Electron microscopical studies of membrane injuries in blue fox spermatozoa subjected to the process of freezing and thawing

Disintegration of blue fox sperm membranes is studied by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). In unfrozen spermatozoa studied by SEM, the plasmalemma and the acrosome appeared to be intact, except for a few cases of disruption of the former structure at the anterior part of the head. In semen frozen in 0.5-ml plastic straws by use of N2 vapor after dilution with Tris-fructose-citric acid with 8 vol % glycerol and 20 vol % egg yolk and thawed at 70 degrees C for 8 sec, the spermatozoa displayed different degrees of membrane damage. These alterations could be classified into three main categories of which the first included only minor changes in the plasmalemma, but vesiculation and disintegration of the outer part of the acrosomal membrane. In the second category (also the most frequent one) the outer part of the acrosomal membrane was extensively vesiculated, and the plasmalemma was discharged proximal to the equatorial segment. Extensive loss of plasmalemma and complete absence of the outer part of the acrosomal membrane characterized the last category of membrane damage. The functional implications of the three categories of membrane alterations are discussed.     

The "prismane" protein resolved: X-ray structure at 1.7 Å and multiple spectroscopy of two novel 4Fe clusters

The three-dimensional structure of the native "putative prismane" protein from Desulfovibrio vulgaris (Hildenborough) has been solved by X-ray crystallography to a resolution of 1.72?Å. The molecule does not contain a [6Fe-6S] prismane cluster, but rather two 4Fe clusters some 12?Å apart and situated close to the interfaces formed by the three domains of the protein. Cluster 1 is a conventional [4Fe-4S] cubane bound, however, near the N-terminus by an unusual, sequential arrangement of four cysteine residues (Cys 3, 6, 15, 21). Cluster 2 is a novel 4Fe structure with two μ₂-sulfido bridges, two μ₂-oxo bridges, and a partially occupied, unidentified μ₂ bridge X. The protein ligands of cluster 2 are widely scattered through the second half of the sequence and include three cysteine residues (Cys 312, 434, 459), one persulfido-cysteine (Cys 406), two glutamates (Glu 268, 494), and one histidine (His 244). With this unusual mixture of bridging and external type of ligands, cluster 2 is named the "hybrid" cluster, and its asymmetric, open structure suggests that it could be the site of a catalytic activity. X-ray absorption spectroscopy at the Fe K-edge is readily interpretable in terms of the crystallographic model when allowance is made for volume contraction at 10?K; no Fe··Fe distances beyond 3.1?Å could be identified. EPR, Mössbauer and MCD spectroscopy have been used to define the oxidation states and the magnetism of the clusters in relation to the crystallographic structure. Reduced cluster 1 is a [4Fe-4S]¹⁺ cubane with S?=?3/2; it is the first biological example of a "spin-admixed" iron-sulfur cluster. The hybrid cluster 2 has four oxidation states from (formally) all Fe^III to three Fe^II plus one Fe^III. The four iron ions are exchange coupled resulting in the system spins S?=?0, 9/2, 0 (and 4), 1/2, respectively, for the four redox states. Resonance Raman spectroscopy suggests that the bridging ligand X which could not be identified unambiguously in the crystal structure is a solvent-exchangeable oxygen.     

Functional Cooperation of the Interleukin-2 Receptor β Chain and Jak1 in Phosphatidylinositol 3-Kinase Recruitment and Phosphorylation

Phosphatidylinositol 3-kinase (PI 3-K) plays an important role in signaling via a wide range of receptors such as those for antigen, growth factors, and a number of cytokines, including interleukin-2 (IL-2). PI 3-K has been implicated in both IL-2-induced proliferation and prevention of apoptosis. A number of potential mechanisms for the recruitment of PI 3-K to the IL-2 receptor have been proposed. We now have found that tyrosine residues in the IL-2 receptor β chain (IL-2Rβ) are unexpectedly not required for the recruitment of the p85 component of PI 3-K. Instead, we find that Jak1, which associates with membrane-proximal regions of the IL-2Rβ cytoplasmic domain, is essential for efficient IL-2Rβ–p85 interaction, although some IL-2Rβ–p85 association can be seen in the absence of Jak1. We also found that Jak1 interacts with p85 in the absence of IL-2Rβ and that IL-2Rβ and Jak1 cooperate for the efficient recruitment and tyrosine phosphorylation of p85. This is the first report of a PI 3-K–Jak1 interaction, and it implicates Jak1 in an essential IL-2 signaling pathway distinct from the activation of STAT proteins.     

N-terminal pro B-type natriuretic peptide levels predict newly detected atrial fibrillation in a population-based cohort

Background B-type natriuretic peptide (BNP) is secreted from cardiomyocytes and may reflect haemodynamic abnormalities predisposing to atrial fibrillation (AF). We aimed to investigate whether N-terminal pro BNP (NT-proBNP) is associated with newly detected AF in subjects obtained from the general population. Methods From the PREVEND programme (n=8592), we selected all subjects with an available baseline and four-year electrocardiogram and NTproBNP levels at baseline. We excluded subjects with AF at baseline and subjects with a serum creatinine >2.0 mg/dl. Results In total, 6494 subjects were eligible for the prospective analysis (aged 49±12 years, 49.7% men). At four years, AF was detected in 41 (0.6%) subjects. Median NT-proBNP levels at baseline in subjects with newly detected AF after four years was 62.2 (22.6 to 208.5) pg/ml as compared with 35.7 (15.9 to 68.7) pg/ml in those with sinus rhythm (p=0.001). Each 1 standard deviation increment in natural log transformed NT-proBNP was associated with a 54% (5% to 126%, p=0.028) increase in risk for AF after adjustment for other risk factors predisposing to AF. NT-proBNP levels above the sex-specific 80th percentile (97 pg/ml in women and 60 pg/ml in men) were associated with a multivariate odds ratio of 2.65 (1.22 to 5.76, p=0.01) for the occurrence of AF. Conclusion Plasma levels of NT-proBNP predict newly detected AF in subjects obtained from the general population independent of cardiovascular risk factors predisposing to AF. (Neth Heart J 2008;16:73-8.)