Plasma levels of tumor necrosis factor-alpha in patients with visceral leishmaniasis (Kala-Azar). Association with activity of the disease and clinical remisson following antimonial therapy

Evaluation of TNF-alpha in patients with Kala-azar has drawn increasing interest due to its regulatory role on the immune system, in addition to its cachetizing activity. The objective of this study was to examine the association between plasma levels of TNF-alpha, measured by immunore-activity (ELISA) and bioactivity (cytotoxicity assay with L-929 cells), and clinical manifestations of visceral leishmaniasis. Plasma samples from 19 patients with Kala-azar were obtained before, during and at the end of antimonial therapy. TNF-alpha determinations was done by using the cytotoxicity assay (all patients) and the enzyme-linked immunoassay (ELISA - 14 patients). A discrepancy between results obtained by ELISA and cytotoxicity assay was observed. Levels of circulating TNF-alpha, assessed by ELISA, were higher in patients than in healthy controls, and declined significantly with improvement in clinical and laboratory parameters. Plasma levels before treatment were 124.7 ± 93.3 pg/ml (mean ± SD) and were higher than at the end of therapy 13.9 ± 25.1 pg/ml (mean ± SD) (p = 0.001). In contrast, plasma levels of TNF-alpha evaluated by cytotoxicity assay did not follow a predicted course during follow-up. Lysis, in this case, might be not totally attributed to TNF-alpha. The discrepancy might be attributed to the presence of factor(s) known to influence the release and activity of TNF-alpha.

Serum levels of interleukin-6, tumor necrosis factor-alpha and interferon-gama in infants with and without dengue

This study compared the serum levels of IL-6, TNF-alpha and IFN-gamma, in children under 1 year of age with and without dengue. Sera were collected from a total of 41 children living in the Department of Antioquia, Colombia (27 patients with dengue and 14 controls). The results showed higher cytokine levels in children with dengue than without dengue, with statistically significant differences for IL-6 and IFN-gamma. No statistically significant differences were found between clinical forms, although IL-6 and IFN-gamma levels were higher in dengue fever cases than in dengue hemorrhagic fever cases. On the other hand, TNF-alpha levels were higher in dengue hemorrhagic fever than in dengue fever. The levels of IL-6 and TNF-alpha were higher in secondary infection than in primary infection, although IFN-gamma levels were higher in primary infection. These results suggest that IL-6, TNF-alpha and IFN-gamma are involved in dengue infection independently of the clinical form.

Detection of circulant tumor necrosis factor-alpha , soluble tumor necrosis factor p75 and interferon-gamma in Brazilian patients with dengue fever and dengue hemorrhagic fever

Pro-inflammatory cytokines are believed to play an important role in the pathogenesis of dengue infection. This study reports cytokine levels in a total of 54 patients examined in Recife, State of Pernambuco, Brazil. Five out of eight patients who had hemorrhagic manifestations presented tumor necrosis factor-alpha (TNF-alpha) levels in sera which were statistically higher than those recorded for controls. In contrast, only one out of 16 patients with mild manifestations had elevated TNF-alpha levels. The levels of interleukin-6 (IL), IL-1beta tested in 24 samples and IL-12 in 30 samples were not significantly increased. Interferon-g was present in 10 out of 30 patients with dengue. The data support the concept that the increased level of TNF-alpha is related to the severity of the disease. Soluble TNF receptor p75 was found in most patients but it is unlikely to be related to severity since it was found with an equivalent frequency and levels in 15 patients with dengue fever and another 15 with dengue hemorrhagic fever.

Increased plasma levels of tumor necrosis factor-alpha in asymptomatic/"indeterminate" and Chagas disease cardiomyopathy patients

We compared plasma tumor necrosis factor-alpha (TNF-alpha) levels among asymptomatic/"indeterminate" Chagas disease patients (ASY) and patients across the clinical spectrum of chronic Chagas disease cardiomyopathy (CCC). Idiopathic dilated cardiomyopathy (DCM) patients and normal controls (NC) were included as controls. ASY Chagas disease patients had significantly higher plasma TNF-alpha levels than NC. TNF-alpha levels among severe CCC patients with significant left ventricular (LV) dysfunction were similar to those of DCM patients, showing average 2-fold higher levels than CCC patients without LV dysfunction and ASY patients, and 8-fold higher levels than NC. In Chagas disease, chronic TNF-a production prior to heart failure may play a role in CCC progression.

Differential expression of notch signaling-related transcripts accompanies Pro-thymocyte proliferation and phenotype transition induced by epidermal growth factor plus insulin in fetal thymus organ cultures

Thymus regression upon stressing stimuli, such as infectious diseases, is followed by organ reconstitution, paralleling its development in ontogeny. A narrow window of thymus development was here studied, encompassing the pro-T lymphoid precursor expansion during specification stages, by the use of epidermal growth factor plus insulin (INS) in murine fetal thymus organ cultures. Aiming to disclose signaling pathways related to these stages, cultured thymus lobes had their RNA extracted, for the search of transcripts differentially expressed using RNAse protection assays and reverse transcriptase-polymerase chain reactions. We found no difference that could explain INS-driven thymocyte growth, in the pattern of transcripts for death/proliferation mediators, or for a series of growth factor receptors and transcriptional regulators known as essential for thymus development. Thymocyte suspensions from cultured lobes, stained for phenotype analysis by fluorescence activated cell sorting, showed a decreased staining for Notch1 protein at cell surfaces upon INS addition. We analyzed the expression of Notch-related elements, and observed the recruitment of a specific set of transcripts simultaneous and compatible with INS-driven thymocyte growth, namely, transcripts for Notch3, for its ligand Jagged2, and for Deltex1, a mediator of a poorly characterized alternative pathway downstream of the Notch receptor.

The balance between the production of tumor necrosis factor-alpha and interleukin-10 determines tissue injury and lethality during intestinal ischemia and reperfusion

A major goal in the treatment of acute ischemia of a vascular territory is to restore blood flow to normal values, i.e. to "reperfuse" the ischemic vascular bed. However, reperfusion of ischemic tissues is associated with local and systemic leukocyte activation and trafficking, endothelial barrier dysfunction in postcapillary venules, enhanced production of inflammatory mediators and great lethality. This phenomenon has been referred to as "reperfusion injury" and several studies demonstrated that injury is dependent on neutrophil recruitment. Furthermore, ischemia and reperfusion injury is associated with the coordinated activation of a series of cytokines and adhesion molecules. Among the mediators of the inflammatory cascade released, TNF-alpha appears to play an essential role for the reperfusion-associated injury. On the other hand, the release of IL-10 modulates pro-inflammatory cytokine production and reperfusion-associated tissue injury. IL-1beta, PAF and bradykinin are mediators involved in ischemia and reperfusion injury by regulating the balance between TNF-alpha and IL-10 production. Strategies that enhance IL-10 and/or prevent TNF-alpha concentration may be useful as therapeutic adjuvants in the treatment of the tissue injury that follows ischemia and reperfusion.

Epidermal growth factor receptors, testosterone levels and parotid gland changes in rats infected with Trypanosoma cruzi

It has been demonstrated that parotid glands of rats infected with Trypanosoma cruzi present severe histological alterations; changes include reduction in density and volume of the acini and duct systems and an increase in connective tissue. We evaluated the association between morphological changes in parotid glands, circulating testosterone levels and epidermal growth factor receptor (EGF-R) expression in experimental Chagas disease in rats. Animals at 18 days of infection (acute phase) showed a significant decrease in body weight, serum testosterone levels and EGF-R expression in the parotid gland compared with a control group. Since decreases in body weight could lead to a reduction in circulating testosterone concentration, we believe that the reduction in EGF-R expression in parotid glands of infected rats is due to alterations in testosterone levels and atrophy of parotid glands is caused by changes in EGF-R expression. Additionally, at 50 days (chronic phase) of infection parotid glands showed a normal histological aspect likely due to the normalization of the body weight. These findings suggest that the testosterone-EGF-R axis is involved in the histological changes.

Gap junction reduction in cardiomyocytes following transforming growth factor-β treatment and Trypanosoma cruzi infection

Gap junction connexin-43 (Cx43) molecules are responsible for electrical impulse conduction in the heart and are affected by transforming growth factor-β (TGF-β). This cytokine increases during Trypanosoma cruzi infection, modulating fibrosis and the parasite cell cycle. We studied Cx43 expression in cardiomyocytes exposed or not to TGF-β T. cruzi, or SB-431542, an inhibitor of TGF-β receptor type I (ALK-5). Cx43 expression was also examined in hearts with dilated cardiopathy from chronic Chagas disease patients, in which TGF-β signalling had been shown previously to be highly activated. We demonstrated that TGF-β treatment induced disorganised gap junctions in non-infected cardiomyocytes, leading to a punctate, diffuse and non-uniform Cx43 staining. A similar pattern was detected in T. cruzi-infected cardiomyocytes concomitant with high TGF-β secretion. Both results were reversed if the cells were incubated with SB-431542. Similar tests were performed using human chronic chagasic patients and we confirmed a down-regulation of Cx43 expression, an altered distribution of plaques in the heart and a significant reduction in the number and length of Cx43 plaques, which correlated negatively with cardiomegaly. We conclude that elevated TGF-β levels during T. cruzi infection promote heart fibrosis and disorganise gap junctions, possibly contributing to abnormal impulse conduction and arrhythmia that characterise severe cardiopathy in Chagas disease.

Vascular endothelial growth factor-A enhances indoleamine 2,3-dioxygenase expression by dendritic cells and subsequently impacts lymphocyte proliferation

Dendritic cells (DCs) are antigen (Ag)-presenting cells that activate and stimulate effective immune responses by T cells, but can also act as negative regulators of these responses and thus play important roles in immune regulation. Pro-angiogenic vascular endothelial growth factor (VEGF) has been shown to cause defective DC differentiation and maturation. Previous studies have demonstrated that the addition of VEGF to DC cultures renders these cells weak stimulators of Ag-specific T cells due to the inhibitory effects mediated by VEGF receptor 1 (VEGFR1) and/or VEGFR2 signalling. As the enzyme indoleamine 2,3-dioxygenase (IDO) is recognised as an important negative regulator of immune responses, this study aimed to investigate whether VEGF affects the expression of IDO by DCs and whether VEGF-matured DCs acquire a suppressor phenotype. Our results are the first to demonstrate that VEGF increases the expression and activity of IDO in DCs, which has a suppressive effect on Ag-specific and mitogen-stimulated lymphocyte proliferation. These mechanisms have broad implications for the study of immunological responses and tolerance under conditions as diverse as cancer, graft rejection and autoimmunity.

Low transformation growth factor-β1 production and collagen synthesis correlate with the lack of hepatic periportal fibrosis development in undernourished mice infected with Schistosoma mansoni

Undernourished mice infected (UI) submitted to low and long-lasting infections by Schistosoma mansoni are unable to develop the hepatic periportal fibrosis that is equivalent to Symmers’ fibrosis in humans. In this report, the effects of the host’s nutritional status on parasite (worm load, egg viability and maturation) and host (growth curves, biology, collagen synthesis and characteristics of the immunological response) were studied and these are considered as interdependent factors influencing the amount and distribution of fibrous tissue in hepatic periovular granulomas and portal spaces. The nutritional status of the host influenced the low body weight and low parasite burden detected in UI mice as well as the number, viability and maturation of released eggs. The reduced oviposition and increased number of degenerated or dead eggs were associated with low protein synthesis detected in deficient hosts, which likely induced the observed decrease in transformation growth factor (TGF)-β1 and liver collagen. Despite the reduced number of mature eggs in UI mice, the activation of TGF-β1 and hepatic stellate cells occurred regardless of the unviability of most miracidia, due to stimulation by fibrogenic proteins and eggshell glycoproteins. However, changes in the repair mechanisms influenced by the nutritional status in deficient animals may account for the decreased liver collagen detected in the present study.

Insulin-like growth factor and growth hormone receptor in postpartum lactating beef cows

The objective of this study was to evaluate the plasma concentrations of insulin-like growth factor-I (IGF-I), and the mRNA hepatic expression of IGF-I and of the growth hormone receptors GHR and GHR 1A, in postpartum beef cows. Four Angus and four crossbred (Angus x Nelore) postpartum suckled beef cows were used. Liver and blood samples were collected every 10 days, from calving to 40 days postpartum, for gene expression and for β-hydroxybutyrate and IGF-I assays, respectively. Samples for progesterone assay were collected every other day, from day 10 to 40 postpartum. Three cows ovulated before 40 days postpartum. IGF-I concentration was higher in Angus x Nelore than in Angus cows. There was no difference in the expression of GHR, GHR 1A and IGF-I according to breed or ovulatory status. IGF-I concentrations were higher in crossbred cows, but have not changed according to postpartum ovulatory status. Moreover, changes in postpartum IGF-I concentrations are not associated with changes in liver GHR, GHR 1A and IGF-I mRNA expression in either breed.

Immunohistochemical evaluation for P53 and VEGF (Vascular Endothelial Growth Factor) is not prognostic for long term survival in end stage esophageal adenocarcinoma

OBJECTIVES: To correlate the expression of p53 protein and VEGF with the prognosis of patients submitted to curative resection to treat esophageal adenocarcinoma. METHODS: Forty-six patients with esophageal adenocarcinoma, submitted to curative resection, were studied. The expressions of p53 protein and VEGF were assessed by immunohistochemistry in 52.2% and 47.8% of tumors, respectively. RESULTS: P53 protein and VEGF expressions coincided in 26% of the cases, and no correlation between these expressions was observed. None of the clinicopathological factors showed a significant correlation with p53 protein or VEGF expressions. There was no significant association between p53 protein and VEGF expressions and long-term survival. CONCLUSION: The expression of p53 protein and VEGF did not correlate with prognosis in esophageal adenocarcinoma patients submitted to curative resection.

Porcine follicular fluid concentration of free insulin-like growth factor-I collected from different diameter ovarian follicles

The study aimed to quantify the concentrations of free IGF-I in serum and fluid of ovarian follicles in pre-pubertal gilts and describe the ovarian morphology by measuring the size of the ovaries and counting the number of surface follicles. Ovaries (n=1,000) from pre-pubertal gilts were obtained immediately after slaughter. A total of 10 samplings were performed, with ovaries obtained from 50 females for each collection. The follicles situated on the surface of each ovary were classified as small (SFs, 2 to 5mm in diameter) or large (LFs 6 to 10mm in diameter) and the follicular fluid was obtained by follicle aspiration. The collection of serum samples was performed after the gilts exsanguination using sterile tubes. From the pool of serum and follicular fluid obtained from 50 females, the concentration of free IGF-I was determined in each sample using an enzyme immunoassay kit (ELISA). The description of ovarian morphometry was performed in 100 ovaries from randomly selected gilts. The larger and smaller lengths of ovaries were measured, and the total number of SFs and LFs present on the surface of each ovary were also counted. The IGF-I concentration was greater (P<0.05) in LFs (170.92±88.29 ng/mL) compared with SFs (67.39±49.90ng/mL) and serum (73.48±34.63ng/mL). The largest and smallest length of the ovaries was 26.0±3.0 and 19.0mm ±2.0mm, respectively. The number of SFs (70.86±25.76) was greater (P<0.01) than LFs (6.54±5.26). The study concluded that LFs present greater levels of IGF-I when compared with SFs and blood, which is related to increased activity of the LFs and its differentiation to ovulation. In addition, ovaries of pre-pubertal gilts have a higher number of SFs compared to LFs. Therefore, our study demonstrated unique data regarding the physiological concentration of free IGF-I in ovarian follicles, that can be used in future research to evaluate the addition of this hormone in the in vitro production media of porcine embryos with the goal to improve the technique efficiency.

Influence of Insulin-like Growth Factor I (IGF-I) on the survival and the in vitro development of caprine preantral follicles

The aim of this study was to investigate the effects of the insulin-like growth factor -I (IGF-I) on survival, activation (transition from primordial to primary follicles) and growth of caprine preantral follicles cultured in vitro. Fragments of ovarian cortex were cultured for one and seven days in the absence or presence of IGF-I (0, 50 and 100ng/ml). The non-cultured and cultured tissues were processed and analyzed by histology and transmission electron microscopy. The culture for one day in a medium with 100ng/ml of IGF-I showed 86.7% of morphologically normal follicles. These results were similar (P>0.05) to the percentage of normal follicles found in the control (96.7%). It was also found that this medium increased the percentage of follicular activation (developing follicles) with one day of culture. The oocyte and follicular diameters remained similar to the control by culturing for one day in a medium containing 100ng/ml of IGF-I. The ultrastructural analysis did not confirm the integrity of the follicular fragments in a medium containing IGF-I (100ng/ml) after one and seven days of culture. In conclusion, this study demonstrated that the addition of 100 ng/ml of IGF-I in the culture medium enables the development of preantral follicles of goats with one day of culture. However, it is not sufficient to maintain the follicular integrity and the follicular survival rate after seven days of culture.