Allergy against chironomids (non biting midges) in adult atopic patients

Summary The aim of this investigation was to evaluate the prevalence of atopic sensitization to chironomids (CHI) in patients with asthma and/or rhinitis (A/R), and to study concomitant sensitization to CHI and other allergens. Skin prick tests were performed with 3 different CHI extracts as well as with common inhalant allergens in 600 consecutive patients, 495 of which had A/R. Allergen specific IgE antibodies in the sera against CHI, shell fish and cockroaches were analyzed with Magic Lite.59 (12%) of the patients with A/R had a positive skin test with CHI. Positive skin tests with house dust mites and a storage mite were more common in CHI allergic patients than in other atopic patients. Nasal or conjunctival provocation tests, performed on 23 of the patients with positive skin test with CHI, were clearly positive in 7 cases (30%), questionable in 8 (35%) and negative in 8 cases (35%).Magic Lite, performed on sera from 50 of the patients with positive skin test against CHI, was positive with CHI in 39 cases (78%), with crayfish in 33 (66%), shrimp 20 (40%), cockroach 21 (40%) and with crab in 3 cases (6%).It is concluded that sensitization against CHI is common in patients with A/R. The clinical relevance of the positive test results is, however, unknown. Concomitant sensitization with CHI, crustaceans and cockroach is common.     

Reduction of 2,4,6-trinitrobenzenesulfonate by glutathione reductase and the effect of NADP+ on the electron transfer

Glutathione reductase has been found to catalyze an NAD(P)H-dependent electron transfer to 2,4,6-trinitrobenzenesulfonate (TNBS). In the presence of oxygen TNBS is not consumed in the reaction, but is rapidly reoxidized with concomitant production of hydrogen peroxide. Cytochrome c can replace oxygen as the final electron acceptor, indicating that a one-electron transfer takes place. The rate is slightly higher in the absence than in the presence of oxygen, ruling out superoxide anion as an obligatory intermediate in cytochrome c reduction. In the absence of oxygen (or cytochrome c), TNBS limits the reaction and accepts a total of four electrons. The TNBS-dependent NADPH (or NADH) oxidation is markedly stimulated by NADP+, and to a smaller extent also by NAD+. The TNBS-dependent reactions are inhibited by excess of NADPH but not by NADH. The kinetics of these reactions are consistent with a branching reaction mechanism in which a pathway including a ternary complex between the two-electron reduced enzyme and NADP+ has the highest turnover. NADPH-dependent reductions of ferricyanide or 2,6-dichloroindophenol catalyzed by glutathione reductase are also markedly influenced by NADP+. Evidently NADP+ facilitates a shift of the catalyzed reaction from the normal two-electron reduction of glutathione disulfide to a more unspecific one-electron reduction of other acceptors. Spectral as well as kinetic data suggest that the rate of radical formation limits the reactions with the artificial electron acceptors and that NADP+ promotes this rate-limiting step.     

Isolation and Partial Purification of Prophenoloxidase from Daucus carota L. Cell Cultures

The enzyme, phenoloxidase, was isolated and partially purified as an inactive enzyme, a proenzyme, from plant cell cultures of Daucus carota, Nicotiana tabacum, and Haplopappus gracilis. The prophenoloxidase was found to be specifically activated by Ca²⁺ or Mn²⁺ ions in concentrations above 1 millimolar. Calmodulin was not involved in this activation. Concentrations of Ca²⁺ or Mn²⁺ below 1 millimolar could not induce activation of the prophenoloxidase, but if trypsin was added simultaneously with Ca²⁺ or Mn²⁺ at a concentration of 1 millimolar or below, the proenzyme was converted to its active form. The inactive form of phenoloxidase was found to be a soluble enzyme, whereas after activation the enzyme aggregated, and a significant amount of the enzyme activity could become pelleted.     

Disruption of estrous cycles in exercise-trained rats

The female Sprague-Dawley rat was evaluated as an animal model for the menstrual irregularities that are common in women athletes. Daily vaginal smears revealed that estrous cycles were markedly disrupted in rats during a 10-week exercise training program, while cycles remained normal in sedentary rats. Compared to 9 sedentary rats, the 10 exercise-trained rats had longer mean cycle lengths and fewer estrus smears. Six of the exercise-trained rats, but none of the sedentary rats, had an "anestrus period" with more than twice the normal interval between estrus smears; one exercise-trained rat became essentially acyclic. Weight gain during the 10-week training program was lower in exercise-trained rats than in sedentary rats. Colonic temperatures, monitored at rest and during 30 min of exercise, were slightly lower in exercise-trained rats with irregular estrous cycles than in exercise-trained rats with regular cycles, indicating that unusually elevated body temperatures during exercise are not responsible for exercise-related reproductive acyclicity. It is concluded that the female Sprague-Dawley rat may be a useful animal model for the study of menstrual irregularities associated with exercise training.     

Protease activities in non-embryogenic and embryogenic carrot cell strains during callus growth and embryo formation

Embryogenic and non-embryogenic cell strains of Daucus carota L. were examined for their protease activity using a wide range of chromogenic synthetic peptides as substrates. High arginine-specific activity was present in all strains, but no protease activity "specific" for embryogenic or non-embryogenic strains could be detected with the substrates tested. The specific protease activity was 5–10 times higher in the non-embryogenic as compared to the embryogenic strain for most tested substrates, and this difference was not due to release of proteases in the latter. All strains showed a decrease in protease activity when cultured in media without 2,4-dichlorophenoxyacetic acid, but the embryos had high protease activity in comparison with the nondifferentiated cell aggregates. In the latter aggregates, hydrolyzing activity towards three of the substrates (H-D-Phe-Pip-Arg- p -nitroanilide, Suc-Ala-Pro-Phe- p -nitro-anilide and Bz-Phe-Val-Arg- p -nitroanilide) was absent, whereas the embryos were able to hydrolyze them.     

Na+, Cl− cotransport in Ehrlich ascites tumor cells activated during volume regulation (regulatory volume increase)

Ehrlich ascites cells were preincubated in hypotonic medium with subsequent restoration of tonicity. After the initial osmotic shrinkage the cells recovered their volume within 5 min with an associated KCl uptake. The volume recovery was inhibited when NO-3 was substituted for Cl-, and when Na+ was replaced by K+, or by choline (at 5 mM external K+). The volume recovery was strongly inhibited by furosemide and bumetanide, but essentially unaffected by DIDS. The net uptake of Cl- was much larger than the value predicted from the conductive Cl- permeability. The undirectional 36Cl flux, which was insensitive to bumetanide under steady-state conditions, was substantially increased during regulatory volume increase, and showed a large bumetanide-sensitive component. During volume recovery the Cl- flux ratio (influx/efflux) for the bumetanide-sensitive component was estimated at 1.85, compatible with a coupled uptake of Na+ and Cl-, or with an uptake via a K+,Na+,2Cl- cotransport system. The latter possibility is unlikely, however, because a net uptake of KCl was found even at low external K+, and because no K+ uptake was found in ouabain-poisoned cells. In the presence of ouabain a bumetanide-sensitive uptake during volume recovery of Na+ and Cl- in nearly equimolar amounts was demonstrated. It is proposed that the primary process during the regulatory volume increase is an activation of an otherwise quiescent, bumetanide-sensitive Na+,Cl- cotransport system with subsequent replacement of Na+ by K+ via the Na+/K+ pump, stimulated by the Na+ influx through the Na+,Cl- cotransport system.     

Role of calcium binding by sarcoplasmic reticulum in the contraction and relaxation of uterine smooth muscle

The binding of calcium by isolated sarcoplasmic reticulum from cow uterus was studied. Sarcoplasmic reticulum was prepared by differential centrifugation. Three fractions were obtained: I, sedimented between 2,500–15,000 x g; II at 40,000 x g; and III, at 150,000 x g. Fraction II was further purified on a sucrose density gradient. All three fractions contained considerable amounts of intrinsic calcium, mostly in fraction I. Calcium binding in the presence of ATP¹ and Mg also was greatest in fraction I, followed by fraction II, with less in fraction III. Without ATP no calcium was taken up. 5 and 10 mM sodium azide partially inhibited calcium binding in fraction I, but not in fraction II, suggesting the presence of some mitochondria or mitochondrial fragments in fraction I. Calcium binding in fraction II was completely inhibited by 3 mM salyrgan; this fraction thus appears to be sarcoplasmic reticulum. ATPase activity was found in all three fractions, highest in fraction II. It is computed that calcium binding in fractions I and II, on the basis of a 50% yield of protein, is sufficient to elicit contraction by supplying calcium to the contractile proteins of the smooth muscle cell and to regulate relaxation and contraction.