Narendranath S. Ranadive

Histamine Levels in Middle Ear Effusions

The presence of histamine in 131 middle ear effusions was determined by the fluorometric assay technique. This potent mediator of inflammation was found in significant amounts in most of the samples, suggesting that it may play an important role in the pathogenesis of otitis media with effusion. It is postulated that mast cells located in the lamina propria of the tympanic mucoperiosteum are triggered to degranulate and release histamine by anaphylatoxin derived from activation of the complement system.

Movement of Calcium Ions and Release of Histamine from Rat Mast Cells

Rat mast cells were stimulated for histamine release in a medium containing radioactive calcium, by band 2 protein (B2) and compound 48/80. It was found that a significant amount of extracellular calcium was taken up by the stimulated cells. When the histamine release process was divided into two stages by activating the cells at 0 degrees C and then washing them prior to suspending them in Tyrodes solution at 37 degrees C, it was found that calcium uptake by the cells took place at the release stage. This suggests that calcium entry into the cells occurs subsequent to the activation stage. Inhibition of histamine release by 2-deoxyglucose (2-DG) and 2,4-dinitrophenol (DNP) also inhibited the calcium influx into the cells. The present studies also suggest that calcium does not diffuse into the cells as a result of degranulation. These findings have been discussed in relation to the mobilization of intracellular calcium.

The visualization of receptors for the Fc portion of the IgG molecule on human neutrophil leukocytes

Abstract Human neutrophils treated with either Fc or with intact IgG and subsequently with fluorescein-labelled anti-IgG showed binding of the Fc or the IgG to the cell membrane. Neutrophils did not appear to bind F(ab′) 2 fragments. Under suitable conditions, polar capping of fluorescence was seen. The data suggest receptors for Fc on the neutrophil membrane and mobility of these receptors.

Quantitation of cutaneous inflammation induced by reactive species generated by UV-visible irradiation of Rose bengal

The present studies were undertaken to quantitate the initial inflammatory response produced by the photo-generated reactive species in rabbit skin. Rose bengal (RB), a photosensensitizer dye, was injected into the skin sites at various concentrations and exposed to UV-visible light for 30–120 min. The increase in vascular permeability and the accumulation of PMNs were investigated using125I-labeled albumin and51Cr-labeled PMNs. RB at a concentration of 1 nmol with 120-min exposure to light enhanced vascular permeability by 3.7 times and accumulation of PMNs by 3.3 times. As low as 0.01 nmol of RB produced discernible effects.β-Carofene (0.1 nmole) inhibited the inflammatory response by 75–100%, suggesting that the reactive species involved in this response was predominantly singlet oxygen. The increase in vascular permeability was inhibited by 48–70% by 25μg of chlorpheniramine maleate. It is therefore suggested that histamine plays a major role in the initial vascular response. The studies demonstrate that this rabbit model is suitable for the quantitation of photoinduced inflammatory response which is not observable by gross anatomic procedures.

Release of basic proteins and lysosomal enzymes from neutrophil leukocytes of the rabbit.

Rabbit neutrophil leukocytes incubated with immune precipitates, with rabbit γ -globulin heat-aggregated at 68 °C, or with rabbit γ -globulin-coated latex particles

Modulation of phagocytosis and intracellular bactericidal activity of polymorphonuclear and mononuclear cells by cationic proteins from human granulocytes: Alternative pathway of phagocytic enhancement

Cationic lysosomal proteins from human polymorphonuclears (PMN) were isolated by column chromatography and divided into five fractions. On acrylamide gel electrophoresis, fraction I had four bands slower than lysozyme (LZM) mobility; fraction II had five or six bands slower than LZM; fraction III had at least seven bands slower and two bands faster than LZM; fraction IV contained LZM, two bands faster and a few faint bands slower than LZM; fraction V was composed of almost pure LZM. Partial characterization of the fractions showed presence of neutral protease in fractions I-IV, chymotrypsin in fraction III, lysozyme in fractions IV and V, and phospholipase A2 mainly in fractions II and III. Modulatory activity of fractions I-V were tested at concentrations up to 50 μg/ml. Enhancement of phagocytosis ofStaphylococcus aureus was observed by fractions I, IV, and V, whereas phagocytic index was enhanced by all but the fraction II. Intracellular bactericidal activity (ICBA) was markedly enhanced by fractions I, II, and V. Addition of DNA or cytochalasine B inhibited or abolished phagocytosis-enhancing activity of cationic fractions. Their influence on ICBA was much less pronounced. Fraction III enhanced phagocytic index and phagocytosis ofE. colt, whereas fractions I and II enhanced intracellular bactericidal activity against this bacteria. Enhancement of phagocytic activity of monocytes has also been observed. The data suggest that some cationic lysosomal fractions from human PMNs enhance phagocytosis and phagocytic index by human PMNs and monocytes and intracellular bactericidal activity of human PMNs. This alternative pathway of phagocytic enhancement is unrelated to the previously described enhancers of phagocytosis and may play a role in defense mechanisms against infection.

Mechanism of histamine release from rat mast cells by compound 48-80. Comparison with the release induced by cationic protein.

Rat peritoneal mast cells incubated with compound 48/80 at temperatures below 10 °C and washed released histamine upon warming to 37 °C. Evidence has been put forward to show that this release of hist

Influence of neutrophil cationic proteins on generation of superoxide by human polymorphonuclear cells during phagocytosis

The cationic proteins from neutrophyl lysosomes have been shown to modulate phagocytic activity of granulocytes. The present study reports the effects of the cationic protein fractions on the generation of O2− by human PMNs during phagocytosis. Human PMNs were reacted win different phagocytic stimuli in the presence and absence of lysosomal cationic proteins and the amount of O2− generated was determined by superoxide dismutase inhibitable reduction of cytochromec. Total cationic protein extract from neutrophil lysosomes enhanced O2− generated by PMNs during the phagocytosis of IgG-coated latex beads and opsonized zymosan particles. The analysis of the fractions of cationic proteins obtained from a Sephadex G-75 column showed that the O2− generation-enhancing activity was associated with the proteins eluted in fractions III and IV. A protein fraction mainly eluted in void volume inhibited the cytochromec reduction by O2− formed during phagocytosis. This was due to the presence of superoxide dismutase-like activity since O2− generated by the xanthine-xanthine oxidase system was also inhibited by this fraction. The cationic protein fractions III and IV from the Sephadex G-75 column were further subfractionated. Although the O2−-enhancing activity was eluted in the same fractions as chymotrypsin activity, there was no quantitative correlation between the amount of O2− generation and chymotrypsin activity. Moreover, commercial chymotrypsin did not enhance O2− generation. Electrophoretic analysis of the isolated protein fractions suggests that O2− generation enhancing protein (SGEP) is different from lysozyme or chymotrypsin and probably represents previously undescribed protein.

Differential effects of antioxidants and indomethacin on compound 48/80 induced histamine release and Ca2+ uptake in rat mast cells

The effect of nordihydroguaiaretic acid (NDGA), vitamin E, butylated hydroxytoluene (BHT) and indomethacin on histamine release and Ca2+ uptake in rat mast cells stimulated with compound 48/80 was studied. NDGA inhibited both the release of histamine and Ca2+ uptake in stimulated cells; however, there was no correlation between inhibition of Ca2+ uptake and the amount of histamine release. At a concentration of 5 microM, NDGA completely inhibited Ca2+ uptake, while histamine release was decreased by less than 50%. BHT (50 microM) inhibited both the Ca2+ uptake and histamine release. On the other hand, vitamin E (50 microM) inhibited histamine release by 70% without impairment in Ca2+ uptake. In the absence of the stimulus, vitamin E increased the cell-associated Ca2+; however, it had no effect on spontaneous release of histamine. Indomethacin (3 microM) inhibited Ca2+ uptake in stimulated cells by 50%, but did not affect the release of histamine. The results suggest that a part of Ca2+-influx may not be related to the coupled activation--secretion response and that lipid peroxidation through the lipoxygenase pathway may be involved in secretion of histamine from mast cells.

Influence of cationic superoxide generation enhancing protein (SGEP) on phagocytic and intracellular bactericidal activity of human polymorphonuclear cells

Cationic fraction III from the lysosomes of normal human peripheral blood polymorphonuclear cells (PMNs) was found to contain superoxide generation enhancing protein (SGEP). Herein, we report on the influence of partially purified SGEP obtained from fraction III (subfractions III-5 and III-6), on various phagocytic functions of human PMNs. SGEP markedly enhanced intracellular bactericidal activity of human peripheral PMNs. The enhancement was time and dose dependent. It also reduced adhesiveness of the PMNs. SGEP did not influence chemotaxis, phagocytosis or phagocytic index. These findings are compatible with our original observation regarding superoxide generation enhancement properties of SGEP.