Generation of carbon monoxide and iron from hemeproteins in the presence of 7,8-dihydroneopterin

7,8-Dihydroneopterin and neopterin are secreted by human and primate macrophages after activation by interferon-gamma in a ratio of 2:1. 7,8-Dihydroneopterin is known to suppress radical-mediated processes, but it is also able in the presence of iron ions to generate superoxide radical anion and hydroxyl radicals from molecular oxygen. Effects of 7,8-dihydroneopterin were investigated on (met)myoglobin and (met)hemoglobin. Addition of 7,8-dihydroneopterin to heme proteins in air-saturated solution resulted in dose-dependent cleavage of the porphyrin moiety. The liberation of non-heme iron and carbon monoxide originating from the cleaved porphyrin was quantified. Both were generated at equimolar concentrations with a linear correlation coefficient of 0.9. Addition of ferrous iron significantly accelerated the pteridine-mediated cleaving of the porphyrin. However, the total yield of porphyrin cleaved was controlled by the pterin rather than by the ferrous ion concentration. 7,8-Dihydroneopterin is assumed to reduce the heme iron in intact protein molecules, thereby preparing the conditions for binding of oxygen and carbon monoxide as ligands. Beyond that, it is concluded that hydroxyl radicals might be generated via reduction of molecular oxygen to superoxide anion in the autoxidation process and dismutation to hydrogen peroxide and subsequent Fenton reaction.     

Procathepsin D expression correlates with invasive and metastatic phenotype of MDA-MB-231 derived cell lines

Procathepsin D (pCD) is a glycoprotein secreted abundantly by cancerous cells with a documented role in tumor development. The levels of pCD in primary tumors are highly correlated with an increased incidence of metastasis. Our earlier studies have shown that pCD exerts its effect on cancer cells through its activation peptide (AP) and involves both autocrine and paracrine modes of action. In this study, we analyzed the expression and role of pCD in MDA-MB-231 and its derived cell lines 1833 and 4175 possessing discrete metastatic abilities. Our results demonstrated a direct relationship between expression and secretion of pCD to the differential invasive potential of these cells. Also, the cell lines responded to AP treatment by enhancing their invasive potential, proliferation and induction of secretion of various cytokines, suggesting that pCD plays a role in metastasis through its AP region.     

Comparison of detection methods for Acarus siro (Acari: Acaridida: Acarididae) contamination in grain

Acarus siro L. 1758 (Acari: Acaridida: Acarididae) is an important pest of stored grain because it contaminates the grain by allergens and transfers pathogenous microorganisms. Rapid detection of contamination enables to intercept an early grain infestation by the pest. In this study, we compared the usability and efficiency of various detection approaches. Under laboratory conditions, grain samples of various sizes were infested by different levels of the following contaminants: eggs, adults, and feces of A. siro. The samples were analyzed by enzyme-linked immunosorbent assay (ELISA) by using anti-A. siro polyclonal antibody (immunochemical method), extracted in Berlese-Tullgren funnels, sieved, and processed by filth-flotation (conventional methods). The adults or juveniles of A. siro could be detected by all the three tested conventional methods and ELISA with detection limits in the range from 221 to 1,157 mites/kg grain. Eggs were detected by filth-flotation only; the detection limit was 1,950 eggs/kg grain. The feces of A. siro were detectable by ELISA test, only. ELISA enabled the detection of the feces with the minimal threshold level of 1.04 microg feces/g grain; it means the assay allowed to trace less than one metabolically active mite per gram of grain. The study thus demonstrated that reliable A. siro detection in grain can only be achieved by combining different detection methods. European Union and U.S. administratives dictate zero or near-zero tolerance level for mite infestation in stored products. This demand is difficult to fulfill, because every detection method is limited by its detection limit; thus, it is hard to reliably detect infestation levels lower than obtained detection limits. This methodical limitation is discussed in context with the determined detection limits of the tested methods.     

Physiological effects of different types of beta-glucan

Background: Numerous types of glucans have been isolated from almost every species of yeast, grain, and fungi. These products have been extensively studied for their immunological and pharmacological effects. Aim: In this paper we evaluated the possibility whether individual glucans will be similarly active against each of the tested biological properties or if each glucan will affect different reactions. Methods: Immunological effects of glucans were measured by evaluation of phagocytosis of HEMA particles by peripheral blood leukocytes and production of IL-2 by mouse splenocytes. Next we measured the effects of long-term treatment with glucan on levels of blood glucose and blood cholesterol. Four different glucans differing in origin (yeast, grain and mushroom) were used. Results: Our results showed that the same glucan, yeast-derived insoluble #300 glucan, stimulated phagocytosis of peripheral blood leukocytes, production of IL-2 by mouse splenocytes, lowered the cholesterol levels in mice with experimentally-induced cholesterolemia and lowered the level of blood sugar after induced hyperglycaemie. The remainder of tested glucans were only marginally active. Conclusion: Taken together, our study showed that with respect to natural glucans, there is a yes-or-no effect suggesting that highly purified and highly active glucans will have pleiotropic impact, whereas poorly isolated and/or less active glucans will have only mediocre biological properties.     

Spontaneous splenic rupture in two patients with hematologic malignancy

BACKGROUND: Spontaneous splenic rupture (SSR) is a very rare complication described in several hundred patients, mainly as case reports. It is defined as a splenic rupture without antecedent injury. The authors of the present paper describe the only two SSR cases diagnosed at the Hemato-oncology department, coincidentally in one year. PATIENTS: The first patient was admitted to hospital because of planned chemotherapy for relapsed hairy cell leukemia. The second was directed to the Hemato-oncology outpatient department because of anemia and painful splenomegaly diagnosed by a physician. The diagnose of hematologic malignancy (diffuse large B-cell lymphoma) was determined subsequently on the basis of histological examination of the spleen. CONCLUSION: It is necessary to consider SSR not only in patients with known diagnosis of malignant disease but in the patients with negative anamnesis, too. The aim of the paper is to draw attention to the existence of this complication.     

Glucan-like synthetic oligosaccharides: iterative synthesis of linear oligo-beta-(1,3)-glucans and immunostimulatory effects

Small reducing and linear oligo-beta-(1,3)-glucans, which are able to act as phytoallexin elicitors or as immunostimulating agents in anticancer therapy, were synthesized according to an iterative strategy that involved a unique key monosaccharidic donor. To avoid anomeric mixtures, the reducing entity of the target oligomers was first locked with benzyl alcohol and further selective deprotection of the 3-OH with DDQ afforded the desired building block as an acceptor. The latter was then used in a second cycle of glycosylation/deprotection to afford the desired disaccharide, and successive reiterations of this process provided the desired oligomers. Unusual conformational behaviors were observed by standard NMR sequences and supported by NOESY studies. Finally, removal of protecting groups afforded free tri-, tetra-, and pentaglucosides in good overall yields. Two oligosaccharides representing linear laminaritetraose and laminaripentaose were compared to the recently described beta-(1,3)-glucan phycarine. Following an intraperitoneal injection, the influx of monocytes and granulocytes into the blood and macrophages into the peritoneal cavity was comparable to that caused by phycarine. Similarly, both oligosaccharides stimulated phagocytic activity of granulocytes and macrophages. Using ELISA, we also demonstrated a significant stimulation of secretion of IL-1beta. Together these results suggest that the synthetic oligosaccharides have similar stimulatory effects as natural beta-(1,3)-glucans.