Human skin chymotrypsin-like proteinase chymase. Subcellular localization to mast cell granules and interaction with heparin and other glycosaminoglycans

The subcellular localization of human skin chymase to mast cell granules was established by immunoelectron microscopy, and binding of chymase to the area of the dermo-epidermal junction, a basement membrane, was demonstrated immunocytochemically in cryosections incubated with purified proteinase prior to immunolabeling. Because heparin and heparan sulfate proteoglycans are major constituents of mast cell granules and basement membranes, respectively, the ability of chymase to bind to glycosaminoglycans (GAG) was investigated. Among a variety of GAGs, only binding of chymase to heparin and heparan sulfate appears physiologically significant. Binding was ionic strength-dependent, involved amino groups on the proteinase, and correlated with increasing GAG sulfate content, indicating a predominantly electrostatic association. Interaction with heparin was observed in solutions containing up to 0.5 M NaCl, and interaction with heparan sulfate was observed in solutions containing up to 0.3 M NaCl. Binding of heparin did not detectably affect catalysis of peptide substrates, but may reduce accessibility of proteinase to protein substrates. Measurements among a series of serine class proteinases indicated that heparin binding was a more common property of mast cell proteinases than proteinases stored in other secretory granules. Binding of chymase to heparin is likely to have a storage as well as a structural role within the mast cell granule, whereas binding of chymase to heparan sulfate may have physiological significance after degranulation.     

Comparative tissue ascorbic acid studies in fishes

Comparative tissue ascorbic acid levels in four species of major carp viz., Labeo rohila, L. calbasu, Cirrhina tnrigala and Catla catla , were investigated. The ascorbic acid level was found to be the highest in the spleen in the four species studied (range 430–380 μg/g) followed by the anterior (adrenal) kidney, gonads, liver, renal kidney, brain and/or eye. Heart and blood had the lowest levels (range 26–18 μg/ml) amongst the tissues studied. Overall tissue ascorbic acid levels were the highest in L. rohita and the lowest in C. mrigala . Investigation on seasonal variations in blood and kidney ascorbic acid levels of Notopterus notopterus revealed peak levels in spring (February-April) and the lowest levels in the postspawning period (August-September).     

Parathyroid hormone stimulates adenylate cyclase in rat cerebral microvessels

We have studied the effect of parathyroid hormone (PTH) on adenylate cyclase of microvessels isolated from rat cerebral cortex. Native bovine (b) PTH-(1–84), the synthetic amino-terminal fragment bPTH-(1–34) and the synthetic analog [Nle⁸, Nle¹⁸, Tyr³⁴]-bPTH- (1–34) amide stimulated adenylate cyclase in a dose-dependent manner with apparent ED₅₀ values of 16 nM, 6.3 nM and 15 nM respectively. The stimulation by bPTH was greatly enhanced by guanosine triphosphate. The PTH antagonist, [Nle⁸, Nle¹⁸, Tyr³⁴]-bPTH-(3–34) amide inhibited the action of bPTH-(1–84) and bPTH-(1–34). In summary, PTH stimulated adenylate cyclase in rat cerebral microvessels in a very similar manner to its stimulation in the renal cortex.     

Effects of Autogamy In Paramecium Tetraurelia On Catalase Activity and On Radiosensitivity to Natural Ionizing Radiations

SYNOPSIS Catalase activity of Paramecium tetraurelia decreased during autogamy and recovered to normal 5 days later. Autogamy also caused changes in the ciliate's sensitivity to natural ionizing radiations—the decrease in cell growth rate previously described in shielded cultures did not occur when autogamous cells were used. Maximum effect of shielding was observed in 11-day-old postautogamous cells. the role of the catalase in the mechanism of natural irradiation effect is discussed.     

An apparatus to measure in vivo biomechanical behavior of dorsi- and plantarflexors of mouse ankle.

We developed an apparatus to quantify the biomechanical behavior of the dorsi- and plantarflexor muscles of the ankle of an anesthetized mouse. When the dorsi- or plantarflexor muscle group is activated by electrical stimulation of either the peroneal or tibial nerve, the apparatus measures the moment developed about the ankle during isometric, isovelocity shortening, or isovelocity lengthening contractions. Displacements may be performed over the full 105 degrees range of ankle motion with an angular resolution of 0.09 degrees. Bidirectional isovelocity ramps in ankle angle up to 1,100 degrees/s are possible and are equivalent to maximum velocities of 2.3 fiber lengths/s (Lf/s) for the fibers in tibialis anterior muscle and 11.9 Lf/s for those in gastrocnemius muscle. During single contractions of the dorsi- and plantarflexor muscle groups at 37 degrees C and with both knee and ankle joint at 90 degrees neutral position, the isometric tetanic force developed was 1.4 and 3.3 N, power output at 2.2 Lf/s was 3.1 and 5.9 mW, and power absorption at 0.5 Lf/s was 4.9 and 9.0 mW, respectively. These values are in reasonable agreement with data from the same muscle groups tested in situ. We conclude that the apparatus provides valid measurements of force and power of the dorsi- and plantarflexor muscle groups.     

Human placental tissue stimulates bovine capillary endothelial cell growth,migration and protease production

A crude extract of human placenta has been demonstrated to stimulate growth, motility and the production of the proteases plasminogen activator and collagenase in cultured bovine capillary endothelial cells. These data are in keeping with the presence of an angiogenic factor(s) in human placenta.     

Initiation of the extrinsic pathway of coagulation. Association of factor VIIa with a cell line expressing tissue factor

We have examined initial assembly of the extrinsic pathway of blood coagulation on cell surfaces with radiolabeled human factor VIIa and a human fetal lung cell line possessing abundant functional tissue factor activity. Binding of factor VIIa to these cells was observed and was time- and temperature-dependent. Binding of factor VIIa was quantitatively equivalent at 37 and 6 degrees C, although the kinetics of binding differed. The radiolabeled ligand bound by the cell was indistinguishable by sodium dodecyl sulfate-polyacrylamide gel analysis from the factor VIIa offered. Factor VIIa binding was influenced by calcium ions. The binding appears to involve at least two classes of calcium-dependent binding sites. Optimal binding occurred at 2 mM calcium for both classes of sites, and there was inhibition of binding to the high affinity sites at higher calcium. Association of factor VIIa was specific, saturable, had a Kd of 123 +/- 37 pm, and factor VIIa interacted with about 100,000 binding sites per cell. Once established, specific binding was rapidly reversible. Direct cellular binding of human factor X also was observed and was calcium, time- and temperature-dependent. Factor X binding was specific and saturable with half-maximal binding at 87.6 +/- 27.4 nM to 6.03 +/- 1.03 X 10(6) sites per cell. Specific high affinity binding of factor VIIa correlated with generation of factor Xa. A direct linear relationship was observed at low factor VIIa binding; however, at higher bound factor VIIa, the relationship was nonstoichiometric, i.e. less factor Xa was formed per mole of factor VIIa. Expression of specific binding sites for factors VIIa and X provides further substantiation for the molecular assembly hypothesized to initiate the extrinsic coagulation protease cascade on cells.