Anti-inflammatory Assessment of Methanol Extract of Acalypha ciliata Leaves and It’s Leucocyte Mobilization in Adult Wistar Rats

Aim: The study's objective was to examine the anti-inflammatory activity and leucocyte mobilization of a methanol extract of leaves from Acalypha ciliata in adult Wistar rats. Study Design: Egg albumin-induced edema was used for anti-inflammatory test and leucocyte mobilization test was carried out to check for total leucocyte cout and differentials. Place and Duration of the Study: This original study was conducted between January and June 2013 at the Department of Biochemistry, Faculty of Biological Sciences, University of Nigeria Nsukka. Methology: The pulverized leaf of Acalypha ciliata was extracted using cold maceration, and the Treas and Evans technique of phytochemical screening was used. The anti-inflammatory study was conducted using the method of Winter et al., and the acute toxicity study was assessed using Lorke's method. ANOVA was used to statistically examine the collected data. Results: Preliminary phytochemical screening revealed the presence of flavonoids, alkaloids, soluble carbohydrates, phenols, glycosides, saponins, terpenoids and steroids. Flavonoids, Alkaloids, and Soluble carbohydrates appeared in abundant concentration (2199.76±10.81, 2141.6±7.583, 913±21.06). Acute toxicity tests showed no toxicity and mortality at doses up to 5000 mgkg-1. Anti-inflammatory study revealed that group treated with 100, 200 and 400 mg/kg b.w of extract significantly (P =0.05) and in a dose dependant manner decreased in TLC when compared with normal control. Moreover, the group treated with 100,200,400 mg/kg b.w of extract and reference drug(Indometacin) mobilized the leucocyte at the rate of 36, 22, 12 and 62 % respectively. A significant (P = 0.05) reduction in the mean paw oedema was observed for all the treatment groups from 1 hour to 5 hours when compared to the toxic group. Conclusion: As a result of this study's findings, it can be concluded that methanol extract of the Acalypha ciliata leaf has anti-inflammatory characteristics. High dosages of the extract resulted in a better decrease of oedema and an increase in leukocyte mobilization in response to agar suspension than low doses. Many anti-inflammatory herbs and substances alter inflammatory reactions by hastening the breakdown of or reversing the effect of the inflammatory mediators. The plant's anti-inflammatory properties could potentially result from the interaction of different photochemical substances present. The findings suggest that, if used, the plant may serve as a source of anti-inflammatory compounds.

Exopolysaccharides from Klebsiella oxytoca: anti-inflammatory activity

Abstract Exopolysaccharides (EPS) produced by Klebsiella oxytoca are of environmental, pharmaceutical, and medicinal interest. However, studies about the anti-inflammatory activity of EPS produced by this microorganism still remain limited. The aim of this study was to produce, characterize, and evaluate the anti-inflammatory activity of EPS from K. oxytoca in a pleurisy model. Colorimetric analysis revealed that precipitated crude exopolysaccharides (KEPSC) and deproteinated exopolysaccharides (KEPS) present high levels of total carbohydrates (65.57% and 62.82%, respectively). Analyses of uronic acid (7.90% in KEPSC and 6.21% in KEPS) and pyruvic acid (3.01% in KEPSC and 1.68% in KEPS) confirm that the EPS are acidic. Gas chromatography-mass spectrometry analyses demonstrated that the EPS consisted of rhamnose (29.83%), glucose (11.21%), galactose (52.45%), and mannose (6.50%). The treatment of an experimental pleurisy model in rats through subcutaneous administration of 50, 100, 200, and 400 mg/kg of KEPS decreased both the volume of inflammatory exudate and the number of leukocytes recruited to the pleural cavity. The present data showed that EPS production by K. oxytoca using the method described is easy to perform and results in a good yield. In addition, we show that KEPS exhibit anti-inflammatory activity when administered subcutaneously in rats.

The Optimum Working Concentration of Antigen in DLST/LMT (in vitro Diagnostic Testing) of Allergic Liver Damage Induced by Turmeric / 日本補完代替医療学会誌

Turmeric-induced liver damage, which is driven mostly by allergic mechanisms, is the most common disease related to health food intake. A positive drug-induced lymphocyte stimulation test (DLST) is considered evidence in the diagnosis of this condition. However, a false positive result may occur with the DLST as the blood concentration of turmeric in the body is unclear, which casts doubt on the reliability of the test. This reliability problem is mainly due to insufficient control data about antigen activity during the testing process. Given these circumstances, this study examined the optimal antigen concentration of turmeric using the leukocyte migration test (LMT).The results of nine volunteer healthy individuals suggested that a reaction at antigen levels of 0.1 µg/mL or higher could produce false positive reactions through the effects of mitogen activation. Therefore, when DLST, LMT, or any other similar test is performed for turmeric, an antigen response of 10 ng/mL or lower is necessary. The results of these tests for curcumin also suggested that the same level of concentration is suitable.

A single administration of fish oil inhibits the acute inflammatory response in rats

Objective To investigate the antiinflammatory effects of a single administration of fish oil (FO) on the acute inflammatory response. Methods The paw edema and pleurisy models were used to evaluate the effects of FO dissolved in olive oil (FOP) orally administered in a single dose in rats. Nitric oxide (NO) concentrations in the pleural exudate were performed according to the Griess method and the cytokine concentrations were determined by Luminex bead-based multiplex assay. Results FOP treatment (30 and 300 mg/kg) significantly reduced paw edema. FOP treatment at 18.75, 37.5, 75.0, 150.0, and 300 mg/kg decreased both the volume of pleural exudate and cellular migration into the pleural cavity and each of these doses presented the same effectiveness. Treatment with FOP (300 mg/kg) reduced NO, TNF-α IL-1β and IL-6 concentrations in the pleural exudate. Conclusions The present data provide evidence that FO has inhibitory effects on the acute inflammatory response when administered in a single dose in rats. This effect might be attributable to a direct inhibitory effect of FO on the production or release of inflammatory mediators that are involved in the pathological processes evaluated herein.

A single administration of fish oil inhibits the acute inflammatory response in rats

OBJECTIVE@#To investigate the antiinflammatory effects of a single administration of fish oil (FO) on the acute inflammatory response.@*METHODS@#The paw edema and pleurisy models were used to evaluate the effects of FO dissolved in olive oil (FOP) orally administered in a single dose in rats. Nitric oxide (NO) concentrations in the pleural exudate were performed according to the Griess method and the cytokine concentrations were determined by Luminex bead-based multiplex assay.@*RESULTS@#FOP treatment (30 and 300 mg/kg) significantly reduced paw edema. FOP treatment at 18.75, 37.5, 75.0, 150.0, and 300 mg/kg decreased both the volume of pleural exudate and cellular migration into the pleural cavity and each of these doses presented the same effectiveness. Treatment with FOP (300 mg/kg) reduced NO, TNF-α, IL-1β, and IL-6 concentrations in the pleural exudate.@*CONCLUSIONS@#The present data provide evidence that FO has inhibitory effects on the acute inflammatory response when administered in a single dose in rats. This effect might be attributable to a direct inhibitory effect of FO on the production or release of inflammatory mediators that are involved in the pathological processes evaluated herein.

Oxidative stress and leukocyte migration inhibition response in cutaneous adverse drug reactions.

Background: Cutaneous adverse drug reactions (CADRs) may either be immunological or non-immunological. The precise mechanisms, however, are largely obscure. Other concomitant mechanisms may amplify and/or contribute to the severity and duration of a reaction. One such mechanism could be oxidative stress, a state of imbalance between reactive oxygen species, and their subsequent detoxification by antioxidants. Aims: (a) to assess the oxidative stress status in the blood of cutaneous drug reaction patients by assaying for reduced glutathione (GSH) and malondialdehyde (MDA) levels, (b) to determine the leukocyte migration inhibition (LMI) response in these patients in response to the suspected drug (s), and (c) to look for the association between oxidative stress parameters and LMI. Methods: Ethical committee approval was obtained for this study. Fresh venous blood samples were obtained from the patients of CADRs (group A) during the acute phase of reaction and healthy control subjects (group B). MDA levels, a measure of oxidative lipid damage, and reduced GSH levels, a measure of anti-oxidant capacity, were assayed in the blood samples of both groups using spectrophotometry. LMI response was measured by challenging the patients' peripheral blood mononuclear cells with the suspected drug to confirm immunological perturbation. Results: Totally 66 participants, 33 cases in group A and equal number of controls in group B, were studied. The mean MDA levels were found to be raised (P < 0.001), but GSH levels were significantly reduced in group A when compared with group B (P = <0.001). LMI response against drug(s) was performed in 33 cases (group A), out of which 25 cases showed a positive LMI response as follows: fixed drug eruption (10/25), SJS (5/25), urticaria (3/25), exfoliative dermatitis (2/25), morbilliform rash (2/25), erythroderma (1/25), vasculitis (1/25), and dapsone syndrome (1/25). The mean MDA levels were found to be significantly higher in the LMI positive CADRs (P < 0.001) when compared with LMI-negative ones, while no significant difference was seen for GSH (P = 0.100). Furthermore, there was a significant positive correlation between MDA levels and LMI response (r = 0.831, P < 0.001). On the other hand, a negative but statistically insignificant correlation was found between GSH and LMI response (r = -0.248, P = 0.271). Conclusion: CADR patients were found to be under oxidative stress based on MDA and GSH levels in the peripheral blood. There is a significant positive correlation of LMI response (against the causative drug) with MDA levels, which strongly associates oxidative stress with the immunopathogenesis in CADRs.

In vivo and in vitro anti-inflammatory activity of red clover Trifolium pratense dry extract

Red clover Trifolium pratense L., Fabaceae, contains four isoflavones, mainly formononetin and biochanin A, and in smaller concentrations, daidzein and genistein. These compounds have gained a lot interest due to its human health benefits, such as estrogenic and progestogenic activities, antioxidant, anti-cancer and others. The objective of this study was to determine in vivo and in vitro anti-inflammatory activity of red clover dry extract. The in vitro anti-inflammatory activity was assayed by the technique using the Boyden chamber method, evaluating the leukocyte migration inhibition (chemotaxis). The in vivo anti-inflammatory activity was tested by a carrageenan-induced rat paw edema test. The results of anti-inflammatory in vitro test showed that there was a significant inhibition of leukocyte migration at the concentrations of 100, 50, 25, 10 and 5 µg/mL of red clover dry extract, these doses resulted in 94.73, 95.39, 94.73, 84.68 and 78.75 percent of inhibition for each dose, respectively. The anti-inflammatory in vivo test resulted in a significant activity in both tested doses (100 and 50 mg/kg of red clover dry extract) and at each tested time. The average percentage of edema inhibition was 63.37 percent. The findings of this study suggested that red clover extract might be suitable for the treatment of inflammatory diseases.

Atividade de tropaeolum majus l. sobre a mobilização e migração leucocitária em modelo de bolsão inflamatório / Activity of tropaeolum majus l. on the mobilization and leukocyte migration in an air pouch model

Tropaeolum majus L. (Tropaeolaceae) é uma importante planta medicinal conhecida popularmente no Brasil como chaguinha, capuchinha ou nastúrcio. Toda parte aérea da planta tem sido utilizada há séculos pela medicina popular. Suas folhas secas ou em infusão são usadas popularmente para o tratamento de várias doenças, incluindo processos inflamatórios. O objetivo do presente estudo foi avaliar o perfil da migração leucocitária em vigência de resposta inflamatória aguda, após um tratamento subcrônico com T. majus. Para isto, ratos Wistar machos foram tratados por cinco dias com três diferentes doses do extrato hidroetanólico (EHTM) obtido de T. majus (75, 150 e 300 mg/kg). Os animais controle receberam volume equivalente de solução salina (5,0 ml/kg) ou indometacina (5,0 mg/kg). Durante este período, os animais receberam 10 ml de ar estéril na cavidade subcutânea (air pouch) em três dias alternados. No quinto dia, uma hora após os tratamentos, a resposta inflamatória foi induzida com a administração de 1 ml de carragenina (solução 1%) na cavidade subcutânea, e 6 horas após, amostras de sangue e do exsudato foram coletadas para a determinação de leucócitos totais e para a realização da contagem diferencial. Os resultados demonstraram que a indometacina reduziu o número de leucócitos totais para o exsudato em aproximadamente 65%. O EHTM nas doses de 75 e 300 mg/kg também reduziram significativamente a migração destas células, com valores estimados em 23 e 40%, respectivamente. Estes resultados indicam uma possível atividade anti-inflamatória do T. majus neste modelo experimental, justificando, pelos menos em parte, o uso popular desta espécie.

Tropaeolum majus L. (Tropaeolaceae) is an important medicinal plant popularly known in Brazil as chaguinha, capuchinha or nasturtium. This species has been used for centuries in popular medicine. Dried leaves are popularly used in infusion for the treatment of various diseases, including inflammatory processes. The aim of this study was to assess the profile of leukocyte migration during an acute inflammatory response after a subchronic treatment with T. majus. For this purpose, male Wistar rats were treated for five days with three different doses of hydroalcoholic extract (HETM) obtained from T. majus (75, 150 and 300 mg/kg). Control animals received equivalent volume of saline solution (5.0 ml/kg) or indomethacin (5.0 mg/kg). During this period, the animals received 10 ml of sterile air in the subcutaneous cavity (air pouch) on three alternate days. On the fifth day, one hour after treatment, the inflammatory response was induced by administration of 1 ml carrageenan solution (1%) in the subcutaneous cavity, and 6 hours after, blood and exudate samples were collected for determination of total and differential leukocytes. The results showed that indomethacin reduced leukocyte migration in the exudates to about 65%. The HETM at doses of 75 and 300 mg/kg also significantly reduced the migration of these cells, with values ??of 23 and 40%, respectively. These results indicate a possible anti-inflammatory activity of T. majus in this experimental model, explaining, at least in part, the popular use of this species.

Cytotoxic, antitumor and leukocyte migration activities of resveratrol and sitosterol present in the hidroalcoholic extract of Cissus sicyoides L., Vitaceae, leaves

Cissus sicyoides L. pertains to the Vitaceae family. It is popularly known as "insulina, cipo-pucá, bejuco caro, puci, anil trepador". A vasoconstrictor effect and an antibacterial activity have also been allocated to it. In Brazil, C. sicyoides was evaluated for its anticonvulsant and anti-diabetc properties. Phytochemistry studies identified and isolated sitosterol and resveratrol compounds from its aerial parts which are pointed out as having antitumor activities. The goal of this study was to investigate the cytotoxic and antitumor activities of Cissus sicyoides hydroalcoholic extract as well as its ability to repair leukocytes cells to injured tissue. Cissus sicyoides did not demonstrate cytotoxic activity but showed an inhibition of tumor growth in face of the tumors tested. The extract had a strong chemotactic effect on the twenty four hours period after treatment. The hidroalchoolic extract of Cissus sicyoides presented antitumor activity which was prompted by T lymphocytes recruitment to the local lesion and suggests a new pathway to antitumor activity by activation of lymphoid lineage.

Cissus sicyoides pertence à família das Vitaceae. É conhecido popularmente como "insulina, cipó-puca, bejuco caro, puci, anil trepador". Esta planta apresenta efeito vasoconstritor eatividade antibacteriana. No Brasil, C. sicyoides foi avaliado pelas suas propriedades anticonvulsivante e anti-diabética. Estudos fitoquímicos identificaram e isolaram a partir de suas partes aéreas o sitoesterol e o resveratrol compostos que são apontados por apresentar atividade antitumoral. O objetivo deste estudo foi investigar as atividades citotóxica e antitumoral do extrato hidroalcoólico Cissus sicyoides bem como a sua capacidade de recrutar leucócitos para os tecidos lesados. Cissus sicyoides não demonstrou atividade citotóxica, mas apresentou uma inibição do crescimento tumoral frente aos tumores testados. O extrato teve um forte efeito quimiotático 24 h após o tratamento. O extrato hidroalcoólico de Cissus sicyoides apresentou atividade antitumoral, relacionada ao recrutamento de linfócitos T para o local da lesão sugerindo que esta atividade esteve relacionada à ativação da linhagem linfóide.

Methotrexate inhibits integrin adhesion molecules in the mouse model of pleurisy induced by carrageenan

The aim of this work was to analyze the effect of methotrexate (MTX) upon leukocyte migration and expression of adhesion molecules CD11a/CD18 in the lung, 4 and 48 h after inflammation induction by carrageenan in mice. The results showed that MTX significantly decreased leukocyte influx and CD11a expression in the lung at 4 and 48 h of pleurisy (P < 0.01). MTX also inhibited CD18 expression at 4 h but not 48 h of pleurisy (P < 0.01). These results proved that MTX at the studied doses had important anti-inflammatory properties, acting primarily on leukocyte migration from the pleural cavity to the lung via inhibition of CD11a/CD18 expression in the mouse model of inflammation.

O modelo experimental da pleurisia induzida pela carragenina, em camundongos é caracterizado pelo aumento da migração de leucócitos às custas de neutrófilos 4 h após a indução da inflamação pela carragenina na cavidade pleural de camundongos.. Após 48 h da indução da inflamação ocorre aumento de leucócitos do tipo mononucleares. O objetivo deste trabalho foi avaliar o efeito do metotrexato (MTX) sobre a migração de leucócitos, e a expressão de moléculas de adesão (CD11a/CD18), no pulmão 4 e 48 h após a inflamação induzida pela carragenina.Os resultados demonstraram que o MTX inibiu significativamente o influxo de leucócitos e a expressão de CD11a no pulmão 4 h e 48 h após a inflamação induzida pela carragenina (P < 0.01). O MTX inibiu a expressão de CD18 no pulmão 48 h após, mas não 4 h após esta resposta inflamatória (P < 0.01). Estes resultados demonstram que o MTX, nas doses estudadas, possui importante efeito antiinflamatório agindo principalmente sobre o influxo de leucócitos da cavidade pleural para os pulmões, via inibição da expressão de moléculas de adesão do tipo CD11a/CD18, no modelo da pleurisia induzida pela carragenina, em camundongos.

Investigation of Chemotactic Activities in Differentiated HL-60 Cells by a Time-lapse Videomicroscopic Assay

BACKGROUND: Chemotaxis is one of the cardinal functions of leukocytes, which enables them to be recruited efficiently to the right place at the right time. Analyzing chemotactic activities is important not only for the study on leukocyte migration but also for many other applications including development of new drugs interfering with the chemotactic process. However, there are many technical limitations in the conventional in vitro chemotaxis assays. Here we applied a new optical assay to investigate chemotactic activities induced in differentiated HL-60 cells. METHODS: HL-60 cells were stimulated with 0.8% dimethylformamide (DMF) for 4 days. The cells were analyzed for morphology, flow cytometry as well as chemotactic activities by a time-lapse videomicroscopic assay using a chemotactic microchamber bearing a fibronectin-coated cover slip and an etched silicon chip. RESULTS: Videomicroscopic observation of the real cellular motions in a stable concentration gradient of chemokines demonstrated that HL-60 cells showed chemotaxis to inflammatory chemokines (CCL3, CCL5 and CXCL8) and also a homeostatic chemokine (CXCL12) after DFM-induced differentiation to granulocytic cells. The cells moved randomly at a speed of 6.99+/-1.24 micrometer/min (n=100) in the absence of chemokine. Chemokine stimulation induced directional migration of differentiated HL-60 cells, while they still wandered very much and significantly increased the moving speeds. CONCLUSION: The locomotive patterns of DMF-stimulated HL-60 cells can be analyzed in detail throughout the course of chemotaxis by the use of a time-lapse videomicroscopic assay. DMF-stimulated HL-60 cells may provide a convenient in vitro model for chemotactic studies of neutrophils.

Influência de extratos hidroetanólicos de plantas medicinais sobre a quimiotaxia de leucócitos humanos / Influence of some medicinal plant hydroethanolic extracts on human leukocyte chemotaxis

Vários métodos in vitro têm sido empregados para a investigação das atividades biológicas de plantas usadas na medicina popular para o tratamento de processos inflamatórios. Neste trabalho, investigou-se a influência dos extratos hidroetanólicos de Rauvolfia sellowii Muell. Arg, Hybanthus bigibbosus (St.-Hil) Hassler e Anchieta pyrifolia (Mart.) G. Don, conhecidas popularmente como pau-pra-tudo, canela-de-veado e cipó-suma, respectivamente, sobre a quimiotaxia de leucócitos humanos, estimulados a migrar contra um gradiente de caseína, utilizando-se o método de Boyden. A dexametasona foi utilizada como substância de referência da inibição da quimiotaxia leucocitária. Os resultados demonstraram efeito inibitório significativo de todos os extratos das plantas testadas, sobre a migração de polimorfonucleares, induzida por caseína. Entretanto, essa atividade variou de intensidade conforme a concentração e a espécie estudada. Efeitos máximos foram observados, nas concentrações de 1000, 10 e 1µg/ml com os extratos de pau-pra-tudo, canela-de-veado e cipó-suma, respectivamente, com migração de 81,6±3,9 por cento; 85,4±2,4 por cento e 91,7±2,2 por cento dos polimorfonucleares, enquanto que, com a dexametasona, este efeito foi de 70,3±5,9 por cento. Embora estudos mais aprofundados sejam necessários, os resultados apresentados podem servir como base preliminar de dados, contribuindo para esclarecer o mecanismo da atividade antiinflamatória atribuída às essas plantas na medicina caseira.

Several in vitro methods have been used for the investigation of the biological activities of plants used in folk medicine for the treatment of inflammatory conditions. In this study, we have investigated the ability of the hydroethanolic extracts from Rauvolfia sellowii Muell. Arg, Hybanthus bigibbosus (St.-Hil) Hassler, and Anchieta pyrifolia (Mart.) G. Don, locally known as pau-pra-tudo, canela-de-veado, and cipó-suma, respectively, in interfering with the human leukocytes migration induced by casein, using the Boyden chamber method. Dexamethasone has been used as a positive control for leukocyte inhibition in the same experimental approach. The data herein presented showed a significant inhibition of the casein-induced polymorphonuclear leukocytes migration for all plants studied. However, the intensity of such activity was variable according to the dose and plant tested. For pau-pra-tudo, canela-de-veado, and cipó-suma extracts the average number of migrated polymorphonuclear leukocytes was 81.6±3.9 percent, 85.4±2.4 percent and 91.7±2.2 percent of the input for the doses of 1000, 10, and 1mg/ml, respectively, while for dexamethasone, the value found was 70.3±5.9 percent. Although further studies are needed, the results presented in this study may be useful to clarify the anti-inflammatory properties of these herbal medicines, supporting their ethnobothanical use for the treatment of inflammatory diseases.

DNCB induces colitis and its relation with LMIF activity / 中国病理生理杂志

AIM: In order to explore the pathogenesis of ulcerative colitis (UC), an experimental colitis in mouse was induced by the hapten dinitrochlorobenzene (DNCB), and the activity of leukocyte migration inhibitory factor (LMIF) was measured at the same time. METHODS: 67 BALB/c mice were randomly divided into control (60% ethanol) and DNCB groups. After they were sensitized by smearing 3.3% DNCB on the abdominal skin, they were challenged with DNCB at concentration of 0.1%, 0.2% and 0.4% respectively by instillation once a day. The weight, stool viscosity and hematochezia were observed and accumulated as disease active index (DAI) score. The pathological changes in colon tissue were judged macropathologically and by means of microscope. LMIF activity was determined by the absorbance (A) of migrated leukocytes. RESULTS: Compared to control group, the increases in DAI accumulate score, pathologic score, and LMIF activity in DNCB groups were observed. CONCLUSION: Mouse colitis was induced by DNCB, which was accompanied by an increase in LMIF activity. [

A Study of Leukocyte Migration Inhibition Factor in Behcet's Syndrome / 대한피부과학회지

This study was undertaken to investigate the immunological mechanism of Behqet s syndrome, considered to be important in the pathogenesis of the disease. Seventy- three patients with complete, incomplete and suspected types of Behget's syndrotne were tested for leukocyte migration ingibition factor(LIF), one of the lymphokines. The results were as follows : 1. There was no difference between the average LIF activity of all the patients and that of eontrol. 2. LIF activity of complete type, according to Shirnizus classification, was significaritly lower than the control value. 3. LIF activity of ocular type, according to Lehners classification, was signficantly lower than the control value. 4. LIF activity for patients with 4 clinical symptoms was well below the value for patients with less symptomes 5. For patients with single clinical symptom, LIF activity of complete type was well below the values of incomplete and suspected types. 6. In suspected and mucocutaneous types, LIF activity was low when the patients showed two clinical symptoms than one. Thus, LIF activity was low for patients with complete, ocular and neurological types and with multiple symptorns.