The natural killer and fungicidal activity of the peripheral blood mononuclear cells in candidiasis

The activity of natural killers and the capacity of peripheral blood mononuclears for the extracellular killing of Candida cells in patients with different forms of candidiasis has been determined in the radiometric test with the use of targets labeled with 3H-uridine RNAase and in the fungicidal test. On the basis of the data thus obtained the conclusion has been made that in candidiasis changes in the activity of natural killers and fungicidal capacity depend on the severity of the disease and the stage of the process. Linear correlation between the activity of natural killers and fungicidal capacity in candidiasis patients at the stage of exacerbation and in healthy persons is absent.     

Immune system function in candidiasis patients

To compare the clinical picture and the immunological characteristics, 58 candidiasis patients differing by the severity and dissemination of the disease were examined. Chronic candidiasis of the skin and mucous membranes, the most severe and disseminated form of the disease, is associated with a decrease in the number of T-lymphocytes and changes in their subpopulations, as well as high titers of Candida albicans antigen and antibodies to it in blood sera. The immune system of patients with visceral candidiasis and chronic vulvovaginal candidiasis was similar to that of healthy persons in the characteristics under study. Immediate and mixed hypersensitivity occurred in candidiasis patients more frequently than in healthy persons. In extremely severe forms of Candida infection immediate hypersensitivity prevailed.     

Specific circulating immune complexes and antibody titers in candidiasis]

Titers of antibodies to Candida albicans and specific circulating immune complexes have been studied in patients with chronic candidiasis of the genitals, skin and mucous membranes. In most patients with candidiasis of the skin and mucous membranes specific immune complexes have been detected in the blood serum; their determination increases the diagnostic value of serological tests in this disease.     

Modifications to the surface of T-lymphocytes in chronic candidiasis of the skin and mucous membranes

27 patients with chronic candidiasis of the skin and mucous membranes have been examined with a view to study the expression of E-receptors and its changes after the incubation of T-lymphocytes with theophylline, levamisole, thymaline and Candida albicans antigen. This study has revealed that, in comparison with healthy persons, most patients show decreased expression of E-receptors, and the character of their changes after incubation with the above-mentioned preparations is essentially different. The deviations revealed in this study have been regarded as the modification of T-cell membranes, which is seemingly one of the reasons of the weak competence of T-cells. In its turn, the expression of E-receptors may be considered as an indicator of the influence of different factors on T-lymphocytes. Substances altering the level of cyclic nucleotides in the cell, the antigenic fractions of the fungal causative agent, the suppression or blocking the action of thymic hormone-like substances may serve as such factors.     

Morphological, ultrastructural, cytochemical and functional analysis of neutrophils deficient in myeloperoxidase

Eighteen patients with established hereditary myeloperoxidase deficiency underwent morphological, ultrastructural, cytochemical and functional analysis in order to correlate the lack of peroxidase from phagocytes with other leucocyte activities. Cytochemical and ultrastructural findings only confirmed the peroxidase defect in neutrophil and monocyte population, whereas normal peroxidase activity was detected in eosinophil granulocytes ("Alius-Grignaschi anomaly"). Morphological analysis, as determined by both ligh and electron microscopy, showed in two patients with total MPO-deficiency a large number of neutrophils (50-60%) with nuclear abnormalities very similar to Pelger-Huet's heterozygous form (two lobed neutrophils having a typical pince-nez appearance and a nuclear chromatin coarser than that of normal PMNL). Other 2 cases displayed a 50-60% five-lobed neutrophils, as occur in congenital nuclear hypersegmentation of PMNL. These findings suggest that Alius-Grignaschi anomaly and Pelger-Huet syndrome can be found associated in the same individuals, since both these abnormalities have a genetic origin. Finally, since an impaired bactericidal and fungicidal activity was observed, no patients displayed particular susceptibility to persistent or severe infections, thus confirming the presence of MPO-independent enzymatic systems.     

The role of neutrophil myeloperoxidase in the development of inflammation induced by thermal skin burns

Luminol-dependent chemiluminescence (CL) of blood neutrophils stimulated with phorbol-12- myristate-13-acetate (PMA) and myeloperoxidase (MPO) activity of neutrophils and plasma have been investigated in children (n = 16) during the early period (1?7 days) after thermal skin burns exceeding 20% of total body surface. The CL level of stimulated neutrophils was higher in burn patients than in healthy children of the reference group (p lt; 0.01). Increased neutrophil MPO activity was found in 40% of patients, while increased plasma MPO activity was detected in 57% of patients. The albumin fraction isolated from plasma of burn patients increased the PMA-stimulated CL response of blood from healthy donors. These results suggest that the acute inflammatory response to the thermal burn causes neutrophil activation and MPO release into plasma. MPO-mediated modification of plasma proteins, particularly albumin, may stimulate neutrophil activation and provoke further inflammatory response of the body to the thermal injury.     

Substantial alterations of the cutaneous bacterial biota in psoriatic lesions

For psoriasis, an idiopathic inflammatory disorder of the skin, the microbial biota has not been defined using cultivation-independent methods. We used broad-range 16S rDNA PCR for archaea and bacteria to examine the microbiota of normal and psoriatic skin. From 6 patients, 19 cutaneous samples (13 from diseased skin and 6 from normal skin) were obtained. Extracted DNA was subjected to the broad range PCR, and 1,925 cloned products were compared with 2,038 products previously reported from healthy persons. Using 98% sequence identity as a species boundary, 1,841 (95.6%) clones were similar to known bacterial 16S rDNA, representing 6 phyla, 86 genera, or 189 species-level operational taxonomic unit (SLOTU); 84 (4.4%) clones with <98% identity probably represented novel species. The most abundant and diverse phylum populating the psoriatic lesions was Firmicutes (46.2%), significantly (P<0.001) overrepresented, compared to the samples from uninvolved skin of the patients (39.0%) and healthy persons (24.4%). In contrast, Actinobacteria, the most prevalent and diverse phylum in normal skin samples from both healthy persons (47.6%) and the patients (47.8%), was significantly (P<0.01) underrepresented in the psoriatic lesion samples (37.3%). Representation of Propionibacterium species were lower in the psoriatic lesions (2.9+/-5.5%) than from normal persons (21.1+/-18.2%; P<0.001), whereas normal skin from the psoriatic patients showed intermediate levels (12.3+/-21.6%). We conclude that psoriasis is associated with substantial alteration in the composition and representation of the cutaneous bacterial biota.     

The effect of cytostatics and corticosteroids on the phagocytic and fungicidal activity of the neutrophilic granulocytes

In vitro tests have revealed that cytostatic agents and corticosteroids, introduced in vivo, produce an inhibitory effect on the phagocytic activity of mouse neutrophil granulocytes and on their capacity for the destruction of Candida albicans blastospores. The action of the above-mentioned preparations may be an important pathogenetic factor which when introduced into the animals, contributes to the development of candidiasis.     

The interaction of neutrophilic granulocytes with Candida albicans fungi in the formation of mycotic foci under conditions of immunodepression

In the electron-microscopic study of the interaction of neutrophil granulocytes with the fungal species C. albicans in the process of the formation of mycotic foci in mice under the conditions of cyclophosphamide-induced immunosuppression, mouse leukocytes have been found to retain their capacity for migration to the focus of inflammation and for the phagocytosis of fungal cells. At the same time the fungicidal activity of leukocytes is decreased, which is manifested by the prevalence of viable fungal cells with the partially digested cell wall in the cytoplasm of leukocytes.     

Acquired neutrophil myeloperoxidase deficiency: an indicator of subclinical activation of blood coagulation?

Using an automated cytochemical analyzer used for routine differential counts, we have been able to demonstrate acquired myeloperoxidase deficiency in 102 patients at our institution. Clinical and laboratory data on these patients showed a high incidence of diabetes mellitus (25.5%) and thrombotic diseases (24.5%), as well as a strikingly constant hyperfibrinogenemia (mean = 635 mg/100 ml; range = 360-1015 mg/100 ml). In 4 additional acute leukemia patients in complete remission, a close time correlation was noted between acquired MPO deficiency, diffuse intravascular coagulation and relapse. These findings indicate the importance of the relationships between neutrophil granulocytes and blood coagulation, and suggest that similar changes in neutrophil MPO activity may represent an early morphological indicator of subclinical activation of blood coagulation.     

Peroxydase activity of the granulocytes in the course of surgical treatment of neoplasm of the alimentary tract

Granulocyte peroxidase activity of patients with alimentary tract neoplasms has been investigated by using the histochemical method. Blood samples were collected before surgical treatment and anaesthesia, during surgical intervention from vessels draining the neoplasms, and 2-3 weeks after surgery from peripheral vessels. The activity of myeloperoxidase in the polymorphonuclear granulocytes of patients was evidently lower in comparison to the activity of this enzyme in the granulocytes of normal individuals. The excision of the tumour did not evidently increase the tested activity in blood from peripheral vein in comparison to the same sample and to the blood from vein draining of tumour. We can suggest that granulocytes may release myeloperoxidase to the surrounding media in the course of neoplasmatic diseases and in this case the cells are exhausted in this enzyme.     

Lymphocyctes Tgammadelta in clinically normal skin and peripheral blood of patients with systemic lupus erythematosus and their correlation with disease activity

Human Tgammadelta lymphocytes constitute from 1 to 15% of all peripheral blood lymphocytes. Recent work has demonstrated that this population plays a major role in the pathogenesis of infectious and immune diseases. Increased numbers of gammadelta T cells have been found in affected skin from systemic sclerosis and chronic cutaneous lupus erythematosus patients. In our study, we have determined the numbers of Tgammadelta lymphocytes and their subpopulations in peripheral blood from 29 patients with systemic lupus erythematosus (SLE) and in 19 healthy volunteers using flow cytometry and specific monoclonal antibodies. The same cells in uninvolved skin from SLE patients and human controls using immunohistochemical analysis were estimated. T-Cell receptor (TCR) delta chain gene rearrangement was identified with primers for Vdelta1, Vdelta2 and Vdelta3 by the polymerase chain reaction. Statistical analysis showed a significantly decreased number of gammadelta T cells in SLE patients (26.4+/-16.9/microl) compared with the control group (55.3+/-20.6/microl (p < 0.001). The number of Vdelta2 TCR+ and Vgamma9 TCR+ subpopulations was also lower in SLE patients than in healthy persons. No statistical correlation between disease activity and the number of gammadelta T cells was demonstrated. The percentage of Tgammadelta lymphocytes in clinically normal skin from SLE patients was twice (22.0+/-9.4%) that found in the skin from healthy persons (11.1+/-5.5%) (p < 0.002). Higher percentages of the Vdelta2 TCR+ and Vgamma9 TCR+ subpopulation of lymphocytes were found in the skin from SLE patients. We have also found positive correlation between the percentage of Tgammadelta lymphocytes in skin and the activity of SLE (r=0.594, p < 0.001), and between subpopulation Vdelta3 TCR+ and disease activity (r=0.659, p< 0.001). In conclusion, the results of our studies demonstrate that, in patients with SLE, accumulation of Tgammadelta lymphocytes can be seen in clinically normal skin, and the percentage of these cells correlates with the activity of the disease.     

Effect of proper-myl on humoral interactions between activated lymphocytes and neutrophils in patients with myeloproliferative diseases

The lymphokine synthesizing function of peripheral blood lymphocytes (PBL) was studied in 22 patients with chronic myeloid leukemia (CML), 28 patients with subleukemic myelosis (SLM) and 15 healthy persons. The index of the neutrophil stimulation (INS) in CML (1.73 +/- 0.068) and SLM (1.458 +/- 0.004) was statistically significantly lower than that in the healthy persons (2.6 +/- 0.07). The use of proper-myl in the complex therapy markedly increased the ability of PBL to produce the factor stimulating phagocytic activity of neutrophils (INS 1.94 +/- 0.04 and 1.832 +/- 0.092). On the basis of the findings it was recommended to use proper-myl for prevention and treatment of infectious complications.     

Optical isomers of a leukotriene B4 antagonist have differential effects on granulocyte diapedesis in the guinea pig dermis.

Leukotriene B4 (LTB4) is a proinflammatory product of arachidonic acid metabolism that has been implicated in a number of inflammatory diseases. When injected intradermally into the guinea pig, LTB4 has been shown to elicit a dose-dependent infiltration of granulocytes as assessed by the level of the neutrophil marker enzyme myeloperoxidase. SC-41930 [7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxy]-3,4-dihydro-8- propyl-2H-1-benzopyran-2-carboxylic acid] is a potent LTB4 receptor antagonist. When compounds were coadministered along with LTB4 (35 ng) into the dermal site, racemic SC-41930, (+)-SC-41930, and (-)-SC-41930 each inhibited granulocyte accumulation with ED50 values of 340 +/- 30, 98 +/- 5.7, and 1000 +/- 142 ng, respectively; when given intravenously inhibited with ED50 values of 0.5 +/- 0.06, 0.3 +/- 0.04, and 1.4 +/- 0.19 mg/kg, respectively; and when given intragastrically inhibited with ED50 values of 1.7 +/- 0.20, 1.4 +/- 0.23, and 3.0 +/- 0.41 mg/kg, respectively.     

Functional and morphological characteristics of the digestive reaction of gastric mucosa

Morphological state of connective tissue (stromal) cells of the stomach mucous membrane has been studied in healthy persons, having a habitual regime of feeding. During digestive period in the stomach mucous membrane, certain changes develop, which are considered as a digestive reaction. Three stages of the digestive reaction, having strict morphological signs are determined, their connections being stated by means of morphometry and mathematical analysis. I stage (preparatory) is characterized with a moderate vascular reaction, degranulation of mast cells under the superficial++ epithelium of the mucous membrane, with a moderate neutrophilic leukopedesis and a moderate lymphocytic infiltration; II stage (developed) is distinguished as a definitely demonstrated reaction of the microcirculatory bed, intensive degranulation of mast cells at all levels of the mucous membrane, massive discharge of neutrophilic granulocytes and lymphocytes into stroma; III stage (restorative) is characterized with a predominance of fibroblasts and fibrocytes, with reparation of mast cells, with decreasing saturation of stroma with neutrophilic granulocytes, lymphocytes, an increased number of eosinophilic granulocytes takes place. The data obtained widen our knowledge on functional morphology of the stomach mucous membrane, normal and at gastroduodenal pathology.     

The characteristics of local immunity in Chlamydia-associated chronic inflammatory diseases of the organs of the lesser pelvis in women

Local immune reactions in 107 women with Chlamydia-associated chronic endometritis and salpingo-oophoritis were studied on endometrial biopsy specimens. Of these women, group 1 consisted 45 patients with Chlamydia-associated chronic endometritis and salpingo-oophoritis and group 2 consisted of 62 such patients having, in addition, bacterial vaginosis and candidiasis. Among associated bacteria, epidermal staphylococci, Escherichia coli and their combinations prevailed. Pathomorphologically, in the biopsy specimens taken from the patients of group 1 a decrease in the activity of mononuclear phagocytes, secondary SIgA deficiency with the level of IgA-producing plasmocytes relatively unchanged, the moderate intensity of the synthesis of IgG and a shift in the ratio of T-helpers/T-suppressors towards the latter were observed. In the biopsy specimens of the uterine mucosa obtained from group 2 a decrease in the phagocytic activity of neutrophil granulocytes and macrophages, the pronounced suppression of IgA production by IgA plasmocytes with a sharp decrease in the amount of SIgA and an essential increase in the number of IgG-producing cells were noted. Disturbances of local immunity, observed in the presence of associated Chlamydia infection, gives grounds for the inclusion of immunomodulating preparations into the complex of therapeutic measures.     

Interaction of Candida albicans with Human Leukocytes and Serum

A quantitative assay of candidacidal activity based on differential staining of non-viable Candida albicans by methylene blue was developed and applied to studies of leukocytes from normal individuals and patients with fungal and other infections. Serum factors were necessary for optimal phagocytosis of C. albicans but lacked direct candidacidal activity. Normal human neutrophils (38 studies) killed 29.0 +/- 7.4% of ingested C. albicans in 1 hr. Eosinophils and monocytes killed a smaller percentage. Neutrophil candidacidal activity did not require protein or ribonucleic acid synthesis by the leukocyte but was inhibited by anaerobic conditions, potassium cyanide, and colchicine. Leukocytes of a patient with hereditary myeloperoxidase deficiency and of three children with chronic granulomatous disease phagocytized C. albicans normally, yet failed to kill them. Our data suggest that the neutrophil can play an important role in resistance to Candida infection and that the lysosomal enzyme myeloperoxidase and its oxidant substrate hydrogen peroxide are the major participants in neutrophil candidacidal activity.     

Myeloperoxidase-catalyzed 3-chlorotyrosine formation in dialysis patients.

Oxidative stress has been implicated in the cardiovascular complications that affect chronic renal failure patients on hemodialysis, though the physiologically relevant pathways mediating oxidative damage are poorly understood. It is known, however, that hemodialysis activates neutrophils, a well-characterized source of hydrogen peroxide and myeloperoxidase. The phagocyte-derived myeloperoxidase-hydrogen peroxide-chloride system generates hypochlorous acid, which reacts with tyrosine residues of proteins to form 3-chlorotyrosine. To explore the role of activated phagocytes in oxidative stress in chronic renal failure, we used 3-chlorotyrosine as a specific marker of myeloperoxidase activity. Utilizing isotope dilution gas chromatography-mass spectrometry, we compared 3-chlorotyrosine levels in plasma proteins of five patients on chronic hemodialysis therapy with those of age- and sex-matched healthy controls. The oxidized amino acid was present in the plasma proteins of 4 of the hemodialysis patients (3.5 +/- 0.8 micromol per mol tyrosine) but was undetectable in the healthy subjects. Therefore, one pathway for oxidative stress in hemodialysis patients appears to involve hypochlorous acid generated by the myeloperoxidase system of activated phagocytes. We also examined intradialytic 3-chlorotyrosine levels using membranes that activate white blood cells and the alternative pathway of complement. Hemodialysis increased plasma myeloperoxidase and the expression of CD11b/CD18 by circulating phagocytes, but failed to demonstrably increase 3-chlorotyrosine levels. 3-chlorotyrosine was detectable in 12 of 19 samples in total, with significant intrasubject variability. Our observations suggest that oxidants generated by myeloperoxidase contribute to the increased oxidative stress observed in renal-failure patients but do not damage plasma proteins during the hemodialysis procedure itself.     

Staphylococci in the microbiocenosis of the skin of the hands]

The skin autoflora on the pad of a forefinger and the back and palm of a hand was studied in 40 healthy males aged 18-60 years by the modified washing and scraping method of P. Williamson and A. Kligman. 638 cultures of aerobic microorganisms, including coccal (55.3%) and bacilliform (44.7%) microbes, were isolated. In 6 persons (15%) coagulase-positive staphylococci were detected. Out of 10 coagulase negative species of this genus, S. epidermidis, S. saprophyticus and S. warneri occurred most frequently on the skin of hands. The highest density of bacterial populations (10.970 +/- +/- 1.845 cells/sq. cm) was registered on the back of hands, the surface of palms was found to have somewhat lower density (8.679 +/- 1.282 cells/sq. sm) and the skin of forefingers, the lowest density of bacterial populations (6.878 +/- +/- 1.137 cells/sq. sm). 17.5% of examined persons were found to be carriers with S. aureus isolated from their nasal mucosa. S. aureus isolated from the skin surface and the nasal cavity of different persons belonged to different phage variants, but S. aureus isolated from the nasal cavity and the skin of the same person belonged to one phage variant.     

Comparison of the antibiotic sensitivity of lipophilic Corynebacterium sp. isolated from patients on the day of admission and during hospitalization

Lipophilic species of Corynebacterium are increasing problem in hospital infections. The aim of this study was to evaluate occurrence of these microorganisms in the materials taken from patients in the day of admission and during the hospitalization as well as comparison of their antibiotic sensitivity. The investigation included 65 strains isolated from hospitalized patients and 48 strains isolated from unchanged skin. Using Api Coryne test 5 species were identified. C. urealyticum dominated, the other were C. subsp. lipophilum and C. jeikeium. Among strains isolated from unchanged diseased skin the most C. jeikeium and C. accolens occurred. All strains were sensitive to glycopeptide, quinupristin/dalphopristin. The strains isolated from hospitalized patients were usually sensitive to fuside acid, doxycycline as well as tetracycline. Strains isolated from unchanged skin were sensitive to almost all tested antibiotics. In the group of 65 strains isolated from hospitalized patients 99.0% were multiresistant. In the group of strains isolated from unchanged skin only two strains were multiresistant. Differences in antibiotic sensitivity among analysed Corynebacterium sp. were confirmed. Majority of the "hospital strains" were characterized by multiresistance. Basing on these results it is possible to suppose, that multiresistance is main factor that favours lipophilic Corynebacterium species in the process of colonization of mucous membranes, skins as well as developing infections.