Phytochemical screening of the dichloromethane–ethanolic extract of Eriosema campestre var. macrophylum roots and its antiproliferative effect on human peripheral blood lymphocytes

ABSTRACT Eriosema campestre var. macrophylum (Grear) Fortunato, Fabaceae, is a native plant of the Brazilian Cerrado and the decoction of its roots has been used by folk medicine for the therapy of inflammatory diseases. In this study we aimed to investigate the effect of the dichloromethane–ethanolic extract of E. campestre roots on the proliferative response of lymphocytes and to examine the profile of IL-2 production. The effect of dichloromethane–ethanolic extract of E. campestre on the proliferation of phytohemagglutinin-stimulated lymphocytes was evaluated by using flow cytometry and the cell supernatants were assayed for IL-2 concentrations by using an enzyme-linked immunosorbent assay. The phytochemical screening of E. campestre roots was performed to determine the main secondary metabolites through chromogenic and precipitation reactions and by using HPLC-PAD. In addition to the presence of subclasses of flavonoids (flavones and flavonols) in dichloromethane–ethanolic extract of E. campestre, we observed that the extract induced a concentration-dependent decrease in IL-2 levels on the supernatant of the cell cultures as well as an antiproliferative effect on T lymphocytes, including CD4+ and CD8+ cells. The anti-inflammatory effects attributed to E. campestre by folk medicine may partly be explained by its antiproliferative action on T lymphocytes.

The effect of insulin resistance and exercise on the percentage of CD16(+) monocyte subset in obese individuals.

UNLABELLED: Obesity is a low-grade chronic inflammation condition, and macrophages, and possibly monocytes, are involved in the pathological outcomes of obesity. Physical exercise is a low-cost strategy to prevent and treat obesity, probably because of its anti-inflammatory action. We evaluated the percentage of CD16(-) and CD16(+) monocyte subsets in obese insulin-resistant individuals and the effect of an exercise bout on the percentage of these cells. Twenty-seven volunteers were divided into three experimental groups: lean insulin sensitive, obese insulin sensitive and obese insulin resistant. Venous blood samples collected before and 1 h after an aerobic exercise session on a cycle ergometer were used for determination of monocyte subsets by flow cytometry. Insulin-resistant obese individuals have a higher percentage of CD16(+) monocytes (14.8 ± 2.4%) than the lean group (10.0 ± 1.3%). A positive correlation of the percentage of CD16(+) monocytes with body mass index and fasting plasma insulin levels was found. One bout of moderate exercise reduced the percentage of CD16(+) monocytes by 10% in all the groups evaluated. Also, the absolute monocyte count, as well as all other leukocyte populations, in lean and obese individuals, increased after exercise. This fact may partially account for the observed reduction in the percentage of CD16(+) cells in response to exercise. Insulin-resistant, but not insulin-sensitive obese individuals, have an increased percentage of CD16(+) monocytes that can be slightly modulated by a single bout of moderate aerobic exercise. These findings may be clinically relevant to the population studied, considering the involvement of CD16(+) monocytes in the pathophysiology of obesity. Copyright © 2016 John Wiley & Sons, Ltd. SIGNIFICANCE OF THE STUDY: Obesity is now considered to be an inflammatory condition associated with many pathological consequences, including insulin resistance. It is proposed that insulin resistance contributes to the aggravation of the inflammatory dysfunction in obesity. The effect of obesity on the percentage of monocytes was previously observed in class II and III obese individuals who presented other alterations in addition to insulin resistance. In this study we observed that insulin-resistant obese individuals, but not insulin-sensitive ones, had an increased percentage of CD14(+) CD16(+) monocytes. This fact shows that a dysfunction of the monocyte percentage in class I obese individuals is only seen when this condition is associated with insulin resistance.

Essential oil from Ageratum fastigiatum reduces expression of the pro-inflammatory cytokine tumor necrosis factor-alpha in peripheral blood leukocytes subjected to in vitro stimulation with phorbol myristate acetate

Abstract Ageratum fastigiatum (Gardner) R.M. King & H. Rob., a member of the Asteraceae family popularly known in Brazil as "matapasto", is indicated in folk medicine as anti-inflammatory and analgesic. Despite its popular use, little is known about its potential effect on the parameters involved in an inflammatory response. The objective of this study was to characterize the chemical composition of the essential oil from A. fastigiatum and to evaluate the frequency of tumor necrosis factor alpha and interferon gamma producing cells in peripheral blood lymphocytes stimulated with phorbol myristate acetate in the presence of essential oil from A. fastigiatum. Non-toxic concentrations of essential oil from A. fastigiatum were evaluated in cultures of peripheral blood leucocytes using the trypan blue exclusion assay by flow cytometry. GC–MS analysis revealed that the prevalent compounds identified in the essential oil from A. fastigiatum sample were α-pinene, limonene, trans-caryophyllene, α-humulene, caryophyllene oxide, 1,2-humulene-epoxide, 1,6-humulanodien-3-ol, and α-cadinol. Results showed that exposure to essential oil from A. fastigiatum at concentrations of 0.5 × 10−2 and 1 × 10−2 µl/ml caused no alterations in leukocyte viability as compared to the control group. Both concentrations lowered the percentage of tumor necrosis factor alpha (+)-lymphocytes and neutrophils. There were no changes in the percentage of lymphocytes positive for the interferon gamma cytokine. Our results suggest that part of the anti-inflammatory activity attributed to A. fastigiatum may be due to the effect of some of its components in decreasing the number of cells that produce the pro-inflammatory cytokine tumor necrosis factor alpha.

Effect of exercise on the plasma BDNF levels in elderly women with knee osteoarthritis.

Knee osteoarthritis is a common disease in the elderly population worldwide. The alleviation of the symptoms associated with this disease can be achieved with physical exercise that induces a cascade of molecular and cellular processes. Of the neurotrophins, brain-derived neurotrophic factor (BDNF) appears to be the most affected by physical activity. Moreover, BDNF seems to have a negative modulatory role in inflammation, and its production by skeletal muscle cells or by cells of the immune system drives the immunoprotective role of physical activity in situations of chronic inflammation. Therefore, the aim of this study was to evaluate plasma BDNF concentrations in elderly individuals presenting with knee osteoarthritis. To accomplish this, sixteen volunteers (mean age 67 ± 4.41 years) presenting with clinically and radiographically diagnosed knee osteoarthritis were evaluated during acute exercise (1 session of 20 min on a treadmill) and after chronic exercise (12 weeks of aerobic training, consisting of a 50-min walk 3 times per week). Additionally, both a functional assessment (during a 6-min walk) and a pain perception assessment were performed at the start and at the end of physical exercises (training). The plasma BDNF concentrations were measured by ELISA. For the population studied, acute exercise increased the levels of BDNF only before the 12-week training period (p < 0.001). Moreover, the training augmented the plasma concentrations of BDNF (p < 0.0001) and improved clinical parameters (functional p < 0.001; pain perception p < 0.01).

Increase in dopaminergic, but not serotoninergic, receptors in T-cells as a marker for schizophrenia severity.

Schizophrenia is characterized by a slow deteriorating mental illness. Although the pathophysiology mechanisms are not fully understood, different studies have suggested a role for the immune system in the pathogenesis of schizophrenia. To date, an altered expression or signaling of neurotransmitters receptors is observed in immune cells during psychiatric disorders. In the present study, we investigated the expression of different serotonin and dopamine receptors in T-cells of schizophrenic and control patients. We used flow cytometry to determine the pattern of expression of dopamine (D2 and D4) and serotonine receptors (SR1A, SR1C, SR2A, SR2B), as well as serotonin transporter (ST), in T-cell subsets (CD4 and CD8). Expression of serotonin receptors and ST in T-cells of schizophrenic patients were not different from controls. However, the percentages of CD4+D4+ and CD8+D4+ were increased in schizophrenic patients as compared to controls. In addition, increased percentages of CD8+D2+ cells were also observed in schizophrenic patients, albeit this population revealed lower CD4+D2+ cells in comparison to controls. Interestingly, a relationship between clinical symptoms and immunological parameters was also observed. We showed that the Brief Psychiatric Rating Scale (BPRS), the Positive and Negative Syndrome Scale (PANSS) and the Abnormal Involuntary Movement Scale (AIMS) were positively related to CD8+D2+ cells, though AIMS was inversely related to CD4+D4+ cells. In conclusion, the alteration in the pattern of cell population and molecules expressed by them might serve as a promising biomarker for diagnosis of schizophrenia.

The crude latex of Euphorbia tirucalli modulates the cytokine response of leukocytes, especially CD4+ T lymphocytes

The plants of the Euphorbiaceae family, especially those of the genus Euphorbia, are frequently used by Brazilian folk communities to treat a wide variety of infectious, tumoral and inflammatory illnesses. Among the species of this genus, Euphorbia tirucalli L. is widely used in some Brazilian regions, such as the Jequitinhonha River Valley. There is evidence that the latex produced by E. tirucalli has antiviral and antitumor activities, but little is known about the mechanisms involved in these effects. It is likely that the mechanism for such activities involves leukocyte activation and cytokine production. In this work, we aimed to evaluate the production of type 1 (TNF-α and IFN-γ) and type 2 (IL-4 and IL-10) cytokines by circulating leukocyte subsets submitted to brief stimulation with the crude latex of E. tirucalli. Peripheral blood leukocytes of twenty healthy subjects were submitted to 4 h incubation with crude E. tirucalli latex diluted in dimethylsulfoxide. After the incubation period, the cells were stained with FITC-conjugated monoclonal antibodies specific to the cell surface receptors CD4, CD8 and CD14, and to PE-conjugated monoclonal antibodies specific to the cytokines TNF-α, IFN-γ, IL-4 and IL-10. The acquisition and analysis of data were performed by flow cytometry. The results showed a significant increase (p<0.05) in the percentage of CD4+ T lymphocytes positive for the type 1 cytokines TNF-α and IFN-γ. Neutrophils and CD8+ T lymphocytes showed a mixed profile of cytokine production, characterized by an increase in the percentage of cells expressing IFN-γ, TNF-α, and IL-10. The data indicate a predominant type 1 cytokine response. The findings presented suggest that the effect popularly attributed to E. tirucalli usage may be attributed to its effect on the production of TNF-α and IFN-γ. However, the relationship between the in vitro and in vivo effects of E. tirucalli needs to be investigated.

Immunologic markers, uveitis, and keratoconjunctivitis sicca associated with human T-cell lymphotropic virus type 1.

PURPOSE: To verify the occurrence of keratoconjunctivitis sicca (KCS) and human T-cell lymphotropic virus type 1 (HTLV-1) associated uveitis (HAU) and to evaluate the immunologic status related to HTLV-1. DESIGN: Cross-sectional study. METHODS: Ophthalmic examination (both eyes) and immunophenotyping of peripheral blood lymphocytes were performed in 207 infected asymptomatic blood donors (AS), 55 controls (NI), and 55 patients with HTLV-1 associated myelopathy (HAM/TSP). Examiner was masked to patient's serologic status. RESULTS: KCS was more frequent in HAM/TSP (30/55, 54.5%) than in NI and AS (07/55, 12.7% and 42/207, 20.3%, respectively). Presence of lacrimal hyposecretion in KCS individuals was higher in the HAM/TSP group (P < .001) as compared with NI and AS. HAU was found in 1/55 (1.82%) of HAM/TSP patients and 4/207 (1.93%) of HTLV-1 seropositive donors. Higher levels of activated CD4(+) and CD8(+) T cells were observed in HAM/TSP. Patients with HAU displayed higher percentage of both CD4(+) HLA-DR(+) and CD8(+)HLA-DR(+) when compared with NI and AS without HAU. CONCLUSIONS: Patients with HAM/TSP manifested more ophthalmologic symptoms than asymptomatic HTLV-1-infected individuals, with significantly higher KCS and immunologic alterations. Levels of activated CD8+ T cells could be used as a prognosis marker of inflammatory disease manifestation to follow-up AS individuals.

Infecçäo e doença pelos vírus linfotrópicos humanos de células T (HTLV-I/II) no Brasil / Infection and disease caused by the human T cell lymphotropic viruses type I and II in Brazil

A infecçäo pelos vírus HTLV-I/II encontra-se presente em todas as regiöes brasileiras, mas as prevalências variam de um estado para outro, sendo mais elevadas na Bahia, Pernambuco e Pará. As estimativas indicam que o Brasil possui o maior número absoluto de indivíduos infectados no mundo. Testes de triagem de doadores e estudos conduzidos em grupos especiais (populaçöes indígenas, usuários de drogas intravenosas e gestantes) constituem as principais fontes de informaçäo sobre essas viroses em nosso país. O HTLV-I causa a leucemia/linfoma de células T do adulto (LLTA), a paraparesia espástica tropical/mielopatia associada ao HTLV (TSP/HAM), uveíte associada ao HTLV (HAU) e anormalidades dermatológicas e imunológicas. O HTLV-II näo se mostrou associado a nenhuma doença até o momento. O diagnóstico é feito com testes de triagem (ELISA, aglutinaçäo) e confirmatórios (Western Blot, PCR). Estes vírus säo transmitidos pelo sangue e agulhas contaminadas, através de relaçöes sexuais e de mäe para filho, especialmente através do aleitamento materno. Medidas de prevençäo devem focalizar a orientaçäo de doadores soropositivos, mäes infectadas e usuários de drogas intravenosas

[Infection and disease caused by the human T cell lymphotropic viruses type I and II in Brazil]. / Infecção e doença pelos vírus linfotrópicos humanos de células T (HTLV-I/II) no Brasil.

HTLV-I/II infection is present in all regions of Brazil, but its prevalence varies according to the geographical area, being higher in Bahia, Pernambuco and Pará. It has been estimated that Brazil has the highest absolute number of infected individuals in the world. Blood donors screening and research conducted with special groups (indigenous population of Brazil, IV drug users and pregnant women) are the major sources of information about these viruses in our Country. HTLV-I causes adult T cell leukemia/lymphoma (ATLL), HTLV associated myelopathy/tropical spastic paraparesis (HAM/TSP), HTLV associated uveitis (HAU), dermatological and immunological abnormalities. HTLV-II is not consistently associated with any disease. Diagnosis is established using screening (enzymatic assays, agglutination) and confirmatory (Western blot, PCR) tests. The viruses are transmitted by blood and contaminated needles, by sexual relations and from mother to child, especially by breast feeding. Prevention efforts should focus on education of positive blood donors, infected mothers and IV drug users.