A longitudinal study of semen quality in pesticide spraying danish farmers

It was hypothesized that occupational exposure to pesticides during a spraying season causes changes in semen quality that might be detected in a longitudinal study. We analyzed the within-person changes in semen quality and reproductive hormones across a spraying season in groups of farmers using and not using pesticides. A total of 248 men collected two semen samples (participation rate: 32%). The median sperm concentration declined significantly from the first to the second sample in both groups, but there was no statistical difference in the decline between the two groups, unadjusted or adjusted. Only minor changes were found in sperm morphology, vitality, motility, sperm chromatin denaturation (SCSA), and reproductive hormones, and the differences in changes between the two groups were nonsignificant, or, in the opposite direction to the expected. There was no relation between the changes in sperm parameters in relation to pesticide exposure variables. In conclusion, use of pesticides by Danish farmers is not a likely cause of short-term effects on semen quality and reproductive hormones.     

Glial cell expression of hepatocyte growth factor in vitreoretinal proliferative disease

The hepatocyte growth factor (HGF) has been crucially implicated in the development of proliferative retinal diseases; however, it is unclear whether retinal glial cells express or respond to HGF. Therefore, we examined the expression of HGF and of the receptor for HGF, c-Met, by immunohistochemical costaining with glial fibrillary acidic protein (GFAP) in epiretinal membranes of patients with proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR), respectively. Furthermore, it was determined whether cells of the human retinal glial cell line, MIO-M1, secrete HGF protein, and whether HGF stimulates proliferation and chemotaxis, and secretion of the vascular endothelial growth factor (VEGF). Neuroretinas of patients with PVR express elevated mRNA level for HGF in comparison to control retinas. In epiretinal membranes of patients with PVR or PDR, immunoreactivity for HGF and for c-Met, respectively, partially colocalized with immunoreactivity for GFAP. Fetal bovine serum and basic fibroblast growth factor, but not heparin-binding epidermal or platelet-derived growth factors, evoked HGF secretion by cultured retinal glial cells. HGF displayed only a marginal effect on cell proliferation while it stimulated chemotaxis. HGF promoted the secretion of VEGF, via activation of the phosphatidylinositol-3 kinase. It is concluded that glial cells in epiretinal membranes express both HGF protein and c-Met receptors. The results suggest an autocrine/paracrine role of HGF in glial cell responses during proliferative vitreoretinal disorders as well as in retinal neovascularization, by stimulating of VEGF release.     

Single-strand breaks, chromosome aberrations, sister-chromatid exchanges, and micronuclei in blood lymphocytes of workers exposed to styrene during the production of reinforced plastics

Chromosome aberrations (CA), micronuclei (MN, cytokinesis-block [CB] method), and sister-chromatid exchanges (SCE) were analysed in blood lymphocytes of 17 workers and 17 control subjects. The mean urinary mandelic acid level (average 9.4 mmol/l) and styrene glycol in blood (average 2.5 mumol/l) implied exposure to about 300 mg/m3 of styrene in the plant. The number of CA was significantly higher in non-smoking workers compared with nonsmoking controls. A significant correlation was observed between duration of exposure and individual CA level of all workers. No significant effects were observed in MN or SCE. Single-strand breaks (SSB) in DNA of isolated lymphocytes were studied in nine of the workers and eight of the controls by the DNA-unwinding technique. The results showed an increase in SSB among the exposed workers. The present findings support earlier reports on the increase of structural CA in blood lymphocytes of workers in the reinforced plastic industry, and also show that SSBs are elevated in such workers.     

A multifunctional bioreactor for three-dimensional cell (co)-culture

Investigation of cell abilities to growth, proliferation and (de)-differentiation in a three-dimensional distribution is an important issue in biotechnological research. Here, we report the development of a new bioreactor for three-dimensional cell culture, which allows for co-cultivation of various cell types with different culture conditions in spatial separation. Preliminary results of neonatal rat cardiomyocyte cultivation are shown. Isolated neonatal rat cardiomyocytes were cultured in spatial separated bioreactor compartments in recirculating medium on a biodegradable fibrin matrix for 2 weeks. Glucose, lactate, and lactate dehydrogenase (LDH), pO2, pCO2, and pH levels were monitored in the recirculated medium, daily. Morphological characterization of matrix and cells was assessed by hematoxylin and eosin staining, and MF-20 co-immunostaining with 4',6-diamidino-2-phenylindole (DAPI). Cell viability was determined by LIVE/DEAD staining before cultivation and on day 3, 7, and 14. The optimized seeding density in the matrix was 2.0 x 10(7) cells retaining cellular proportions over the cell culture period. The bioreactor allows the maintenance of physiologic culture conditions with aerobic cell metabolism (low release of lactate, LDH), a high oxygen tension (pO2-183.7 +/- 18.4 mmHg) and physiological pH values (7.4 +/- 0.02) and a constant level of pCO2 (43.1 +/- 2.9) throughout the experimental course. The cell viability was sufficient after 2 weeks with 82 +/- 6.7% living cells. No significant differences were found between spatial separated bioreactor compartments. Our novel multifunctional bioreactor allows for a three-dimensional culture of cells with spatial separation of the co-cultured cell groups. In preliminary experiments, it provided favorable conditions for the three-dimensional cultivation of cardiomyocytes.     

Quantitative pharmacological characterization of beta-receptors and two types of alpha-receptors mediating sympathomimetic smooth muscle response in the human Fallopian tube at various cyclic stages

The dissociation constants for adrenoceptor-antagonist complexes (KB) were determined in vitro in circular and longitudinal smooth musculature from the ampullary and isthmic regions of the human Fallopian tube. High extracellular potassium concentrations were used to eliminate the spontaneous contractile activity. Neuronal and extraneuronal amine uptake mechanisms were blocked. The parallel shift of the log dose-response curves was secured in Arunlakshana-Schild plots. KB for the beta-receptor, mediating sympathomimetic relaxation, were determined during alpha-receptor blockade: the values for propranolol were the same (approximately 10(-6) M) in all preparations and at all cyclic stages, as determined from plasma estradiol and progesterone levels. KB for the complex between the alpha-receptor (mediating contraction) and phentolamine were determined during beta-receptor blockade. The values were the same in all types of smooth musculature, but varied with cyclic stage: they were around 7 x 10(-8) M when plasma estradiol and progesterone were both minimum, and around 2 x 10(-7) M when these steroid levels were moderate to high, suggesting that the properties of the contractile receptors of the human Fallopian tube are modified during the menstrual cycle.     

Influence of scaffold thickness and scaffold composition on bioartificial graft survival

Biological scaffolds exhibit advantageous properties for tissue engineering of small diameter vessels. The influence of their extracellular matrix (ECM) components during in vivo repopulation is unknown. We implanted different xenogenic vascular matrices in a rat model to determine the influence of scaffold-thickness and ECM composition on in vivo repopulation. Decellularized ovine jugular vein (JV, n=42), carotid artery (CA, n=42) and aorta (AO, n=42) were implanted subcutaneously in the neck of adult male rats. Animals were sacrificed 2, 4 and 8 weeks after implantation. Cell and matrix morphology of explanted scaffolds were characterized by hematoxylin-eosin and pentachrome staining. Monoclonal anti-rat-CD31 was used to identify revascularization. Quantification of cell density was done by DNA-isolation.THICKNESS OF IMPLANTED XENOGENIC SCAFFOLDS VARIED ACCORDING TO THE MATERIAL USED (AO: 3.0-3.8mm; CA: 0.7-0.88mm; JV: 0.35-0.61mm). Immunohistology revealed complete repopulation of AO, CA, and JV scaffolds with endothelial cells and myofibroblasts within 2 weeks. After 8 weeks of implantation, AO scaffolds were completely covered by an endothelial monolayer and showed signs of a central matrix degeneration. JV scaffolds were completely degenerated at this stage. In contrast, CA scaffolds showed preserved ECM with a normal myofibroblast population and endothelial cell coverage.     

Decrease of some metabolic and physiologic effects of diazoxide by propranolol in rat

Summary Diazoxide significantly decreased the blood pressure and relaxed the uterine muscle in anaesthetized normotensive rats. A marked elevation of blood glucose followed the intravenous injection of diazoxide. The hyperglycemic and the uterine relaxing response could be significantly decreased by injection of propranolol prior to diazoxide. The hypotensive effect was not diminished by propranolol, however. In liver and uterus the content of cAMP was increased following diazoxide treatment in vivo. The rise in cAMP could be completely inhibited by propranolol, indicating a -receptor stimulation being the cause of the cAMP elevation.     

Expression pattern of the cell cycle promoter cyclin e in benign extravillous trophoblast and gestational trophoblastic lesions: correlation with expression of Ki-67.

In this study, a specific monoclonal antibody was used to immunohistochemically investigate correlated expression of the cell cycle promoter cyclin E and the proliferation marker Ki-67 in benign extravillous trophoblast and gestational trophoblastic lesions. Our data show that cyclin E is expressed in the normal extravillous trophoblast, with strongest levels of expression in the cell columns of anchoring villi. Differences could be observed in expression of Ki-67 in both normal extravillous trophoblast and gestational trophoblastic lesions. In the extravillous trophoblast of the cell columns, expression of cyclin E started more distal compared with Ki-67 and was maintained (with less intensity) into the deeper layers of interstitial trophoblast. In the benign trophoblastic lesions (exaggerated placental site [EPS] and placental site nodule [PSN]) and in the trophoblast proliferations on the surface of hydropic villi of hydatidiform moles (HM), the percentage of cells expressing cyclin E was higher than of those expressing Ki-67. The same observation could be made for a case of placental site trophoblastic tumor (PSTT). In contrast, choriocarcinomas (N=8), which are definitely malignant tumors, showed an opposite pattern, with a much higher percentage of strongly Ki-67-positive cells compared with cyclin E-positive cells. We conclude that cyclin E is expressed in benign extravillous trophoblast and gestational trophoblastic lesions, where a ratio cyclin E/Ki-67<1 characterizes choriocarcinomas, whereas PSTT and the benign lesions (HM, EPS, PSN) show expression of cyclin E in a higher percentage of cells than Ki-67 (cyclin E/Ki-67 ratio >1).