Interaction of an opportunistic fungus Purpureocillium lilacinum with human macrophages and dendritic cells

IntroductionPurpureocillium lilacinum is emerging as a causal agent of hyalohyphomycosis that is refractory to antifungal drugs; however, the pathogenic mechanisms underlying P. lilacinum infection are not understood. In this study, we investigated the interaction of P. lilacinum conidia with human macrophages and dendritic cells in vitro.MethodsSpores of a P. lilacinum clinical isolate were obtained by chill-heat shock. Mononuclear cells were isolated from eight healthy individuals. Monocytes were separated by cold aggregation and differentiated into macrophages by incubation for 7 to 10 days at 37°C or into dendritic cells by the addition of the cytokines human granulocyte-macrophage colony stimulating factor and interleukin-4. Conidial suspension was added to the human cells at 1:1, 2:1, and 5:1 (conidia:cells) ratios for 1h, 6h, and 24h, and the infection was evaluated by Giemsa staining and light microscopy.ResultsAfter 1h interaction, P. lilacinum conidia were internalized by human cells and after 6h contact, some conidia became inflated. After 24h interaction, the conidia produced germ tubes and hyphae, leading to the disruption of macrophage and dendritic cell membranes. The infection rate analyzed after 6h incubation of P. lilacinumconidia with cells at 2:1 and 1:1 ratios was 76.5% and 25.5%, respectively, for macrophages and 54.3% and 19.5%, respectively, for cultured dendritic cells.ConclusionsP. lilacinum conidia are capable of infecting and destroying both macrophages and dendritic cells, clearly demonstrating the ability of this pathogenic fungus to invade human phagocytic cells.

Activity of natural killer cells during HIV-1 infection in Brazilian patients

Natural killer cells are increasingly being considered an important component of innate resistance to viruses, but their role in HIV infection is controversial. Some investigators have found that natural killer cells do not confer a protective effect during the progression of HIV disease, whereas others have shown that natural killer cells may be protective and retard the progression of the disease, either through their lytic activity or by a chemokine-related suppression of HIV replication. In this study, we analyzed functional alterations in the activity of natural killer cells during HIV-1 infection using a natural killer cells activity assay with K562 cells as targets. RESULTS: Our results show that the activity of natural killer cells decreases only in the advanced phase of HIV infection and when high (40:1) effector cell-target cell ratios were used. The depression at this stage of the disease may be related to increased levels of some viral factors, such as gp120 or gag, that interfere with the binding capacity of natural killer cells, or to the decreased production of natural killer cells -activity-stimulating cytokines, such as IFN-a and IL-12, by monocytes, a subset of cells that are also affected in the late stage of HIV infection. The data suggest that decreased natural killer cells cell activity may contribute to the severe impairment of the immune system of patients in the late stages of HIV infection.

An experimental model for the transplantation of fetal central nervous system cells to the injured spinal cord in rats

INTRODUCTION: Traumatic spinal cord injury is one of the most disabling conditions occurring in man and thus stimulates a strong interest in its histopathological, biochemical, and functional changes, primarily as we search for preventive and therapeutic methods. PURPOSE: To develop an experimental model for transplantation of cells from the fetal rat central nervous system to the site of an injured spinal cord of an adult rat in which the transplanted cells survive and become integrated. This experimental model will facilitate investigations of factors that promote regeneration and functional recovery after spinal cord trauma. MATERIAL AND METHODS: Fifteen adult Wistar rats underwent laminectomy, and an spinal cord lesion was made with microdissection. Fetal spinal cord tissue was then transplanted to the site of the injury. The rats were monitored over a 48-hour period, and then their vertebral column was completely removed for histological analysis. RESULTS: In 60% of transplanted rats, the fetal tissue at the injured site remained viable in the site of the lesion.

C cells in the thyroid of pinealectomized rats

PURPOSE: To study quantitatively C cells in the thyroids of non-isogenic rats to determine the possible effects of pinealectomy on the number of these cells, and consequently on the synthesis and secretion of calcitonin. METHODS: Twenty male rats of an outbred strain (200-300 g) were used in the present study. One group of 10 animals was pinealectomized 50 days prior to sacrifice. Thyroid tissue was stained for calcitonin (Dako Corporation) at a 1:1500 dilution. The number of C cells observed was expressed as number of cells/cm². Data were analyzed statistically by Mann-Whitney test. RESULTS: The number of C cells in pinealectomized and normal animals ranged from 489 to 2084 per cm² and 227 to 1584 per cm², respectively, a difference that was statistically significant (P <0.05). CONCLUSIONS: These results showed consistent differences in the number of C cells after pinealectomy when compared to controls. We believe that pinealectomy increases the number of C cells in the rat thyroid.

C-cells in colloid goiter

PURPOSE: The aim of this investigation was to quantitatively evaluate C-cells in colloid goiters, analyzing 36 thyroids that were obtained through thyroidectomy from 24 patients with goiter and 12 normal glands from adult patients without thyroid disease, which were used as the control group. MATERIAL AND METHODS: On average, 6 different thyroid areas were sampled and labeled by immunohistochemistry with a monoclonal anticalcitonin antibody, utilizing the avidin-biotin-peroxidase complex. C-cells were counted in fields measuring 1 square centimeter, and the mean number of cells per field was then calculated. Data were statistically analyzed using the Mann-Whitney test. RESULTS: In the colloid goiter group, the number of C-cells ranged from 0 to 23 per field, while in normal controls they ranged from 20 to 148 per field. CONCLUSIONS: These results demonstrate a significant decrease of C-cell number in the colloid goiter group compared with control group, indicating that the hyperplastic process is restricted to follicular cells, to the detriment of C-cells, which probably cease to receive trophic stimuli.

Cytotoxic evaluation of essential oil from Casearia sylvestris Sw on human cancer cells and erythrocytes

A Casearia sylvestris (Flacourtiaceae) é uma planta popularmente conhecida como "guaçatonga" e é usada por povos indígenas da América do sul (Brasil, Peru e Bolivia) no tratamento de muitas doenças, incluindo câncer. Estudos citotóxicos mostraram que esta planta apresenta um possível e interessante potencial antitumoral devido à presença de moléculas chamadas casearinas. Além disso, a composição do óleo essencial mostrou uma alta concentração de sesquiterpenos de alto potencial citotóxico. Neste trabalho, nós verificamos que o óleo essencial da C. sylvestris apresentou uma boa citotoxicidade seletiva contra as linhagens de células tumorais HeLa, A-549 and HT-29 (CD50 63,3, 60,7 e 90,6 µg.ml-1, respectivamente) quando comparada às células não-tumorais Vero (CD50 210,1 µg.ml-1) e macrófagos de camundongos (CD50 234,0 µg.ml-1). Além disso, o óleo causou hemólise em sete diferentes tipos de eritrócitos, indicando que a C. sylvestris precisa ser usada com cuidado. Também foram testados padrões de β-cariofileno e α-humuleno que mostraram citotoxicidade similar àquelas apresentadas pelo óleo, indicando que estes compostos podem ser os responsáveis pelos efeitos tóxicos que foram observados neste estudo.

Puberty in male collared peccary (Pecari tajacu) determined by quantitative analysis of spermatogenic cells

Biological studies are necessary for the management of wildlife in captivity, and knowledge of reproduction is one of the important features for increasing production. The objective of the research was to determine the age at which male collared peccaries reach puberty. Testicular samples of 15 animals, aged 7 to 16 months, distributed into five groups (G1, G2, G3, G4 and G5) were used. The testes showed considerably increased weight, length and width (p < 0.05) from G1 to G3, whereas, from this group onward, the development of this organ was slower. There was positive correlation (p < 0.001) between the following testicular parameters: weight and length (r = 0.97), weight and width (r = 0.88), length and width (r = 0.92). Regarding the diameter of seminiferous tubules, an increase was observed (p < 0.05) from G1 to G4. The total number of spermatogenic cells increased significantly (p < 0.05) until G3 and then it stabilized. There was also positive correlation between testis weight and tubular diameter (r = 0.99, p < 0.001), and testis weight and spermatogenic cells (r = 0.98, p < 0.001). The number of Sertoli cells decreased significantly (p < 0.05) from G1, when they were undifferentiated as support cells, to G5, when they occurred together with the complete line of spermatic cells. The results demonstrate that the reproductive development of peccaries can be classified into the following stages: impuberty (G1, 7-8 months); pre-pubertal (G2, 9-10 months); pubertal (G3, 11-12 months); post-pubertal 1 (G4, 13-14 months); and post-pubertal 2 (G5, 15-16 months). Based on the histological analyses, puberty in the male collared peccary was determined to occur between 11 and 12 months of age.

Autologous Transplantation of Bone Marrow Adult Stem Cells for the Treatment of Idiopathic Dilated Cardiomyopathy

Background:Morbimortality in patients with dilated idiopathic cardiomyopathy is high, even under optimal medical treatment. Autologous infusion of bone marrow adult stem cells has shown promising preliminary results in these patients.Objective:Determine the effectiveness of autologous transplantation of bone marrow adult stem cells on systolic and diastolic left ventricular function, and on the degree of mitral regurgitation in patients with dilated idiopathic cardiomyopathy in functional classes NYHA II and III.Methods:We administered 4,54 x 108 ± 0,89 x 108 bone marrow adult stem cells into the coronary arteries of 24 patients with dilated idiopathic cardiomyopathy in functional classes NYHA II and III. Changes in functional class, systolic and diastolic left ventricular function and degree of mitral regurgitation were assessed after 3 months, 6 months and 1 year.Results:During follow-up, six patients (25%) improved functional class and eight (33.3%) kept stable. Left ventricular ejection fraction improved 8.9%, 9.7% e 13.6%, after 3, 6 and 12 months (p = 0.024; 0.017 and 0.018), respectively. There were no significant changes neither in diastolic left ventricular function nor in mitral regurgitation degree. A combined cardiac resynchronization and implantable cardioversion defibrillation was implanted in two patients (8.3%). Four patients (16.6%) had sudden death and four patients died due to terminal cardiac failure. Average survival of these eight patients was 2.6 years.Conclusion:Intracoronary infusion of bone marrow adult stem cells was associated with an improvement or stabilization of functional class and an improvement in left ventricular ejection fraction, suggesting the efficacy of this intervention. There were no significant changes neither in left ventricular diastolic function nor in the degree of mitral regurgitation.

Role of microRNAs 221/222 on Statin Induced Nitric Oxide Release in Human Endothelial Cells

Background: Nitric oxide (NO) has been largely associated with cardiovascular protection through improvement of endothelial function. Recently, new evidence about modulation of NO release by microRNAs (miRs) has been reported, which could be involved with statin-dependent pleiotropic effects, including anti-inflammatory properties related to vascular endothelium function. Objective: To evaluate the effects of cholesterol-lowering drugs including the inhibitors of cholesterol synthesis, atorvastatin and simvastatin, and the inhibitor of cholesterol absorption ezetimibe on NO release, NOS3 mRNA expression and miRs potentially involved in NO bioavailability. Methods: Human umbilical vein endothelial cells (HUVEC) were exposed to atorvastatin, simvastatin or ezetimibe (0 to 5.0 μM). Cells were submitted to total RNA extraction and relative quantification of NOS3 mRNA and miRs -221, -222 and -1303 by qPCR. NO release was measured in supernatants by ozone-chemiluminescence. Results: Both statins increased NO levels and NOS3 mRNA expression but no influence was observed for ezetimibe treatment. Atorvastatin, simvastatin and ezetimibe down-regulated the expression of miR-221, whereas miR-222 was reduced only after the atorvastatin treatment. The magnitude of the reduction of miR-221 and miR-222 after treatment with statins correlated with the increment in NOS3 mRNA levels. No influence was observed on the miR-1303 expression after treatments. Conclusion: NO release in endothelial cells is increased by statins but not by the inhibitor of cholesterol absorption, ezetimibe. Our results provide new evidence about the participation of regulatory miRs 221/222 on NO release induction mediated by statins. Although ezetimibe did not modulate NO levels, the down-regulation of miR-221 could involve potential effects on endothelial function.

Comportamento do Heterocromossômio em alguns Ortópteros do Brasil

In the present paper the behavior of the heterochromoso-mes in the course of the meiotic divisions of the spermatocytes in 15 species of Orthoptera belonging to 6 different families was studied. The species treated and their respective chromosome numbers were: Phaneropteridae: Anaulacomera sp. - 1 - 2n = 30 + X, n +15+ X and 15. Anaulacomera sp. - 2 - 2n - 30 + X, n = 15+ X and 15. Stilpnochlora marginella - 2n = 30 + X, n = 15= X and 15. Scudderia sp. - 2n = 30 + X, n = 15+ X and 15. Posldippus citrifolius - 2n = 24 + X, n = 12+X and 12. Acrididae: Osmilia violacea - 2n = 22+X, n = 11 + X and 11. Tropinotus discoideus - 2n = 22+ X, n = 11 + X and 11. Leptysma dorsalis - 2n = 22 + X, n = 11-J-X and 11. Orphulella punctata - 2n = 22-f X, n = 11 + X and 11. Conocephalidae: Conocephalus sp. - 2n = 32 + X, n = 16 + X and 16. Proscopiidae: Cephalocoema zilkari - 2n = 16 + X, n = 8+ X and 8. Tetanorhynchus mendesi - 2n = 16 + X, n = 8+X and 8. Gryliidae: Gryllus assimilis - 2n = 28 + X, n = 14+X and 14. Gryllodes sp. - 2n = 20 + X, n = 10- + and 10. Phalangopsitidae: Endecous cavernicola - 2n = 18 +X, n = 94-X and 9. It was pointed out by the present writer that in the Orthoptera similarly to what he observed in the Hemiptera the heterochromosome in the heterocinetic division shows in the same individual indifferently precession, synchronism or succession. This lack of specificity is therefore pointed here as constituting the rule and not the exception as formerly beleaved by the students of this problem, since it occurs in all the species referred to in the present paper and probably also m those hitherto investigated. The variability in the behavior of the heterochromosome which can have any position with regard to the autosomes even in the same follicle is attributed to the fact that being rather a stationary body it retains in anaphase the place it had in metaphase. When this place is in the equator of the cell the heterochromosome will be left behind as soon as anaphase begins (succession). When, on the contrary, laying out of this plane as generally happens (precession) it will sooner be reached (synchronism) or passed by the autosomes (succession). Due to the less kinetic activity of the heterochromosome it does not orient itself at metaphase remaining where it stands with the kinetochore looking indifferently to any direction. At the end of anaphase and sometimes earlier the heterochromosome begins to show mitotic activities revealed by the division of its body. Then, responding to the influence of the nearer pole it moves to it being enclosed with the autosomes in the nucleus formed there. The position of the heterochromosome in the cell is explained in the following manner: It is well known that the heterochromosome of the Orthoptera is always at the periphery of the nucleus, just beneath the nuclear membrane. This position may be any in regard of the axis of the dividing cell, so that if one of the poles of the spindle comes to coincide with it, the heterochromosome will appear at this pole in the metaphasic figures. If, on the other hand, the angle formed by the axis of the spindle with the ray reaching the heterochromosome increases the latter will appear in planes farther and farther apart from the nearer pole until it finishes by being in the equatorial plane. In this way it is not difficult to understand precession, synchronism or succession. In the species in which the heterochromosome is very large as it generally happens in the Phaneropteridae, the positions corresponding to precession are much more frequent. This is due to the fact that the probabilities for the heterochromosome taking an intermediary position between the equator and the poles at the time the spindle is set up are much greater than otherwise. Moreover, standing always outside the spindle area it searches for a place exactly where this area is larger, that is, in the vicinity of the poles. If it comes to enter the spindle area, what has very little probability, it would be, in virtue of its size, propelled toward the pole by the nearing anaphasic plate. The cases of succession are justly those in which the heterochromosome taking a position parallelly to the spindle axis it can adjust its large body also in the equator or in its proximity. In the species provided with small heterochromosome (Gryllidae, Conocephalidae, Acrididae) succession is found much more frequently because here as in the Hemiptera (PIZA 1945) the heterochromosome can equally take equatorial or subequatorial positions, and, furthermore, when in the spindle area it does offer no sereous obstacle to the passage of the autosomes. The position of the heterochromosome at the periphery of the nucleus at different stages may be as I suppose, at least in part a question of density. The less colourability and the surface irregularities characteristic of this element may well correspond to a less degree of condensation which may influence passive movements. In one of the species studied here (Anaulacomera sp.- 1) included in the Phaneropteridae it was observed that the plasmosome is left motionless in the spindle as the autosomes move toward the poles. It passes to one of the secondary spermatocytes being not included in its nucleus. In the second division it again passes to one of the cells being cast off when the spermatid is being transformed into spermatozoon. Thus it is regularly found among the tails of the spermatozoa in different stages of development. In the opinion of the present writer, at least in some cases, corpuscles described as Golgi body's remanents are nothing more than discarded plasmosomes.

Ensaio sobre o controle de "damping-off' do cafeeiro

Testes de 3 fungicidas-Vapam, PCNB e Cupravit - foram conduzidos em 2 tipos de substrato, visando controlar "damping-off" do cafeeiro. Os substratos se comportaram de maneira diferente quanto ao número de plantas sadias obtidas. Os fungicidas agiram de maneira diferente em diferentes substratos. No substrato A os fungicidas diferiram na sua ação protetora enquanto que no solo B, não. No resultado finai, alguns tratamentos se mostraram superiores que outros.

Cytochemical characterization of gill and hepatopancreatic cells of the crab Ucides cordatus (Crustacea, Brachyura) validated by cell metal transport

Ucides cordatus (Linnaeus, 1763) is a hypo-hyper-regulating mangrove crab possessing gills for respiratory and osmoregulatory processes, separated in anterior and posterior sections. They also have hepatopancreas, which is responsible for digestion and absorption of nutrients and detoxification of toxic metals. Each of these organs has specific cells that are important for in vitro studies in cell biology, ion and toxic metals transport. In order to study and characterize cells from gills and hepatopancreas, both were separated using a Sucrose Gradient (SG) from 10 to 40% and cells in each gradient were characterized using the vital mitochondrial dye DASPEI (2-(4-dimethylaminostyryl)-N- ethylpyridinium iodide) and Trichrome Mallory's stain. Both in 20 and 40% SG for gill cells and 30% SG for hepatopancreatic cells, a greater number of cells were colored with DASPEI, indicating a larger number of mitochondria in these cells. It is concluded that the gill cells present in 20% and 40% SG are Thin cells, responsible for respiratory processes and Ionocytes responsible for ion transport, respectively. For hepatopancreatic cells, the 30% SG is composed of Fibrillar cells that possess larger number of membrane ion and nutrient transporters. Moreover, the transport of toxic metal cadmium (Cd) by isolated hepatopancreatic cells was performed as a way of following cell physiological integrity after cell separation and to study differences in transport among the cells. All hepatopancreatic cells were able to transport Cd. These findings are the first step for further work on isolated cells of these important exchange epithelia of crabs, using a simple separation method and to further develop successful in vitro cell culture in crabs.

Cytochemical study of Rhodnius neglectus and Rhodnius prolixus salivary gland cells (Hemiptera, Triatominae)

Triatomines are hematophagous bugs of medical interest in South and Central America, where they may act as invertebrate hosts of the hemoflagellate protozoa Trypanosoma cruzi (the causative of Chagas’ disease) and Trypanosoma rangeli (Tejera, 1920). Triatomines of Rhodnius genus have salivary gland formed by two close and independent units: the principal and the accessory. This gland secretes saliva that abounds in substances that facilitate and permit feeding. Despite this importance, there are few reports on its cytochemistry. In purpose of amplifying this understanding, in this work it was investigated the nuclear structures (chromatin and nucleolar corpuscles) of salivary gland cells of Rhodnius neglectus (Lent, 1954) and Rhodnius prolixus (Stål, 1859). The salivary glands were removed from adult insects, fixed and submitted to different cytochemical methods: lacto-acetic orcein, silver ion impregnation, Feulgen reaction, Toluidine Blue, Variant method of critical electrolyte concentration and C-banding. The results evidenced predominance of binucleated cells, with bulky and polyploid nucleus, decondensed chromatin and a large nucleolar area. In addition, cytoplasmic metachromasy and a clear association between nucleolar and heterochromatic corpuscles were observed. Such characteristics were associated with intense synthesis activity to produce saliva. Besides, the heterochromatic corpuscles observed with C Banding permitted the differentiation of sexes and species.

A study of the endocervical columnar cells I - morphology and frequency

After the observation of many thousands of histological sections of the endocervical mucosa it became evident that its columnar cells present a great variety of aspects not only those of the surface of the canal but also those of the glands. A classification of these cells was made taking into account the staining affinity, the intensity staining of the cytoplasm, the presence or absence of cilia, the shape and location of the nucleus. The various combinations of these different data made possible the characterization of 26 types of cells which we labelled by the alphabetical letters. Two hundred and fifty cervices obtained by cervical amputation and by hysterectomy were studied. The uteri presented lesions in the course of routine laboratory examination. In each of the 250 histological sections there were specifically counted 2,000 columnar cells which cover the cervical canal and 2,000columnar cells which form the glands. A graphic representation of the frequency of both the superficial and glandular columnar cells was presented; this was given the name EPITHELIOGRAM. The variation of the cellular "composition" of each epithelium is discussed and the frequency of the various cellular types after the count of one million of cells is presented.

A study of the endocervical columnar cells II - effects of physiological changes and of oathological lesions of the uterus on the specific numerical composition of the endocervical epithelium

In the second part of this paper we nalysed the correlation between the clinical pathological alterations and the sum of the types of columnar cells of 300 histological sections of cervix. Fifty histological sections of normal cervix of sexually mature women were selected and considered as normal in pattern. The specific counts of the columnar cells which line the endocervical mucosa and those of the glands of 50 normal cervices were compared with other similar counts made in 50 histological sections of cervices of old women and emphasized the differences. Comparisons were made also between 50 normal cervices and 50 sections of cervices with chronic inflammation, 50 cervices with epidermoid metaplasia and 50 cervices with myoma of the corpus. Counts were made from 50 cervices of patients who on the occasion of the surgical operation were in the proliferative phase of the menstrual cycle; these were compared with the counts of 50 cervices of uteri in the luteal phase. Finally, the numerical frequency of the following data encountered in the 300 cervices was recorded: 1. aspects of the ectocervical epithelium; 2. number of Nabothian cysts; 3. number of cervical glands; 5. number of deliveries and 6. aspect of the material within the cervical canal.