Different defense strategies of Dendrolimus pini, Galleria mellonella, and Calliphora vicina against fungal infection.

The resistance of Galleria mellonella, Dendrolimus pini, and Calliphora vicina larvae against infection by the enthomopathogen Conidiobolus coronatus was shown to vary among the studied species. Exposure of both G. mellonella and D. pini larvae to the fungus resulted in rapid insect death, while all the C. vicina larvae remained unharmed. Microscopic studies revealed diverse responses of the three species to the fungal pathogen: (1) the body cavities of D. pini larvae were completely overgrown by fungal hyphae, with no signs of hemocyte response, (2) infected G. mellonella larvae formed melanotic capsules surrounding the fungal pathogen, and (3) the conidia of C. coronatus did not germinate on the cuticle of C. vicina larvae. The in vitro study on the degradation of the insect cuticle by proteases secreted by C. coronatus revealed that the G. mellonella cuticle degraded at the highest rate. The antiproteolytic capacities of insect hemolymph against fungal proteases correlated well with the insects' susceptibility to fungal infection. The antiproteolytic capacities of insect hemolymph against fungal proteases correlated well with the insects' susceptibility to fungal infection. Of all the tested species, only plasmatocytes exhibited phagocytic potential. Exposure to the fungal pathogen resulted in elevated phagocytic activity, found to be the highest in the infected G. mellonella. The incubation of insect hemolymph with fungal conidia and hyphae revealed diverse reactions of hemocytes of the studied insect species. The encapsulation potential of D. pini hemocytes was low. Hemocytes of G. mellonella showed a high ability to attach and encapsulate fungal structures. Incubation of C. vicina hemolymph with C. coronatus did not result in any hemocytic response. Phenoloxidase (PO) activity was found to be highest in D. pini hemolymph, moderate in G. mellonella, and lowest in the hemolymph of C. vicina. Fungal infection resulted in a significant decrease of PO activity in G. mellonela larvae, while that in the larvae of D. pini remained unchanged. PO activity in C. vicina exposed to fungus slightly increased. The lysozyme-like activity increased in the plasma of all three insect species after contact with the fungal pathogen. Anti E. coli activity was detected neither in control nor in infected D. pini larvae. No detectable anti E. coli activity was found in the control larvae of G. mellonella; however, its exposure to C. coronatus resulted in an increase in the activity to detectable level. In the case of C. vicina exposure to the fungus, the anti E. coli activity was significantly higher than in control larvae. The defense mechanisms of D. pini (species of economic importance in Europe) are presented for the first time.

Basic properties of the pyruvate dehydrogenase complex isolated from aurochs heart.

The purified aurochs (Bison bonasus, European bison) heart pyruvate dehydrogenase complex (PDC) has a set of subunits typical of mammalian PDC. PDC from aurochs heart contains firmly bound tiamine pyrophosphate in the amount providing over 50% of the maximal activity of the complex. The apparent value for activation energy of PDC is 60 kJ/mol. The Michaelis constant values for aurochs heart PDC are 22.4 +/- 1.0, 3.3 +/- 0.1 and 24.4 +/- 3.6 microM for pyruvate, CoA and NAD, accordingly. Acetyl-CoA is a competitive inhibitor with respect to CoA (Ki = 14.2 +/- 0.4 microM), whereas NADH gives the same inhibition with respect to NAD (Ki = 46.9 +/- 10.0 microM). The Km for CoA and NAD of the aurochs heart PDC are lower than that of domestic animals PDC.

Cooperation of divalent ions and thiamin diphosphate in regulation of the function of pig heart pyruvate dehydrogenase complex.

The role of Mg2+, Ca2+, and Mn2+ in regulation of purified pig heart pyruvate dehydrogenase complex (PDC) containing endogenous thiamin diphosphate (TDP) was studied. It was found that the effects of the cations depended on the presence of exogenous TDP. In the absence of added TDP, the divalent cations led to a shortening of a lag phase of the PDC reaction and a strong reduction of the Km value for pyruvate. The relative efficiency of the three types of ions are presented as follows: Mn2+>Ca2+>Mg2+. The other sources claim that in the presence of exogenous TDP, which alone strongly increased the affinity of PDC for pyruvate, any significant additional effects of the cations were not observed. However, Mg2+, Ca2+, and Mn2+ decreased the Km value for CoA in both cases, the absence and presence of exogenous TDP, in approximately a similar extent (about twofold). The affinity of PDC for NAD+ seems to be not sensitive to the presence of the divalent cations. The data obtained suggest that Mg2+, Ca2+, and Mn2+ can cooperate with TDP as positive regulatory effectors of pig heart PDC on the level of pyruvate dehydrogenase and lipoamide acetyltransferase components of the complex.

[Phagocytosis and killing of Staphylococcus aureus by granulocytes in mice after granulocyte-macrophage colony stimulating factor (GM-CSF) injection]. / Fagocytoza i zabijanie komórek Staphylococcus aureus przez granulocyty myszy poddanych dzialaniu stymulatora kolonii granulocytarno-makrofagowych (GM-CSF).

GM-CSF is a hematopoietic growth factor. In vitro it stimulates the proliferation of myeloid progenitors and formation of granulocyte and macrophage colonies. It was found that GM-CSF in vitro is also stimulated the function of mature granulocytes, but we have no information about such influence in vivo. The purpose of this investigation was the evaluation in vivo of the GM-CSF effect on phagocytosis, bactericidal activity, and lysosome enzyme activities in granulocytes. GM-CSF was injected into mice subcutaneously during 5 consecutive days in the dose of 1 microgram/kg/d. The examination of the percent of cell phagocytizing bacteria (Staphylococcus aureus), NBT test, bactericidal activity and activation of acid phosphatase, alkaline phosphatase, peroxidase and esterase was performed every day and an evident increase of the tested parameters was found. These results prove in vivo activation of granulocytes by GM-CSF.

[Phagocytosis and oxidative metabolism of granulocytes in mice after stem cell factor (SCF) injection]. / Fagocytoza i metabolizm tlenowy granulocytów myszy poddanych dzialaniu czynnika wzrostowego komórek pnia (SCF).

Stem Cell Factor (SCF) can stimulate the growth and development of primitive multipotential and unipotential hematopoietic stem cells, either alone or in combination with other cytokines such as Granulocyte-Macrophage Colony Stimulating factor (GM-CSF) and Granulocyte-CSF (G-CSF). It was found that these cytokines can also stimulate the function of granulocytes but there is no information concerning SCF influence on the function of these mature cells. SCF was injected into mice subcutaneously during 5 consecutive days in a dose of 1 microgram/kg/d. An examination of the percentage of phagocytic granulocytes and NBT test was performed. The activity of acid phosphatase (AcP), alkaline phosphatase (AP), peroxidase (MPO) and esterase were determined by cytochemistry methods. On the basis of obtained results we can conclude that SCF evidently increases all tested parameters connected with the metabolism of phagocytosis.

[Phagocytosis and killing of Staphylococcus aureus by mouse granulocytes after interleukin-3 (IL-3) injection]. / Fagocytoza i zabijanie komórek Staphylococcus aureus przez granulocyty myszy poddanych dzialaniu interleukiny-3 (IL-3).

Interleukin-3 is a multipotential hematopoietic growth factor, which like other colony stimulating factors (CSFs) is effective "in vitro" stimulation of the mature cells function. It was found that IL-3 synergistically with GM-CSF and G-CSF stimulated the proliferation of the granulocytes. Therefore the purpose of this investigation was the evaluation "in vivo" of the influence of IL-3 on the phagocytosis, bactericidal activity, and enzyme activities of granulocytes. IL-3 was injected into mice subcutaneously during 5 days in dose 1 microgram/kg/d. The examination of the percent of cells phagocytizing bacteria (Staphylococcus aureus), NBT test and bactericidal activity, were performed every day and evident increase of the tested parameters was found. Additionally the enzyme activities in primary granules were measured and showed on increase of acid phosphatase and peroxidase activity.

[Pentoxifylline and verapamil influence on the phagocytosis of peritoneal macrophages from women with endometriosis]. / Wplyw pentoksyfiliny i werapamilu na aktywnosc fagocytarna makrofagów plynu otrzewnowego u kobiet z endometrioza.

OBJECTIVES: Disturbances in topical immunological response during endometriosis, cell and humoral, is of particular interest nowadays. Increase in the number and activity of peritoneal macrophages depends to a large extent on calcium concentration. Pentoxifylline and verapamil are both known for their ability to regulate calcium homeostasis. DESIGN: Evaluation of the pentoxifilline and verapamil influence on the phagocytic activity of the peritoneal macrophages from women with endometriosis. MATERIALS AND METHODS: Incubation of the peritoneal fluid with latex in four sets: 1) control with medium for 30 minutes; 2) with pentoxyfilline (3.8 mmol/l) for 30 minutes; 3) with verapamil 0.4 mmol/1; 4) with pentoxyfilline and verapamil together. Percentage of phagocyting cells, phagocytosis index and a score of phagocyting activity was calculated after 30 minutes incubation. Verapamil showed to increase all parameters of phagocytosis in comparison with control. CONCLUSIONS: Verapamil is effective reagent in activity of peritoneal fluid macrophages. Further studies on larger group of patients are necessary to see if the changes are statistically relevant.

[Interleukin 3 (IL-3) in diagnosis and monitoring of non-small-cell lung cancer]. / Interleukina 3 (IL-3) w diagnostyce i monitorowaniu niedrobnokomórkowego raka pluca.

Several circulating tumour markers for non-small-cell lung cancer (NSCLC) have been identified. Recent studies have focused on a new family of markers--hematopoietic growth factors. Some clinical investigations have shown cell surface receptors for interleukin 3 (IL-3) in lung cancer cells and autologous production of IL-3 in various human cell lines derived from NSCLC. The purpose of this investigation was to compare serum levels of IL-3 in non-small-cell lung cancer with a control group, to assess pre- and post treatment levels of IL-3 in relation to levels of commonly accepted tumour markers such as carcino-embryonic antigen (CEA) and cytokeratin fragment 19 (CYFRA 21-1), and to define the diagnostic sensitivity and specificity of IL-3 in NSCLC. In this study, the serum level of tumour markers was measured in 34 patients with NSCLC and in 20 healthy subjects. Serum samples were drawn before surgery and 10, 30, 90, 180 and 270 days after surgery. IL-3 and CEA were assayed using ELISA system and CYFRA 21-1 was measured by radioimmunoassay (RIA). Preoperative level of IL-3 was significantly increased in cancer patients relative to the control group. Concentrations of tumour markers were decreased after surgery and then increased (IL-3 and CEA) during chemio- or radiotherapy. The diagnostic sensitivity of IL-3 was 44% and the diagnostic specificity--85%. This investigation is one of the first studies assessing serum levels of IL-3 in the cancer patients. These results suggest that IL-3 may be useful in diagnostics of NSCLC, but this subject needs further studies.

Changes in some properties of the aurochs heart pyruvate dehydrogenase complex induced by its partial phosphorylation.

Comparative studies on the properties of the dephosphorylated and partially phosphorylated (to 35% activity reduction) pyruvate dehydrogenase complex (PDC) from aurochs heart muscle have been made. Data have been obtained indicating that the partial phosphorylation of PDC abolishes the kinetic attributes of a positive cooperativity of the pyruvate binding sites (nH = 1.5) featuring at low substrate concentrations. In addition, the partially phosphorylated PDC is inactivated slower at 50 degrees C.

[Predacious nematode-destroying fungi]. / Drapiezne grzyby nicieniobójcze.

Gastrointestinal nematodes are considered a serious economic problem affecting the livestock industry around the world. Current methods of their control, relaying mainly on organic drugs, are not sustainable because parasites develop resistance to anthelmintic and bacause of increasing public concern about chemicals residues in livestock products and environment. Nematode-trapping fungi offer a very promissing, nonchemotherapeutic approach to nematode parasite control. Their potential in preventing nematodosis is well documented. In this paper we outline the present knowlege on mechanisms involved in trapping and killing nematodes by the predacious nematode-destroying fungi.

[Influence of free fatty acids on growth, sporulation and virulence of the parasitic fungus Conidiobolus coronatus]. / Wplyw wolnych kwasów tluszczowych na wzrost, sporulacje i wirulencje pasozytniczego grzyba Conidiobolus coronatus.

Five free fatty acids (FFA): C16:0, C18:0, C18:1, C18:2 and C18:3 were introduced into culture media in order to investigate differential development of pathogenic fungus Conidiobolus coronatus as a function of FFA concentration. All tested FFA showed fungistatic action inhibiting hyphae growth and sporulation. Fungal colonies grown in the presence of FFA showed decreased virulence.

Parenteral nutrition and preOp preparation in prevention of post-operative insulin resistance in gastrointestinal carcinoma.

PURPOSE: The aim was to compare preventive effect of total parenteral nutrition (TPN) and oral nutrition (preOp) on the perioperative insulin resistance prevention in surgical gastrointestinal cancer patients. MATERIAL/METHODS: The study was conducted in a group of 75 elective gastric and large intestine cancer patients. Patients were randomly divided into 3 study groups, 25 patients each: group I (NIL) - no preparations influencing tissue sensitivity to insulin, group II (TPN) - total parenteral nutrition in its preoperative stage and group III (TPN + preOp) parenteral nutrition and preOp in the preoperative phase. RESULTS: Immediately after the surgery, no statistically significant differences in insulin resistance level between groups were observed. During the first 6 postoperative hours, a statistically significant decrease of insulin resistance level in the TPN+ preOp group in comparison to others, was observed. During the first 24 postoperative hours, the NIL group was the only one to keep the insulin resistance level the same as in the preoperative phase. CONCLUSIONS: Application of TPN in the preoperative phase leads to shortening of perioperative insulin resistance time. Combining TPN with oral application of carbohydrate before surgical procedure is an effective and the best method in postoperative insulin resistance syndrome prevention.

The effect of granulocyte colony-stimulating factor (G-CSF) on the activity of granulocyte enzymes in children with cancer who developed neutropenia after chemotherapy.

PURPOSE: G-CSF is a cytokine that stimulates the proliferation and maturation of granulocyte precursor cells. The results of in vitro and in vivo investigations conducted on animal models revealed that this cytokine influences the functions of mature granulocytes increasing the activities of the granulocyte enzymes participating in phagocytosis. MATERIAL AND METHODS: The investigation was conducted on a group of 26 children (age: 1.5-17 years) with cancer who developed neutropenia after chemotherapy and were treated with G-CSF. The control group included 29 healthy children (age: 5-17 years). The heparinized blood samples were taken before the injection of the stimulator (time 0) and after the 2nd and 5th injection of G-CSF (on day 3 and 6). Activities of granulocyte enzymes involved in the process of phagocytosis (myeloperoxidase, acid and alkaline phosphatase and esterase) in blood smears were evaluated. RESULTS: It has been found that G-CSF affects the activity of granulocyte enzymes by the normalization of decreased values of myeloperoxidase, acid phosphate and increasing the normal values of alkaline phosphate activity. The enzyme activities increased during the following days of treatment. CONCLUSION: Based on the obtained results, we can conclude that G-CSF activates the formation of fully competent granulocytes in cytostatic-treated children with various neoplastic diseases.

The effect of granulocyte colony stimulating factor on neutrophil functions in children with neutropenia after chemotherapy in the course of neoplasma.

PURPOSE: Granulocyte-colony stimulating factor stimulates proliferation and maturation of granulocyte precursor cells. The influence of this hematopoietic factor on phagocytic function of granulocytes was performed in in vitro experiments. The aim was to find, whether G-CSF applicated to children with neutropenia after chemotherapy influences phagocytic functions of neutrophils and whether evaluated parameters depend on a time of G-CSF injection? MATERIAL AND METHODS: The investigation was conducted on a group of 26 children with cancer, treated with granulocyte-colony stimulating factor in the cause of neutropenia after chemotherapy. The control group included 29 healthy children. The blood was taken before the stimulator injection and after 2 and 5 granulocyte-colony stimulating factor injections. The percentage of phagocyting cells and the phagocytic index of granulocytes were determined in heparinized whole blood samples. Oxygen metabolism was evaluated in the absence and presence endotoxin by nitroblue tetrazolium reduction. RESULTS: It was found that granulocyte-colony stimulating factor activates phagocytic functions of neutrophils by normalizing low values of phagocytic index and number of granulocytes, reducing dye nitroblue tetrazolium reduction and increasing the number of phagocytic cells. CONCLUSION: Based on obtained results we can conclude that granulocyte-colony stimulating factor apart from granulopoiesis stimulation can also increase phagocytic and oxidative capacity of granulocytes after chemotherapy.

An investigation of the kinase activity of aurochs heart pyruvate dehydrogenase complex.

The purified aurochs heart pyruvate dehydrogenase complex (PDC) saturated to approximation 60% with endogenous thiamine pyrophosphate (TPP), was slowly and incompletely inactivated by its kinase in the presence of ATP. Exogenous TPP or ADP, but not pyruvate, strongly inhibited the kinase activity. The kinetic properties of the aurochs heart PDC kinase suggested the occurrence of two active sites each with different affinities for ATP (K'm - 1.7 microM, K''m = 48 microM).

[Granulocyte colony stimulating factor (G-CSF) in diagnosis and monitoring of non-small-cell lung cancer (NSCLC)]. / Czynnik wzrostu kolonii granulocytarnych (G-CSF) w diagnostyce i monitorowaniu niedrobnokomórkowego raka pluca (NSCLS).

Granulocyte-colony stimulating factor (G-CSF) is one of the glycoproteins called colony-stimulating factors (CSFs). It has been shown that the target of the actions of CSFs are not limited to hematopoietic cells but can also affect the proliferation of nonhematopoietic cells. Some clinical investigations have shown the presence of cell surface receptors for G-CSF in lung cancer cells and autologous production of G-CSF in various human cell lines derived from non-small-cell lung cancer (NSCLC). The purpose of this investigation was to compare serum levels of G-CSF in NSCLC patients to a control group, to assess pre- and post treatment levels of G-CSF in relation to levels of commonly accepted tumour markers such as carcinoembryonic antigen (CEA) and cytokeratin fragment 19 (CYFRA 21-1), and to define the sensitivity of G-CSF in NSCLC. In this study, the serum levels of tumour markers were measured in 34 patients with NSCLC and in 20 healthy subjects. Serum samples were drawn before surgery and 10, 30, 90, 180 and 270 days after surgery. G-CSF and CEA were assayed using ELISA system and CYFRA 21-1 was measured by radioimmunoassay (RIA). Preoperative level of G-CSF was significantly increased in cancer patients relative to the control group. Concentrations of G-CSF and CYFRA 21-1 were decreased on the 10th day, but CEA on the 30th day after operation. The diagnostic sensitivity of G-CSF was 66%, CEA--62% and CYFRA 21-1--51%. Combined use of two markers increased the sensitivity in comparison to the use of G-CSF only. These results suggest that G-CSF may be useful in diagnostic and monitoring of NSCLC, but they need further studies.

[Role of hematopoietic factors in regulation of proliferation and function of phagocytes]. / Rola czynników hematopoetycznych w regulacji rozwoju oraz funkcji fagocytów.

Hematopoietic growth factors belong to the family of glycoproteins responsible for regulation, differentiation and proliferation of myelopoietic cells. In this paper the role of these factors in regulation of the function of phagocytic cells (granulocyte, monocyte, macrophage) is discussed on the basis of literature data.

Different extent of inhibition of pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase both containing endogenous thiamine pyrophosphate, by some anticoenzyme analogues.

Inhibitory effects of 4'-oxythiamine pyrophosphate (OTPP) and tetrahydrothiamine pyrophosphate (ThTPP) on the purified bison heart pyruvate dehydrogenase complex (PDC) semisaturated with endogenous thiamine pyrophosphate (TPP) and 2-oxoglutarate dehydrogenase complex (OGDC) saturated about 85% with endogenous TPP, were studied. It has been established that the thiamine derivatives strongly inhibit not only the PDC apoenzyme moiety, but also the PDC holoenzyme moiety. The apparent I50 values for the holoenzyme were 0.006 microM and 0.046 microM for OTPP and ThTPP, respectively. The inhibition of the PDC is reversible. After removal of the anticoenzyme analogues by gel filtration the endogenous TPP within the PDC is retained as it is evidenced by complete recovery of the enzyme activity without added TPP. In contrast with the PDC, OGDC holoenzyme form is weakly inhibited by the anticoenzyme analogues.

[Stem cell factor (SCF) in diagnosis and monitoring of non-small-cell lung cancer]. / Czynnik wzrostu komórek krwiotwórczych (SCF) w diagnostyce i monitorowaniu niedrobnokomórkowego raka pluca.

Lung cancer, of which non-small-cell lung cancer (NSCLC) constitutes about 80%, is the greatest cause of cancer-related death worldwide. Serum tumour markers may be helpful in early diagnosis, in the initial assessment of the progress of the disease and in monitoring of the tumour growth or tumour volume reduction. Recent studies have focused on a new family of markers--hematopoietic growth factors. Some clinical investigations have shown autologous production of stem cell factor (SCF) in various human cell lines derived from lung cancer and the expression of SCF mRNA in these lines. In this study, the serum level of SCF was measured using a sensitive sandwich ELISA system in 34 patients with non-small-cell lung cancer before and 10, 30, 90, 180 and 270 days after operation. Additionally common accepted tumour markers such as CEA and CYFRA 21.1 were also assayed. Preoperative level of SCF was increased in cancer patients in comparison to the normal sera. Concentrations of SCF and CYFRA 21.1 were decreased on 10th day, but CEA on 30th day after surgical treatment, although upon comparison of pre- and postoperative tumour markers serum levels significant difference was observed for SCF and CYFRA 21.1 (p < 0.05). Levels of SCF were increased in 79%, CEA in 62% and CYFRA 21.1 in 51%. The diagnostic sensitivity of SCF were related to the stage of the disease and the combined use of two markers increased the sensitivity compared with the use of only one. These results suggest that SCF may be useful in the diagnostic and monitoring of patients with NSCLC.

[Granulocyte macrophage-colony stimulating factor (GM-CSF) in diagnosis and monitoring of non-small cell lung cancer]. / Czynnik wzrostu kolonii granulocytarno-makrofagowych (GM-CSF) w diagnostyce i monitorowaniu niedrobnokomórkowego raka pluca.

Serum tumor markers may be helpful in early diagnosis of cancer, in the initial assessment of the extent of the disease, and in monitoring the tumor growth or tumor volume reduction once cancer has been diagnosed and treatment started. Recent studies have focused on a new family of markers -hematopoietic growth factors, especially on granulocyte-macrophage colony-stimulating factor (GM-CSF). A number of investigations have shown autologous production of GM-CSF in various human cell lines derived from melanoma, gastric or ovarian cancer, and in certain tumors of nonhematopoietic origin. In this study serum level of GM-CSF was measured using a sensitive sandwich ELISA system in 34 patients with non-small cell lung cancer (NSCLC) before and 10, 30, 90, 180 and 270 days after surgical operation. Additionally common accepted tumor markers such as CEA and CYFRA 21.1 were also assayed. Preoperative level of GM-CSF was significantly increased in cancer patients relative to the normal sera (p < 0.02). Concentration of GM-CSF and CYFRA 21.1 were decreased on 10th day, but CEA on 30th day after surgical treatment, although upon comparison of pre- and postoperative tumor markers serum levels significant difference was observed for CYFRA 21.1 (p < 0.05). Levels of GM-CSF were increased in 85%, CEA in 62% and CYFRA 21.1 in 51%. The diagnostic sensitivity and serum levels of GM-CSF were related to the stage of the disease and the combined use of two markers increased the sensitivity compared with the use of only one. These results suggest that GM-CSF, especially in the combination with CYFRA 21.1., may be useful in the diagnostic and monitoring of patients with NSCLC.