Chromatin structural changes in sperm after scrotal insulation of Holstein bulls

The reported effects on semen quality ascribed to testicular heat stress generally relate to traits impacting sperm transport and fertilizing ability but not to the genetic material contained by the sperm. To characterize the effects of testicular heat stress on sperm chromatin, susceptibility of DNA in sperm nuclear chromatin to in situ acid denaturation was measured by flow cytometry after staining with acridine orange using the sperm chromatin structure assay (SCSA). Semen was collected from Holstein bulls at 3-day intervals, before and after 48-hour scrotal insulation, until the morphologically abnormal sperm content in raw semen exceeded 50%. After cryopreservation in egg yolk-citrate extender, semen was thawed and sampled during incubation in vitro at 38.5 degrees C. Overall, SCSA results showed that chromatin susceptibility to denaturation was increased for sperm collected post- vs. preinsulation and was more pronounced for sperm presumably in the testes during insulation than for those sperm presumably in the epididymides. Increased susceptibility was detected as early as the first collection postinsulation; however, chromatin of sperm presumably in the proximal epididymis during insulation did not appear to have been detrimentally affected. Chromatin susceptibility to denaturation increased with increased incubation time in vitro, but the rate of change in susceptibility during incubation did not differ among pre- vs. postinsulation specimens. We conclude that elevated scrotal temperatures adversely affect both epididymal and testicular sperm by reducing sperm chromatin stability. The effects of heat stress on the chromatin of epididymal sperm were more subtle than those exhibited by testicular sperm but detectable within close proximity to the heat stress event.     

"Aggrecanase" activity is implicated in tumour necrosis factor alpha mediated cartilage aggrecan breakdown but is not detected by an in vitro assay

An accurate analysis of the morphological changes which take place during pathological processes of the posterior pole is important for a correct diagnosis and therapeutic approach. The purpose of the study was to determine the intraobserver and interobserver reproducibility of the Image-net system 100 (Topcon, Japan) to take measurements on the retina. The program 'Linear/Areal Measurement functions' of Image-net system 100 which is an image digitalization technique, was tested. Twelve patients were consecutively selected from the patients of the Retina Center of the Department of Ophthalmology, University of Genoa. Three images of each eye were taken from each subject and only the best image was used in this study. The intraobserver and interobserver reproducibility of both the distance between two pre-set points (linear measurement), and the perimeter and area of preselected retinal zones were calculated. The repeatability (or intraobserver reproducibility) of the linear sizes was measured by the coefficient of variation and ranged from 0.32% to 7.38%, while the interobserver reproducibility ranged from 0.46% to 5.22%. The repeatability and reproducibility of the perimeters ranged from 0.72% to 9.63% and from 0.6% to 5.7%, respectively, while the repeatability and reproducibility of the areas ranged from 0.72% to 9.63% and from 0.6% to 5.7%, respectively. Although the results were quite good, the quality of the image of the fundus and the number of observers influenced the coefficient of variation; furthermore, the anatomy of the areas to be measured and the computer 'mouse' could increase the value of the coefficient of variation.     

Crystal structure of the complex between VEGF and a receptor-blocking peptide

Vascular endothelial growth factor (VEGF) is a specific and potent angiogenic factor and, therefore, a prime therapeutic target for the development of antagonists for the treatment of cancer. As a first step toward this goal, phage display was used to generate peptides that bind to the receptor-binding domain (residues 8-109) of VEGF and compete with receptor [Fairbrother, W. J., Christinger, H. W., Cochran, A. G., Fuh, G., Keenan, C. J., Quan, C., Shriver, S. K., Tom, J. Y. K., Wells, J. A., and Cunningham, B. C. (1999) Biochemistry 38, 17754-17764]. The crystal structure of VEGF in complex with one of these peptides was solved and refined to a resolution of 1.9 A. The 20-mer peptide is unstructured in solution and adopts a largely extended conformation when bound to VEGF. Residues 3-8 form a beta-strand which pairs with strand beta6 of VEGF via six hydrogen bonds. The C-terminal four residues of the peptide point away from the growth factor, consistent with NMR data indicating that these residues are flexible in the complex in solution. In contrast, shortening the N-terminus of the peptide leads to decreased binding affinities. Truncation studies show that the peptide can be reduced to 14 residues with only moderate effect on binding affinity. However, because of the extended conformation and the scarcity of specific side-chain interactions with VEGF, the peptide is not a promising lead for small-molecule development. The interface between the peptide and VEGF contains a subset of the residues recognized by a neutralizing Fab fragment and overlaps partially with the binding site for the Flt-1 receptor. The location of the peptide-binding site and the hydrophilic character of the interactions with VEGF resemble more the binding mode of the Fab fragment than that of the receptor.     

Primary malignant fibrous histiocytoma of spleen

We report a case of primary malignant fibrous histiocytoma of spleen, which also emphasizes the difficulties in diagnosing such a rare condition.     

Giant coronary artery pseudoaneurysm causing pulmonary artery obstruction: a rare complication of coronary bypass surgery--a case report

The authors report the diagnosis and successful management of a 57-year-old man with right ventricular outflow tract obstruction from a large pseudoaneurysm of the left anterior descending coronary artery 5 years after he had undergone redo coronary artery bypass grafting.     

Multiple temporal components of optic flow responses in MST neurons

Activation of the hypothalamus-pituitary-interrenal (HPI) axis is characteristic of stress responses, which may result from a variety of environmental challenges. To investigate whether the stress response, and in particular the HPI axis, in tilapia (Oreochromis mossambicus) is compromised by short-term exposure to PCB 126, fish of both sexes were fed diets containing PCB 126 (50 microg/kg fish . day) for 5 days. In the first approach, which was performed twice, fish were acutely stressed for periods varying between 1 and 30 min at the end of the exposure period; in the second approach fish were sampled at the end of the exposure period either at rest or after 2 h of stress (confinement). After 5 days, the body weights in all experiments were significantly lower in PCB-fed fish than in control fish. There were no changes in basal plasma glucose levels, plasma ion concentrations, or branchial, renal, and intestinal Na,K-ATPase activity following PCB exposure. In the first experimental approach, in which fish experienced acute sampling stress, plasma cortisol levels reached lower levels in PCB-fed fish than in controls. This suggests an impaired ability to acutely activate interrenal steroidogenesis in PCB-treated tilapia. Adrenocorticotropic hormone (ACTH)- and cAMP-stimulated in vitro cortisol release from superfused head kidneys was lower in tissues from tilapia exposed to PCB 126 than in tissues from control animals. This effect persisted after 24 h in vitro, which, together with the high PCB 126 concentrations measured in the head kidneys of PCB-fed fish, may indicate direct toxic effects on the interrenal cells. The second experimental approach demonstrated that basal plasma cortisol and ACTH levels were not influenced by PCB treatment, but that the basal ACTH content of the rostral pars distalis (RPD) of the pituitary gland of PCB-fed fish was lower than that of control fish. After 2 h confinement, plasma cortisol levels and ACTH content of the RPD rose to similar values in both groups, whereas plasma ACTH levels were higher in confined PCB-fed fish than in confined controls. PCB-fed fish showed a lower hyperglycemic response to confinement than control fish. Confinement resulted in similarly elevated renal and intestinal Na,K-ATPase activities in both PCB-fed and control fish; branchial enzyme activities were not affected. Since PCB did not affect Na,K-ATPase activities and plasma ion concentrations, it is concluded that the effects of PCB 126 on the HPI axis in tilapia are not secondary to ionoregulatory dysfunction.     

Effects of oxygen, positive end-expiratory pressure, and carbon dioxide on oxygen delivery in an animal model of the univentricular heart

OBJECTIVE: Respiratory manipulations are a mainstay of therapy for infants with a univentricular heart, but until recently little experimental information has been available to guide their use. We used an animal model of a univentricular heart to characterize the physiologic effects of a number of commonly used ventilatory treatments, including altering inspired oxygen tension, adding positive end-expiratory pressure, and adding supplemental carbon dioxide to the ventilator circuit. RESULTS: Lowering inspired oxygen tension decreased the ratio of pulmonary to systemic flow. This ratio was 1.29 +/- 0.08 at an inspired oxygen tension of 100%, 0.61 +/- 0.09 at an inspired oxygen tension of 21%, and 0.42 +/- 0.09 at an inspired oxygen tension of 15% (p < 0.05 compared with an inspired oxygen tension of 100% and a positive end-expiratory pressure of 0 cm H2O). High-concentration supplemental carbon dioxide (carbon dioxide tension of 80 to 90 mm Hg) added to the ventilator circuit decreased inspired oxygen tension from 1.29 +/- 0.11 to 0.42 +/- 0.12 (p < 0.05 compared with baseline). A mixture of 95% oxygen and 5% carbon dioxide (carbon dioxide tension of 50 to 60 mm Hg) did not decrease the pulmonary/systemic flow ratio significantly. All three types of interventions influenced systemic oxygen delivery, which was a function of the pulmonary/systemic flow ratio. As the pulmonary/systemic flow ratio decreased from initially high levels (greater than 1), oxygen delivery first increased and reached an optimum at a flow ratio slightly less than 1. As the pulmonary/systemic flow ratio decreased further, below 0.7, oxygen delivery decreased. The ability of systemic arterial and venous oxygen saturations to predict the pulmonary/systemic flow ratio was examined. Venous oxygen saturation correlated well with both pulmonary/systemic flow ratio and systemic oxygen delivery, whereas arterial oxygen saturation did not accurately predict either pulmonary/systemic flow ratio or oxygen delivery. CONCLUSION: This model demonstrated the value of estimating the pulmonary/systemic flow ratio before initiating therapy. When the initial ratio was greater than about 0.7, interventions that decreased the ratio increased oxygen delivery and were beneficial. When the initial pulmonary/systemic flow ratio was below 0.7, interventions that decreased the ratio decreased oxygen delivery and were detrimental. We conclude by presenting a framework to guide therapy based on the combination of arterial and venous oxygen saturations and the estimate of the pulmonary/systemic flow ratio that they provide.     

The junction-associated protein, zonula occludens-1, localizes to the nucleus before the maturation and during the remodeling of cell-cell contacts

The junction-associated protein zonula occludens-1 (ZO-1) is a member of a family of membrane-associated guanylate kinase homologues thought to be important in signal transduction at sites of cell-cell contact. We present evidence that under certain conditions of cell growth, ZO-1 can be detected in the nucleus. Two different antibodies against distinct portions of the ZO-1 polypeptide reveal nuclear staining in subconfluent, but not confluent, cell cultures. An exogenously expressed, epitope-tagged ZO-1 can also be detected in the nuclei of transfected cells. Nuclear accumulation can be stimulated at sites of wounding in cultured epithelial cells, and immunoperoxidase detection of ZO-1 in tissue sections of intestinal epithelial cells reveals nuclear labeling only along the outer tip of the villus. These results suggest that the nuclear localization of ZO-1 is inversely related to the extent and/or maturity of cell contact. Since cell-cell contacts are specialized sites for signaling pathways implicated in growth and differentiation, we suggest that the nuclear accumulation of ZO-1 may be relevant for its suggested role in membrane-associated guanylate kinase homologue signal transduction.     

Can clinical response to cyclosporin in chronic severe asthma be predicted by an in vitro T-lymphocyte proliferation assay?

This study tests the hypothesis that the clinical response to cyclosporin therapy of patients with chronic severe asthma is related to the sensitivity of their T-lymphocytes to the antiproliferative effects of cyclosporin in vitro. In a previous study, we observed such a relationship with glucocorticoids and the same lectin-driven proliferation assay was used in the present study. Peripheral blood mononuclear cells were obtained from 33 patients participating in a cross-over trial of oral cyclosporin therapy during both cyclosporin and placebo treatment periods, and cultured in the presence of phytohaemagglutinin and serial dilutions of cyclosporin and dexamethasone. Proliferation was measured by tritiated thymidine uptake. Both cyclosporin and dexamethasone inhibited T-lymphocyte proliferation in a concentration-dependent manner in vitro at concentrations encompassing those achieved in peripheral blood during therapy in vivo. T-lymphocytes from the asthmatic patients showed a range of sensitivity to the antiproliferative effects of cyclosporin, but this could not be correlated with improvements in peak expiratory flow rate (PEFR) or forced expiratory volume in one second (FEV1) during cyclosporin therapy as compared with placebo. In contrast to previous observations with glucocorticoids, this in vitro T-lymphocyte proliferation assay is not predictive of clinical response to cyclosporin therapy in chronic severe asthmatics.