Widespread expression of ecto-apyrase (CD39) in the central nervous system

We have shown that ecto-apyrase protein is expressed in primary neurons and astrocytes in cell culture (T.-F. Wang, P.A. Rosenberg, G. Guidotti, 1997. Mol. Brain Res. 1997, 47: 295-302). Here we present immunohistochemical studies showing that ecto-apyrase protein is widely distributed in rat brain, as it is present in neurons of the cerebral cortex, hippocampus and cerebellum as well as in glial cells and endothelial cells. Ecto-apyrase is enriched in brain postsynaptic density membrane fractions and is localized in proximity to synaptophysin, the marker of synaptic vesicles. These results together with the observation that P2 purinergic receptors are present throughout the brain suggest that ecto-apyrase is involved in regulating synaptic transmission mediated by extracellular ATP.     

Malignant oncocytoma of the lacrimal sac: ultrastructure and immunocytochemistry

Malignant oncocytoma of the lacrimal sac is a rare tumor. A third example is presented here. In addition to oncocytes, tumor acini also harbored a few mucin cells and myoepithelial cells. The presence of immunoreactive lactoferrin and secretory piece suggested a resemblance to parotid acinic cell carcinomas. The difference was marked by a compact acinar structure and myoepithelial cell differentiation in the oncocytoma.     

Impaired production of IL-12 in systemic lupus erythematosus. I. Excessive production of IL-10 suppresses production of IL-12 by monocytes

Currently, primary osteoporosis is the most frequent metabolic disease in women after menopause [1]. The resulting loss of bone mass is accompanied by an increased risk of skeletal fragility. One reason for the development of osteoporosis might be an impaired function of mature osteoblasts. To evaluate the involvement of specific growth factors in bone remodeling, cell cultures of osteoblastic cells derived from nonosteoporotic and osteoporotic postmenopausal women were established. The influences of TGF beta-1 and IGF-I on proliferation and mRNA expression of TGF beta-1 were investigated by [3H]-thymidine incorporation and competitive RT-PCR. We found IGF-I to have no significant effect on proliferation in cells of osteoporotic and nonosteoporotic patients. In contrast, differences were found in TGF beta-1 mRNA expression after application of IGF-I. Application of TGF beta-1 enhanced its own mRNA expression in both groups in a similar manner. Whereas the proliferation of cells of nonosteoporotic patients was inhibited by (10(-10) M) TGF beta-1, this treatment led to an increased proliferation of cells of osteoporotic patients.     

Angiotensin II and insulin induce growth of ciliary artery smooth muscle: effects of AT1/AT2 antagonists

PURPOSE: Abnormal growth of smooth muscle cells (SMCs) in small arteries of the eye is associated with hypertension and diabetes, and the complications that they induce. Migration and proliferation of SMCs into the intima are primary mechanisms involved in neointima formation. In aortic SMCs, angiotensin II (AII)-induced proliferation is inhibited by angiotensin type 1 (AT1) receptor antagonist. However, in small artery SMCs, in particular in the circulation of the eye, the effects of AII on migration and proliferation are unknown. METHODS: The effects of AII (10(-6) to 10(-10) M) on migration and proliferation of growth-arrested SMCs of porcine ciliary arteries were studied in the presence and absence of insulin (5 x 10(-10) M) by assaying DNA synthesis (3H-thymidine incorporation), cell number, and movement of SMCs across the membrane of a modified Boyden chamber. RESULTS: In the absence of insulin, only high concentrations (10(-6) to 10(-8) M) of AII induced DNA synthesis and increased cell number (P < 0.05); however, in the presence of insulin (5 x 10(-10) M), AII induced DNA synthesis and cell number at low concentrations (10(-10) M) and in a concentration-dependent manner (P < 0.05). In contrast to proliferation, AII induced SMC migration in a concentration-dependent manner in the absence of insulin (P < 0.05). The AT1 antagonist CGP48933 (10(-8) to 10(-12) M), but not the AT2 antagonist CGP42112 (10(-8) to 10(-12) M), inhibited AII (10(-8) M)-induced proliferation and migration in a concentration-dependent manner (P < 0.05). CONCLUSIONS: Our results suggest that AII is a potent mitogen for SMCs of ophthalmic arteries, an effect that is enhanced in the presence of insulin, and that it may be an important contributor to structural vascular changes in the ophthalmic circulation in hypertension associated with non-insulin dependent diabetes. The inhibition of AII-induced growth by an AT1 antagonist suggests that these drugs may be important therapeutic tools to prevent structural vascular changes in the ophthalmic vasculature under these conditions.     

Mixed population approach for vaccination with live recombinant Salmonella strains

Attenuated strains of enteropathogenic species, such as Salmonella, represent useful carries for the delivery of heterologous recombinant antigens to the immune system. A frequently encountered obstacle, however, is the negative influence of high-level antigen production on the stability of carrier strains and the maintenance of their specific properties concerning tissue colonization and viability during infection. To solve this problem we have established an expression system based on genetic variation. This generates two sub-populations of a recombinant vaccine strain, i.e., one consisting of viable cells which maintain all characteristics of the native carrier strain and generate a second population of cells producing antigen(s) of interest at a very high level. This novel expression system offers unique applications and advantages over common live recombinant vaccine approaches.     

Endothelial function as an end-point in interventional trials: concepts, methods and current data

SURROGATE END-POINTS: Pharmacotherapy of cardiovascular disease has been increasingly validated in large interventional trials to assess its efficacy, safety and costs. As endpoints, morbidity and mortality are evaluated. More recently, surrogate end-points have been included in interventional trials, both to increase our understanding of pathogenic mechanisms and for their potential use as markers of risk in patients. ENDOTHELIAL FUNCTION: The endothelium lies in a strategic anatomical position between the circulating blood and vascular smooth muscle and hence is a major local mediator of cardiovascular function. Also, endothelial cells are a target for mechanical forces and cardiovascular risk factors in the circulation. Thus, it is not surprising that their function becomes impaired at an early stage in the disease process. The cells are able to produce numerous proteins and mediators. This review focuses on nitric oxide and endothelin-1, which are endothelium-derived relaxing and constrictor factors, respectively. Nitric oxide also prevents platelet adhesion and aggregation and the adhesion of monocytes. Both substances also affect vascular structure in that nitric oxide inhibits while endothelin stimulates vascular smooth muscle proliferation and migration. MEASUREMENTS OF ENDOTHELIAL FUNCTION: Endothelial function and the effects of nitric oxide and endothelin in particular can be evaluated in the coronary circulation by quantitative coronary angiography and Doppler flow wire, and in the peripheral circulation with plethysmography and new ultrasound/Doppler devices. In these experimental set-ups, lipid-lowering drugs and angiotensin converting enzyme (ACE) inhibitors have been evaluated. Lipid-lowering drugs improve endothelium-dependent vasodilation in the coronary and forearm circulation of patients with hyperlipidemia and atherosclerosis. Similarly, ACE inhibitors improve coronary vasomotion in patients with coronary artery disease and normal lipid levels. In hypertension, ACE inhibitors have failed to improve endothelium-dependent vasodilation, while studies with other drugs are planned. CONCLUSIONS: Endothelial function can now be assessed precisely in patients in vivo, in both the coronary and the peripheral circulation. Tests can detect early dysfunction in patients with a risk of cardiovascular disease and the possible effects of drugs on endothelial function. Large interventional trials are needed to establish how far endothelial dysfunction can or cannot predict clinical outcome.     

Efficient lymphocyte migration across high endothelial venules of mouse Peyer's patches requires overlapping expression of L-selectin and beta7 integrin

Lymphocyte migration into lymphoid organs is regulated by adhesion molecules including L-selectin and the beta7 integrins. L-selectin and alpha4beta7 are predominantly hypothesized to direct the selective migration of lymphocytes to peripheral lymph nodes and the gut-associated lymphoid tissues, respectively. To further characterize interactions between L-selectin and beta7 integrins during lymphocyte recirculation, mice deficient in both receptors (L-selectin/beta7 integrin-/-) were generated. The simultaneous loss of L-selectin and beta7 integrin expression prevented the majority of lymphocytes (>95% inhibition) from attaching to high endothelial venules (HEV) of Peyer's patches and other lymphoid tissues during in vitro binding assays. Moreover, the inability to bind HEV eliminated the vast majority of L-selectin/beta7 integrin-/- lymphocyte migration into Peyer's patches during short-term and long-term in vivo migration assays (>99% inhibition,p < 0.01). The lack of lymphocyte migration into Peyer's patches correlated directly with the dramatically reduced size and cellularity (99% reduced) of this tissue in L-selectin/beta7 integrin-/- mice. High numbers of injected L-selectin/beta7 integrin-/- lymphocytes remaining in the blood of wild-type mice correlated with markedly increased numbers of circulating lymphocytes in L-selectin/beta7 integrin-/- mice. Loss of either L-selectin or the beta7 integrins alone resulted in significant but incomplete inhibition of Peyer's patch migration. Collectively, the phenotype of L-selectin/beta7 integrin-/- mice demonstrates that these two receptors primarily interact along the same adhesion pathway that is required for the vast majority of lymphocyte migration into Peyer's patches.     

Losartan but not verapamil inhibits angiotensin II-induced tissue endothelin-1 increase: role of blood pressure and endothelial function

Endothelin partially mediates angiotensin (Ang) II-induced vascular changes in vivo. This study investigated the effects of the angiotensin type 1 receptor antagonist losartan and the calcium channel blocker verapamil on vascular reactivity and tissue endothelin-1 levels in aortas of Wistar-Kyoto rats treated for 2 weeks with Ang II (200 ng x kg(-1) x min(-1)). Ang II increased systolic blood pressure (39+/-4 mm Hg, P<0.05). Concomitant treatment with losartan abolished the Ang II-induced pressure increase (P<0.05), whereas verapamil reduced it only partially (P<0.05). In the aortas of rats with Ang II-induced hypertension, tissue endothelin-1 content was increased threefold and contractions to endothelin-1 were impaired (P<0.05). Interestingly, these alterations were normalized by losartan (P<0.05) but not by verapamil. Hence, there was a strong, negative correlation between contractions to endothelin-1 and tissue endothelin-1 content (r=-0.733, P<0.0001). In contrast, both antihypertensive drugs normalized impaired endothelium-dependent relaxations to acetylcholine and reduced the sensitivity of vascular smooth muscle to sodium nitroprusside compared with Ang II-treated rats (P<0.05). Ang II-induced hypertension enhanced endothelium-dependent contractions to acetylcholine, and these were normalized by either drug. In conclusion, these findings suggest that long-term treatment with Ang II modulates endothelin-1 protein expression in the rat aorta. Although both antihypertensive agents lowered blood pressure and normalized endothelial function, only losartan prevented the increase in tissue endothelin-1 content, suggesting that angiotensin type 1 receptor antagonists but not calcium antagonists modulate tissue endothelin-1 in vivo.     

Delivery of salbutamol to nonventilated preterm infants by metered-dose inhaler, jet nebulizer, and ultrasonic nebulizer

An isocratic reversed-phase high-performance liquid chromatographic method with ultraviolet detection at 230 nm has been developed for the determination of paclitaxel in human plasma. Plasma samples were prepared by a selective one-step liquid-liquid extraction involving a mixture of acetonitrile-n-butyl chloride (1:4, v/v). Paclitaxel and the internal standard docetaxel were separated using a column packed with ODS-80A material, and a mobile phase consisting of water-methanol-tetrahydrofuran-ammonium hydroxide (37.5:60:2.5:0.1, v/v). The calibration graph for paclitaxel was linear in the range 10-500 ng/ml, with a lower limit of quantitation of 10 ng/ml, using 1 ml plasma samples. The extraction recoveries of spiked paclitaxel and docetaxel to drug-free human plasma were 89.6+/-8.52 and 93.7+/-5.0%, respectively. Validation data showed that the assay for paclitaxel is sensitive, selective, accurate and reproducible. The assay has been used in a single pharmacokinetic experiment in a patient to investigate the applicability of the method in vivo.