Probing the role of the proximal heme ligand in cytochrome P450cam by recombinant incorporation of selenocysteine

The unique monooxygenase activity of cytochrome P450cam has been attributed to coordination of a cysteine thiolate to the heme cofactor. To investigate this interaction, we replaced cysteine with the more electron-donating selenocysteine. Good yields of the selenoenzyme were obtained by bacterial expression of an engineered gene containing the requisite UGA codon for selenocysteine and a simplified yet functional selenocysteine insertion sequence (SECIS). The sulfur-to-selenium substitution subtly modulates the structural, electronic, and catalytic properties of the enzyme. Catalytic activity decreases only 2-fold, whereas substrate oxidation becomes partially uncoupled from electron transfer, implying a more complex role for the axial ligand than generally assumed.     

Functional analysis of an Orc6 mutant in Drosophila

The origin recognition complex (ORC) is a 6-subunit complex required for the initiation of DNA replication in eukaryotic organisms. ORC is also involved in other cell functions. The smallest Drosophila ORC subunit, Orc6, is important for both DNA replication and cytokinesis. To study the role of Orc6 in vivo, the orc6 gene was deleted by imprecise excision of P element. Lethal alleles of orc6 are defective in DNA replication and also show abnormal chromosome condensation and segregation. The analysis of cells containing the orc6 deletion revealed that they arrest in both the G₁ and mitotic stages of the cell cycle. Orc6 deletion can be rescued to viability by a full-length Orc6 transgene. The expression of mutant transgenes of Orc6 with deleted or mutated C-terminal domain results in a release of mutant cells from G₁ arrest and restoration of DNA replication, indicating that the DNA replication function of Orc6 is associated with its N-terminal domain. However, these mutant cells accumulate at mitosis, suggesting that the C-terminal domain of Orc6 is important for the passage through the M phase. In a cross-species complementation experiment, the expression of human Orc6 in Drosophila Orc6 mutant cells rescued DNA replication, suggesting that this function of the protein is conserved among metazoans.     

Correlations between postural stability and strength of lower body extremities of women population living in long-term care facilities

The present study was aimed at analyzing correlation between strength of lower body extremities and postural stability in function of age. A pool of 180 women divided into 6 age groups (65-69, 70-74, 75-79, 80-84, 85-89 and 90-94 years) was examined. They all were informed-consent participants. The results suggest that age is negatively correlated with lower body strength and positively correlated with decrease of postural balance. There is also an exponential correlation between the strength of lower body part and postural balance. The conclusion was derived that postural stability is at least partially controlled by the strength of lower body. The age of 75 in women population seems to be a threshold above which the neurodegenerative and muscle degeneration processes are responsible for significant increase of risk of fall.     

Dual myocardial scintigraphy mismatch in an infant with Bland-White-Garland syndrome

Bland-White-Garland syndrome (BWG) is a rare disorder that includes abnormalities of the coronary arteries that cause severe myocardial ischemia or infarction in infancy. In this case report, we describe an infant with BWG evaluated by dual single photon emission computed tomography using thallium-201 and I-123 beta-methyl-p-iodophenyl-pentadecanoic acid before and after surgery.     

Consecuencias de las contracciones excéntricas del diafragma sobre su función

Introduction and Objectives

Eccentric contractions are those that occur after a muscle has been stretched, and they can predispose the muscle to damage. Most previous studies have been performed on limb muscles, and the potential consequences of eccentric contractions on the respiratory muscles are therefore unknown. The aim of this study was to evaluate the effects of repeated eccentric contractions on diaphragmatic function.

Methods

In 6 dogs, the diaphragm was stretched by applying pressure on the abdominal wall, and consecutive series of eccentric contractions were induced by bilateral supramaximal stimulation. The effect of these contractions on the diaphragm was then evaluated by applying bilateral twitch and tetanic stimulation of the phrenic nerves and measuring the changes in abdominal pressure and the shortening of the right and left hemidiaphragms (by sonomicrometry). Structural study of the muscle was also performed in 4 animals.

Results

Eccentric contractions were successfully achieved in all cases. Stimulation-induced diaphragmatic pressures became lower immediately after these contractions: twitch pressure fell by 53% and tetanic pressure by 67% after the first 10 eccentric contractions (P<.001 in both cases). Tetanic stimulation also demonstrated an early deterioration in contractility, which fell by 29% in the right hemidiaphragm (P<.05) and by 14% in the left hemidiaphragm (P<.001). Functional impairment was persistent, lasting at least 12 hours, and was associated with sarcomeric and sarcolemmal damage.

Conclusions

This experimental model, which enabled the effects of eccentric contractions to be studied in the diaphragm, revealed a deterioration of muscle function that persisted for hours and that appeared to be partly due to structural damage. In the clinical setting, physiologic or therapeutic maneuvers that increase the resting length of the diaphragm should be used with caution.     

Use of modified McMaster method for the diagnosis of intestinal helminth infections and estimating parasitic egg load in human faecal samples in non-endemic areas

Summary  A modified McMaster method has been used for the diagnosis and estimating helminth egg load in human faecal samples obtained from random consecutive patients in the areas non-endemic for helminth infections (Slovak Republic, North West Russia). Both positive and negative findings were in a 100 % concordance to those obtained with a reference method accepted in clinical diagnostic laboratories (microscopy of the native stool smear). The McMaster method was efficient in detecting nematode eggs in patients’ stool samples with egg load varying from very low (15–60 epg for T. trichiura) to moderate (1650–4500 epg for A. lumbricoides). Therefore, this method may be successfully (and with a better technical feasibility) used for the diagnosis of intestinal helminth infections in non-endemic areas, with further quantitative analysis of the sample when required.