Oxidant/antioxidant status in lambs and sheep with liver and lung cystic echinococcosis diagnosed by ultrasonography and necropsy.

The aim of this study was to evaluate total antioxidant capacity (TAC), total oxidant status (TOS), and oxidative stress index (OSI) in sheep and lambs with cyctic eccinocoocosis (CE) diagnosed by ultrasonography and necropsy findings. A total of 9 sheep and 17 lambs with CE were used in this study and the findings were compared to those of 6 healthy control sheep. Ultrasonography were used for the diagnosis of CE in sheep and lambs, and necropsy was performed to check the presence of cysts in liver and lungs. Serum TOS and TAC were measured by a novel colorimetric method. The TOS-to-TAC ratios were also calculated as OSI values. Serum biochemical profiles were determined by conventional measurement methods as well. The mean values for TOS, TAC and OSI were significantly (p<0.001) lower in sheep and lambs with CE when compared with those of the control sheep, and they were also significantly lower in lambs with CE in comparison to the mean values obtained in sheep with CE. The levels of serum albumin, total cholesterol, creatinine, and triglycerides in lambs with CE were found out to decrease significantly (p<0.001) when compared with those of both sheep with EC and the control group. There were no significant differences between the groups in terms of other serum parameters. In addition, when clinically and some biochemical values were evaluated, CE was found to be more severe in lambs than in sheep. It was concluded that although common diagnostic cyst detection is performed by postmortem examination, ultrasonography could successfully be used in conjunction with serum biochemical profile detection and serum TOS, TAC and OSI measurements for diagnosis of cysts in liver and lungs of severely infected living sheep and lambs. Serum albumin, total cholesterol, creatinine, total protein and triglycerides might be used as indicators in sheep and particularly in lambs for the diagnosis of CE.

[Transformation of the fungus Paecilomyces variotii and the causes of host cell lysis at the boundary with a fungal mycelium-containing Echinococcus capsule].

Experiments were carried out on animal species. The experiments used 30-day chicks, 80 rats, and 70 rabbits. Three hundred and twenty-nine patients with echinococcus complicated by paecilomycosis were meticulously examined. The fungi of the genus Paecilomyces undergo two transformation directions: the saprotrophic mycelial form of the fungus Paecilomyces variotii transforms to the tissue parasitic one as a globular form of spherules that transforms to the mycelial form in larval Echinococcus infection because the cyst capsule is a favorable environment for growth of fungal mycelia. The growth and aggressiveness of larval Echinococcus in the human lung are associated with the fact that fungal mycelial fibrous tunic contains Paecilomyces that have been first used to isolate active hyaluronidase that lyses host cells. Pulmonary echinococcosis complicated by the tissue form of paecilomycosis can be complicated by the mycelial form of the fungus of the genus Paecilomyces, by afflicting the nails and skin of patients, which requires particular treatment after surgery for hydatid disease. The chicks that had been brooded in an incubator and grown under special conditions to rule out fungal infection were first contaminated with the fungal mycelium labeled with methionine, sodium sulfate, sodium phosphate, or iodine. Each chick received 0.5 g of the labeled fungal mycelium. Regardless of the contamination mode, all the chicks from 3 groups were infected with Paecilomyces; the spherules exhibited labeled isotopes. Thus, it has been first conclusively proven that the diagnosis of paecilomycosis based on the blood detection of fungal globular spherules is valid and easy-to-use in any health care facility.

Accumulation of F-18 FDG in the infected pulmonary cyst in a patient with hydatid disease.

Echinococcosis is endemic in certain parts of the world, including Turkey. When the pulmonary cyst has characteristic features, it can be easily diagnosed, but when the appearance changes as a result of complications, the cyst may resemble a malignant lesion. We presented a complicated pulmonary hydatid disease patient who was referred to our department for PET-CT, after the detection of 2 lung lesions.

Long-term follow-up of metabolic activity in human alveolar echinococcosis using FDG-PET.

AIM: [(18)F]fluoro-deoxyglucose positron-emission-tomography (FDG-PET) detects metabolic activity in alveolar echinococcosis (AE). The slow changes in metabolic and morphological characteristics require long-term follow-up of patients. This is the first study to evaluate metabolic activity over may years, hereby assessing the utility of FDG-PET for the evaluation of disease progression and response to treatment. PATIENTS, METHODS: 15 patients received a follow-up FDG-PET combined with computed tomography (integrated PET/CT) with a median of 6.5 years after the first PET in 1999. Number and location of enhanced metabolic activity in the area of AE lesions was determined. Quantification of intensity of metabolic activity was assessed by calculation of mean standardized uptake values. RESULTS: AE lesions in 11/15 patients had been metabolically inactive initially, but only two showed permanent inactivity over the course of 81 months. Interestingly, in two patients metabolic activity was newly detected after 80 and 82 months. Benzimidazole treatment was intermittently discontinued in seven cases. Persisting activity at FDG-PET demanded continued benzimidazole treatment in four patients. Neither treatment duration, lesional size, calcifications nor regressive changes correlated with metabolic activity. CONCLUSION: Treatment responses are heterogeneous and vary from progressive disease despite treatment to long-term inactive disease with discontinued treatment. Lack of metabolic activity indicates suppressed parasite activity and is not equivalent to parasite death. However, metabolic activity may remain suppressed for years, allowing for temporary treatment discontinuation. Relapses are reliably detected with PET and restarting benzimidazole treatment prevents parasite expansion.

Apoptosis as a possible mechanism of infertility in Echinococcus granulosus hydatid cysts.

Echinococcus granulosus is a parasitic cestode causing hydatidosis in intermediate hosts (human and herbivorous). Most symptoms of the disease occur by the pressure exerted on viscera by cysts that are formed upon ingestion of the parasite eggs excreted by definitive hosts (canines). Protoscoleces, the developmental form of the parasite infective to definitive hosts, are formed in the germinal nucleated layer of fertile hydatid cysts. For unknown reasons, some cysts are unable to produce protoscoleces (infertile hydatid cysts). In this study, analysis of DNA fragmentation using TUNEL and agarose gel electrophoresis showed higher levels of apoptosis in infertile cysts as compared to fertile cysts. Additionally, caspase 3 was detected both in fertile and infertile cysts; the activity of this enzyme was found to be higher in infertile cysts. We conclude that apoptosis may be involved in hydatid cyst infertility. This is the first report on the presence of programmed cell death in E. granulosus.

The level of the collagen cross-link pyridinoline reflects the improvement of cutaneous lesions in one case of skin alveolar echinococcosis.

Cutaneous parasitic lesions, associated with a dense fibrous reaction, markedly improved under albendazole treatment in one case of supraumbilical skin localization of alveolar echinococcosis. Since collagen cross-linking increases during fibrogenesis and contributes to the stability of fibrotic lesions, we monitored the level of the cross-links pyridinoline and pentosidine in skin lesions from this patient to determine if they would reflect the changes occurring during treatment. We looked at the deposition of cross-linked type I collagen by immunohistochemistry and also measured the serum concentrations of pentosidine and of a fragment of type I collagen (ICTP), which contains a site of pyridinoline formation. Albendazole treatment did not affect either the collagen content of skin lesions or the serum concentrations of ICTP and pentosidine, but it led to a pronounced decrease in pyridinoline level concomitant with the disappearance, observed by immunohistochemistry, of extensively cross-linked fibrotic type I collagen. The follow-up of collagen cross-linking by pyridinoline in skin tissue thus appears to be useful in reflecting the improvement of fibrotic skin diseases during therapy.

Both murine SAA1 and SAA2 yield AA amyloid in alveolar hydatid cyst-infected mice.

Amyloid susceptible C57BL/6 and partially amyloid resistant A/J mice, infected intraperitoneally with 250 alveolar hydatid cyst (AHC), the larval stage of a cestode parasite Echinococcus multilocularis, develop multiple organ amyloid deposits at approximately 1 and 4 weeks post infection (p.i.), respectively. Pooled spleens and livers from each mouse strain, at 8 and 10 weeks p.i., were used for the purification of protein AA utilizing a HiLoad Superdex 200 column equilibrated with 5 M guanidine-HCl. Protein AA from each mouse strain was separated on 16% Tris-tricine SDS-PAGE gels and immunoblotted with monospecific rabbit anti-mouse AA IgG; five and six immunoreactive AA subspecies were detected in the C57BL/6 and A/J materials, respectively. N-Terminal amino acid sequence analysis was performed on the bulk column-purified protein AA as well as on the electroblotted AA subspecies from each mouse strain. The results show a mixture of serum amyloid A1 (SAA1) and (SAA2)-derived AA protein from each mouse strain; SAA1-derived AA, although alluded to, has never been demonstrated as tissue deposits in mice. These findings suggest that the intense and persistent inflammatory processes in AHC-infected mice may have induced conversion of weakly amyloidogenic SAA1 to AA. This conversion could be detected by amino acid sequencing of electrophoretically separated AA subspecies.

Pathogenesis of secondary amyloidosis in an alveolar hydatid cyst-mouse model: histopathology and immuno/enzyme-histochemical analysis of splenic marginal zone cells during amyloidogenesis.

C57BL/6 mice infected with 10, 50 or 250 alveolar hydatid cysts (AHC) were used to study the pathogenesis of secondary amyloidosis. Immuno and enzyme-histochemical analyses on spleen sections were performed to investigate the temporal relationship between AHC antigen, serum amyloid A protein (SAA) and amyloid (AA) deposition and concomitant qualitative and quantitative changes in the concentration of lysosomal acid phosphatase (AP) and nonspecific esterase (NSE) in splenic marginal zone (MZ) and red pulp reticuloendothelial (RE) cells prior to and during amyloidogenesis. AA-induction period was reduced from 5 weeks in the 50 cyst group to 6 or 7 days in the 250-cyst group; the 10-cyst group mice remained negative for splenic AA. Splenic RE cell hyperplasia, deposition of AHC antigen and SAA and peak AP and NSE activities occurred in splenocytes prior to AA deposition. AA-deposition in the MZs coincided with reduced RE cell AP and NSE activities and degenerative changes in the MZs. AA-induction period in the 250-cyst group was further reduced from 7 days to 4 days after intravenous injection of silica which is cytotoxic to RE cells. We suggest that defective clearance of SAA from tissue sites as a result of progressive reduction in lysosomal enzymes coupled with degenerative changes in splenic MZ monocytoid cells trigger amyloidogenesis in the extracellular matrix.

Albendazole treatment of echinococcosis in humans: effects on microsomal metabolism and drug tolerance.

We prospectively studied the effect of albendazole on microsomal reserve and on first-pass activation to albendazole sulfoxide in patients with hydatid disease. An aminopyrine breath test was performed in 12 patients while they were receiving albendazole treatment and while they were not. Excretion of 14CO2 in breath averaged 0.70%.kg.mmol-1 +/- 0.20%.kg.mmol-1 without treatment and 0.54%.kg.mmol-1 +/- 0.14%.kg.mmol-1 with treatment (p less than 0.005). Plasma levels of albendazole sulfoxide were measured 4 hours after the morning dose during the first and second half of the 4-week treatment cycles. In nine of the 12 patients albendazole sulfoxide levels decreased during the second half of the cycle by an average of 0.84 +/- 0.76 mumol/L (p less than 0.02). Transaminase levels increased in 10 of the 12 patients during long-term albendazole treatment, and major side effects, including hepatotoxicity, neutropenia, and alopecia, were observed in three patients. We conclude that albendazole partially inhibits microsomal enzyme function but induces its own metabolism. Hepatotoxicity and other possible severe side effects necessitate close therapeutic monitoring of patients who are given albendazole.

Lipids in hydatid fluid collected from lungs and livers of sheep and man.

Hydatid fluid collected from the lungs and livers of sheep and humans was analysed for protein and lipid composition. There were no marked differences in the composition of these parameters and the major lipids were triglycerides and diglycerides. The phospholipids, which formed the minor fraction, were mainly phosphatidyl choline and phosphatidyl inositol. Cholesterol present was in the free form.

Induction of amyloid enhancing factor and its biological properties in murine alveolar hydatidosis.

The biological activity and time of appearance of alveolar hydatid cyst induced splenic amyloid enhancing factor (AEF) with respect to amyloid deposition in the spleens were determined in C57BL/6J mice. Mice were infected intraperitoneally with 100 alveolar hydatid cysts (AHC) and killed bi-weekly between 2 and 14 weeks postinfection (p.i.). AHCs and spleens were excised, weighed and a portion of each spleen was sectioned and stained for quantitation of amyloid deposits and histological studies. The remaining spleen pieces were sonicated separately in cold phosphate buffered saline, pH 7.4 (I g/10 ml), centrifuged (27,000 g, 60 min, 4 degrees C) and the supernatant tested for AEF activity. Splenomegaly followed the progressive increase in the AHC biomass and AEF activity coincided with the appearance of amyloid deposits at 6 weeks p.i. A 2.5 mg intraperitoneal protein dosage of AEF in conjunction with a subcutaneous injection of 0.5 ml of a 0.11 M AgNO3 solution in mice, induced the maximum amount of splenic amyloid deposition at 48 h; the amount of splenic amyloid deposits decreased by either increasing or decreasing the AEF dosage. In vivo, 70% of the AEF activity was abolished by day 4 post-injection of AEF and completely by 3 weeks. These findings indicate that AHC-induced AEF is functionally analogous to casein-induced AEF and its appearance in the spleen coincides with neutrophilia, histiocytosis and amyloid deposition.

Alveolar hydatid cyst induced amyloid enhancing factor (AEF): physicochemical properties and abolition of AEF activity by serine protease inhibitors.

Cell free supernatants prepared from amyloidotic liver, unfractionated and fractionated peritoneal and spleen cells from casein stimulated (16 h post-injection) or alveolar hydatid cyst-infected (7 or 12 weeks post-infection, p. i.) C57BL/6J mice were used to assess amyloid enhancing factor (AEF) activity and to determine its cell-source and physicochemical properties. Of the various supernatants used, the plastic adherent spleen cell lysate (95% macrophages) from 7 weeks p.i mice showed greater AEF activity, on a cell to cell basis, than the lysates prepared from whole spleen cells, peritoneal exudate cells or nonadherent (96% lymphocytes) spleen cells. Culture supernatants obtained from Con A, LPS or the parasite antigen stimulated amyloidotic spleen cells but not the normal mouse spleen cells contained AEF activity; the supernatant from unstimulated amyloidotic spleen cells was negative for AEF activity. AEF was precipitated with 25% and 50% saturation with (NH4)2SO4 and after gel-filtration the low molecular weight fraction contained the AEF activity which on SDS-PAGE resolved into three peptides ranging between mol. wts 15,000 and 31,000. Of the various specific and nonspecific protease inhibitors tested, AEF activity was completely abolished by 30 min preincubation with 10 mM phenylmethylsulphonyl fluoride. Taken together, these results indicate that AEF may be a small molecular weight lysosomal neutral protease of neutrophil/macrophage origin.

Lipids in the laminated layer of liver, lung and daughter hydatid cysts of equine Echinococcus granulosus (Cestoda).

Lipids extracted from the laminated layers of horse liver and lung hydatids, including a daughter liver cyst, were analysed using TLC. No differences in lipid composition was detected in 11 liver cysts, whether from the same or different livers, and di- and triacylglycerols, cholesterol, wax and steryl esters, oleic acid, sphingomyelin, phosphatidyl choline, phosphatidyl inositol and ceramide hexosides were detected. The daughter cyst differed from its "parent" cyst in lacking diacylglycerols and wax and steryl esters. The lung cyst differed from the liver cysts in that cholesterol, wax and steryl esters and diacylglycerols were not detected.

The use of equilibrium density-gradient ultracentrifugation in the isolation and characterisation of glycoproteins with blood group P1 activity from sheep hydatid-cyst fluid.

Equilibrium density-gradient ultracentrifugation in caesium choride and caesium sulphate has been used in the isolation and fractionation of the glycoproteins specific for blood-group P1 from hydatid cyst fluids. The fractions obtained have distinct and systematic differences specifically related to their buoyant densities, chemical compositions and specific-activities for group P1. High levels of specific-activity were maintained over a large range of chemical compositions. The peptide content varied systematically from 2.5% for the densest fraction to 37% for the least dense fraction. The amino acid composition was essentially constant over all fractions. The proportion of glucosamine decreased and the proportions of galactosamine, mannose and glucose increased with increasing peptide content of the fractions. The data presented suggest the present of oligosaccharide side-chains of various lengths and compositions and/or the presence of oligosaccharide side-chains with very different chemical compositions, of which only some are associated with the specificity for group P1. The properties of the glycoproteins from hydatid cyst fluids have been compared with those of the glycoproteins from human ovarian cysts. Although some similarities have been demonstrated there are significant differences.

Similarities of human hydatid cyst fluid components and the host serum.

Nine lung hydatid cyst fluid of Echinococcus granulosus species from man were analysed by electrophoresis, immunoelectrophoresis and by biochemical tests. In addition respective sera of the host were analysed for comparison. Analysis revealed striking similarities in cellulose acetate and agarose gel electro- and immunoelectrophoretic patterns of proteins from hydatid cyst fluid and the serum of the respective hosts. It is presumed that serum proteins (albumin and globulins) occur in hydatid cyst fluid, but in smaller amount than in serum, an we believe that the host proteins can penetrate the membranes of the hydatid cyst.