A translational approach for limb vascular delivery of the micro-dystrophin gene without high volume or high pressure for treatment of Duchenne muscular dystrophy

Background  

Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder with monogenic mutations setting the stage for successful gene therapy treatment. We have completed a study that directly deals with the following key issues that can be directly adapted to a gene therapy clinical trial using rAAV considering the following criteria: 1) A regional vascular delivery approach that will protect the patient from widespread dissemination of virus; 2) an approach to potentially facilitate safe passage of the virus for efficient skeletal muscle transduction; 3) the use of viral doses to accommodate current limitations imposed by vector production methods; 4) and at the same time, achieve a clinically meaningful outcome by transducing multiple muscles in the lower limb to prolong ambulation.     

Human natural killer cell lysis of Salmonella typhi intracellularly-infected U937 cells

Peripheral blood mononuclear cell (PBMC) cytotoxicity against S. typhi (wild type or mutant strain TYT1231)-infected U937 cells was significantly higher than its lytic effect against noninfected cells (control) at the various effector-to-target cell ratio used (30:1, 50:1 and 70:1). Natural killer cell activity [expressed as % specific lysis (mean +/- SEM); 30:1 (25.4 +/- 3.6, 25.1 +/- 4.2 and 16.3 +/- 3.3); 50:1 (27.8 +/- 3.7, 26.7 +/- 4.5 and 20.9 +/- 2.9) and 70:1 ratio (33.2 +/- 5.9, 29.4 +/- 4.2 and 22.8 +/- 2.8), respectively] appeared to be dependent on such ratios and independent of the S strain studied. Most (80%) of individual samples tested showed at least a 20% specific lysis increase over their own control; essentially no changes or smaller increases in NKC activity were observed in all other samples. Similar results were obtained when using highly purified NKC (HPNKC) preparations as effector cells [NKC activity (mean +/- SEM); 5:1 (46.2 +/- 4.7, 43.2 +/- 5.0 and 25.2 +/- 2.3) and 10:1 effector-to-target cell ratio (49.3 +/- 4.9, 44.7 +/- 5.2 and 27.2 +/- 2.6, respectively)]. All individual samples tested showed at least a 20% specific lysis increase over their own control. These results show that S. typhi-infected U937 cells are a significantly better target for NKCs than control cells and indicate that intracellular bacteria survival capacity is not a critical factor for infected cells becoming a NKC target.     

Cardiovascular effects of cerebroventricular ouabain perfusion in the adult dog.

Cerebroventricular perfusion with artificial cerebrospinal fluid containing 10(-5) M ouabain was performed in adult dogs in order to describe the time course of the cardiovascular effect of intraventricular ouabain and to evaluate treatments to eliminate the cardiovascular effect. The central effect of ouabain caused a 56% increase in blood pressure above control values and a 35% increase in heart rate with various cardiac arrhythmias. Both alpha- and beta-adrenergic blocking drugs given intravenously.altered the pressure and rate effects ou ouabain, whereas vagotomy attenuated the effect.     

HIV infection among patients in U.S. acute care hospitals. Strategies for the counseling and testing of the hospital patients. The Hospital HIV Surveillance Group.

BACKGROUND. Routine, voluntary testing of hospital patients for the human immunodeficiency virus (HIV) has been proposed in order to identify those with early HIV infection in a setting where there is ready access to counseling, appropriate clinical referral, evaluation, and therapy. We studied the pattern of HIV infection among patients in 20 U.S. hospitals, in order to evaluate possible national strategies for the routine, voluntary HIV counseling and testing of hospital patients. METHODS. Blood specimens remaining after clinical use from a systematically selected sample of patients at 20 hospitals in 15 U.S. cities were tested anonymously for antibody to HIV type 1 (HIV-1). Multivariate regression was used to determine which variables best predicted HIV seroprevalence in individual hospitals. Using these data, we estimated the number of HIV-positive patients in all U.S. hospitals and considered the efficiency of routine counseling and testing in different subgroups of patients and hospitals. RESULTS. From September 1989 through October 1991, 9286 of 195,829 specimens (4.7 percent) were positive for HIV-1 in the 20 hospitals. The seroprevalence of HIV at these institutions ranged from 0.2 percent to 14.2 percent. Among HIV-positive patients, 32 percent had symptomatic HIV infection or the acquired immunodeficiency syndrome (AIDS) at the time of admission or evaluation. In the 20 hospitals, HIV seroprevalence was 10.4 times (95 percent confidence interval, 8.8 to 12.0) the AIDS-diagnosis rate (the annual number of patients with new diagnoses of AIDS per 1000 discharges in 1990). In a multivariate model that included 13 hospital-specific variables, only the AIDS-diagnosis rate was associated with the hospital-specific HIV-seroprevalence rate (P less than 0.001). Using these data and the AIDS-diagnosis rates for all U.S. acute care hospitals, we estimated that 225,000 HIV-positive persons were hospitalized (95 percent confidence interval, 190,000 to 260,000) in all 5558 such hospitals in 1990, including 163,000 persons presenting with conditions other than HIV or AIDS (95 percent confidence interval, 130,000 to 196,000). In 1990, in 593 U.S. hospitals with AIDS-diagnosis rates of 1.0 or more per 1000 discharges, HIV testing of patients 15 to 54 years old (3 million patients, or 12.0 percent of all patients in U.S. acute care hospitals) would have identified an estimated 68 percent of all HIV-positive patients (110,000 patients) who were admitted with conditions other than symptomatic HIV infection or AIDS. CONCLUSIONS. We estimate that about 225,000 HIV-positive persons were hospitalized in 1990, of whom only one third were admitted for symptomatic HIV infection or AIDS. Routine, voluntary HIV testing of patients 15 to 54 years old in hospitals with 1 or more patients with newly diagnosed AIDS per 1000 discharges per year could potentially have identified as many as 110,000 patients with HIV infection that was previously unrecognized.     

Prevalence of molecular types and mecA gene carriage of coagulase-negative Staphylococci in a neonatal intensive care unit: relation to nosocomial septicemia

Molecular typing of isolates revealed that neonatal coagulase-negative staphylococcal (CONS) septicemia is most frequently caused by predominant, antibiotic-resistant CONS types, which are widely distributed among both neonates and staff of the neonatal unit, suggesting cross-contamination. Therefore, infection control measures may be valuable in the prevention of this common nosocomial septicemia.     

Differences in assembly or stability of complex I and other mitochondrial OXPHOS complexes in inherited complex I deficiency

NADH-ubiquinone oxidoreductase (complex I) deficiency is amongst the most encountered defects of the mitochondrial oxidative phosphorylation (OXPHOS) system and is associated with a wide variety of clinical signs and symptoms. Mutations in complex I nuclear structural genes are the most common cause of isolated complex I enzyme deficiencies. The cell biological consequences of such mutations are poorly understood. In this paper we have used blue native electrophoresis in order to study how different nuclear mutations affect the integrity of mitochondrial OXPHOS complexes in fibroblasts from 15 complex I-deficient patients. Our results show an important decrease in the levels of intact complex I in patients harboring mutations in nuclear-encoded complex I subunits, indicating that complex I assembly and/or stability is compromised. Different patterns of low molecular weight subcomplexes are present in these patients, suggesting that the formation of the peripheral arm is affected at an early assembly stage. Mutations in complex I genes can also affect the stability of other mitochondrial complexes, with a specific decrease of fully-assembled complex III in patients with mutations in NDUFS2 and NDUFS4. We have extended this analysis to patients with an isolated complex I deficiency in which no mutations in structural subunits have been found. In this group, we can discriminate between complex I assembly and catalytic defects attending to the fact whether there is a correlation between assembly/activity levels or not. This will help us to point more selectively to candidate genes for pathogenic mutations that could lead to an isolated complex I defect.     

Application of simple anthropometry in the assessment of health risk: implications for the Canadian Physical Activity, Fitness and Lifestyle Appraisal.

Incremental improvements in our knowledge of the associations between human body composition and disease have been facilitated by advances in research technology. Magnetic resonance imaging and computerized tomography are among the technological advances that have helped unravel the mechanisms that link body composition and disease. However, because the use of these methods in large-scale studies and field settings is impractical, the potential relationships between body composition and health risk rely on the use of anthropometric tools. Indeed, the application of simple anthropometry to identify relationships between body composition and health risk in clinical practice is no less valuable than the use of advanced technologies to gain insight into the mechanistic links between body composition and disease in the laboratory. Accordingly, the purpose of this review is to summarize current knowledge regarding the ability of anthropometry to predict health risk and to act as surrogate measures of total and abdominal fat distribution. Because the ultimate objective is to make recommendations for revision to the Healthy Body Composition section of the Canadian Physical Activity, Fitness and Lifestyle Appraisal (CPAFLA) manual, we focus on those anthropometric methods specific to CPAFLA. Consistent with this objective, when necessary we present original data to reinforce important concepts not suitably addressed in the literature.     

Kinetics of NDV-specific antibodies in hens: I. Methods and reproducibility of HAH tests

1. It is recommended to use for the hemagglutination inhibition test (HI test) 0,85% saline solution (pH 6,8), for dilution of sera (log2 dilution), NDV (4 HA units) and for suspension of chicken red blood cells (1%). 2. No evidence was obtained that the use of various buffers for the HI test influenced the titer. 3. Various strains of NDV (G 35, Herts, Italia, Montana, R 1111) have similar HI titers. However for standardization purposes it is recommended to use known and characterized strains of NDV such as Montana or Italia. 4. Similar results were obtained using the macro method according to Salk and the micro method according to Takatsy. 5. The use of the recommended procedure results in high reproducibility of the HI titers obtained.     

Rapid detection of cucumber vein yellowing virus by tissue-print hybridisation with digoxigenin-labelled cDNA probes

Hybridisation of tissue prints with nonradioactive cDNA probes was developed to detect cucumber vein yellowing virus (CVYV) in cucurbit plants. Results showed irregular distribution of the virus within cucumber, zucchini or melon plants without defined tropism for a specific tissue. Therefore, reliable diagnosis of CVYV requires analysis of tissue prints from at least five different plant sites. This detection procedure allows rapid analysis of large numbers of plants and it can be useful for epidemiological studies of CVYV and to control virus spread via eradication of early foci.