonary hypertensive animals, the latter produced by acute IV injection of 676-microm diameter polystyrene beads, raising pulmonary artery pressure from normal (15 mmHg) to 33 mmHg. Optison-injected animals were compared with albumin-injected controls. Two animals developed severe hypotension in response to albumin and could not be used. Lung compliance and wet/dry weight ratio were unaffected by Optison and no effects on gas exchange were seen at any dose or time in either group of dogs. In the healthy group, there was slight (1 mmHg per mL Optison, transient and delayed pulmonary hypertension without change in cardiac output, suggesting a vasoconstrictor rather than mechanical basis for these small effects. No such changes occurred in the pulmonary hypertensive group. These results imply that usual human doses of Optison (0.5 mL) will produce no significant hemodynamic or gas exchange effects in either healthy or pulmonary hypertensive dogs.