Inflammatory response induced by Helicobacter pylori infection in lung.

Helicobacter pylori is a microorganism that in the last years has been associated with extragastric disorders such as respiratory diseases, however, its impact on lung is partially understood. The aim of this work was to study infection impact of H. pylori on the inflammatory markers expression at the pulmonary level using an animal model. Infection was performed by BALB/c wild type (WT) mice orotracheal instillation with 20 µl of 1 × 108H. pylori reference strain suspension once per day throughout 3 days. Inflammatory response was evaluated at 3, 7, 14, 21 and 30 days post infection. Lung was aseptically removed and pulmonary edema index values showed a significant change at 30 days of infection. Hematoxylin-Eosin (H-E) stain allowed to visualizing H. pylori presence in lung samples at 3 days of infection near the phagocytic cells or in the alveoli lumen. Bronchoalveolar lavage (BAL) was used for inflammatory response evaluation. Lactate dehydrogenase values showed a gradual increase in infected animals along infection time. Protein concentrations in mg/ml from BAL increased significantly at 7 days in infected animals. Macrophages viability obtained from BAL, decreased at the first moment of infection, maintaining constant values along contamination time. Results obtained demonstrate an inflammatory response in lung after orotracheal H. pylori infection and suggest that the pathogenic mechanism is strongly evidenced by tissue damage, endothelial dysfunction inflammatory mediators and markers expression at the pulmonary level.

Study of Helicobacter pylori infection on lung using an animal model.

Helicobacter pylori infection has been reported to be associated with extra-digestive disorders such as respiratory diseases; however, the impact of H. pylori on lung is incompletely understood. Inflammatory response is mediated by the release of cytokines, interferon, and enzymes such as metalloproteinases (MMPs). This may contribute to collagen accumulation during the early phase of infection. MMP expression is an important factor for the proliferation and infiltration of lung cells in the process of fibrosis formation. The aim of this work was to study the impact of the infection with H. pylori on lung using a mouse model. We looked for histological lesions of lung infected with the microorganism as well as the expression of inflammatory and of endothelial dysfunction markers. C57BL/6 wild type (WT) mice were infected by orotracheal instillation with 20 µl of 1 × 108H. pylori reference strain suspension once per day for 3 days. Animals infected and controls were sacrificed at 3, 7, 14, 21 and 30 days. The lung from mice were stained with Hematoxylin-Eosin (H-E), Masson's Trichromic and Periodic Acid Schifft (PAS) for histological study. Also, lipid hydroperoxides and enzime catalase (CAT) activity were determined. Expression level of multiple markers implicated in inflammation (tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-4, IL-6, IL-8, IL-10; metalloproteinase MMP-9) and markers of endothelial dysfunction (I-CAM and V-CAM) was determined from lung tissues mRNA using RT-PCR. Results showed that H. pylori induced morphological changes in the lung tissue with recruitment of inflammatory cells and lung parenchymal cell degradation. The mRNA of IL-1ß and TNF-α; MMP-9, I-CAM and V-CAM increased at 3-7 days infections. Also, iNOS, IL-8 and Phosphocholine cytidyltransferase (CCT) increased with lung injury. Anti-inflammatory interleukin as: IL-4, and IL-10 increased at 7 and 14 days post infection respectively. The results obtained suggest that the pathogenic mechanism of H. pylori on lung could be strongly associated with lung injury as indicate the expression increased of inflammatory mediators and markers of endothelial dysfunction.

Allopregnanolone modulates striatal dopamingergic activity of rats under different gonadal hormones conditions.

OBJECTIVES: Progesterone modulates dopamine (DA) release in corpus striatum. Our objective was to evaluate the effect of the i.c.v injection of the neurosteroid allopregnanolone (ALL), a progesterone metabolite on dopaminergic activity in the corpus striatum of rats under different gonadal hormonal conditions. METHODS: We have measured the concentrations of DOPA, DA and DOPAC (main metabolite of DA) in the corpus striatum in estrus and diestrus rats and in ovariectomized rats without hormonal replacement (OVX group) and primed with estrogen and progesterone (OVX(i) group). Additionally, we have used the aromatic acid decarboxylase inhibitor NSD in order to evaluate the function of tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine synthesis. RESULTS: ALL significantly decreased the striatal concentrations of both DA and DOPAC in the estrus. On the other hand, ALL increased significantly the levels of DA in the OVX(i) group. The DOPA accumulation in OVX(i) after NSD treatment in the ALL-treated groups was greater than in the vehicle group. However, the estrus group did not modify the DOPA accumulation after NSD injection. DISCUSSION: Our results suggest that ALL could modulate the dopaminergic transmission in the corpus striatum by causing changes in the activity of TH and/or in the pre- and post-synaptic dopaminergic terminals in the corpus striatum. This neurosteroidal mechanism could be a new kind of neurotransmitter systems modulation accomplished on TH activity itself and/or on the second messengers not related to ionic channels. Additionally, our results reinforce the idea of a close relationship between the fast non-genomic mechanism of ALL and the genomic actions of estrogen and progesterone.

TNFa increases in vitro migration of human HPV18-positive SW756 cervical carcinoma cells

TNFa has been associated with both, tumor survival and apoptosis. This cytokine is also involved in promoting cell migration during wound healing and tumorigenesis. SW756 is a HPV18-positive cervical carcinoma cell line, which has been used to study different mechanisms of cervical cancer progression. An in vitro assay of scratch wound healing onto monolayers of SW756 cells was used to assess the effect of TNFa on cell migration into a wound space. It was found that SW756 cells have the ability to migrate, but not proliferate in response to scratch wounding in a serum-free medium supplemented with TNFa. RT-PCR analysis showed that SW756 cells express TNFa mRNA when incubated in medium with and without serum. Wound closure and migration rate of SW756 cells were significantly increased in the presence of serum-free media supplemented with TNFa (10 ng/mL) as compared to serum-free media, and media supplemented with either anti-TNFa antibody or both TNFa and anti-TNFa antibody (p<0.05). The results showed a stimulatory effect of TNFa on the migration of SW756 cervical carcinoma cells, suggesting a novel and important role for TNFa in cervical cancer progression.

Seasonal variations of gonadotropins in the pars distalis male viscacha pituitary. Effect of chronic melatonin treatment.

The gonadotropes, LH and FSH cells, were immunohistochemically identified in the pituitary pars distalis of the adult male viscacha (Lagostomus maximus maximus) using specific antibodies against hLHbeta and hFSHbeta with the streptavidin-biotin-peroxidase complex. The distribution, size and percentage immunopositive area of these cells were analyzed by image analysis in viscachas captured during the annual reproductive cycle and after the chronic administration of melatonin. The LHbeta and FSHbeta cells showed seasonal changes in the distribution, size and percentage immunopositive area. The LHbeta cells were found widely distributed throughout the pars distalis during the reproductive period, and they were found in the ventro-medial region in the pars distalis during the gonadal regression and gonadal recovery periods. The LHbeta cells reached the largest size and immunopositive area during the reproductive period and the smallest size and immunopositive area during the gonadal regression period. The FSHbeta cells were found in the ventro-medial region during reproductive and gonadal regression periods. The FSHbeta cells were found widely distributed throughout the pars distalis during the gonadal recovery period when they showed the maximum percentage immunopositive area. A decrease in the size of LHbeta and FSHbeta cells was observed after the chronic administration of melatonin. Moreover, it produces a decrease in the immunopositive area occupied by the LHbeta cells but not in the immunopositive area occupied by the FSHbeta cells. Our results show great activity of LHbeta and FSHbeta cells in different moments of the annual reproductive cycle demonstrating that these cells do not secrete in parallel. Moreover, melatonin acts differentially on the activity of the gonadotrope cells.

TNFalpha increases in vitro migration of human HPV18-positive SW756 cervical carcinoma cells.

TNFalpha has been associated with both, tumor survival and apoptosis. This cytokine is also involved in promoting cell migration during wound healing and tumorigenesis. SW756 is a HPV18-positive cervical carcinoma cell line, which has been used to study different mechanisms of cervical cancer progression. An in vitro assay of scratch wound healing onto monolayers of SW756 cells was used to assess the effect of TNFalpha on cell migration into a wound space. It was found that SW756 cells have the ability to migrate, but not proliferate in response to scratch wounding in a serum-free medium supplemented with TNFalpha. RT-PCR analysis showed that SW756 cells express TNFalpha mRNA when incubated in medium with and without serum. Wound closure and migration rate of SW756 cells were significantly increased in the presence of serum-free media supplemented with TNFalpha (10 ng/mL) as compared to serum-free media, and media supplemented with either anti-TNFalpha antibody or both TNFalpha and anti-TNFalpha antibody (p < 0.05). The results showed a stimulatory effect of TNFalpha on the migration of SW756 cervical carcinoma cells, suggesting a novel and important role for TNFalpha in cervical cancer progression.

TNFalpha increases in vitro migration of human HPV18-positive SW756 cervical carcinoma cells.

TNFalpha has been associated with both, tumor survival and apoptosis. This cytokine is also involved in promoting cell migration during wound healing and tumorigenesis. SW756 is a HPV18-positive cervical carcinoma cell line, which has been used to study different mechanisms of cervical cancer progression. An in vitro assay of scratch wound healing onto monolayers of SW756 cells was used to assess the effect of TNFalpha on cell migration into a wound space. It was found that SW756 cells have the ability to migrate, but not proliferate in response to scratch wounding in a serum-free medium supplemented with TNFalpha. RT-PCR analysis showed that SW756 cells express TNFalpha mRNA when incubated in medium with and without serum. Wound closure and migration rate of SW756 cells were significantly increased in the presence of serum-free media supplemented with TNFalpha (10 ng/mL) as compared to serum-free media, and media supplemented with either anti-TNFalpha antibody or both TNFalpha and anti-TNFalpha antibody (p < 0.05). The results showed a stimulatory effect of TNFalpha on the migration of SW756 cervical carcinoma cells, suggesting a novel and important role for TNFalpha in cervical cancer progression.

Uterine mast cells: a new hypothesis to understand how we are born

Birth is the result of complex, well-defined, and coordinated events, that are tightly regulated by endocrine, nervous, and immune responses, and take place primarily in the female reproductive tract. Various mechanisms and mediators involved in pregnancy, labor, and delivery, are highly conserved among different mammalian species and mast cells emerge as potential and crucial participants in these processes, as it is discussed in this review.

Uterine mast cells: a new hypothesis to understand how we are born

Birth is the result of complex, well-defined, and coordinated events, that are tightly regulated by endocrine, nervous, and immune responses, and take place primarily in the female reproductive tract. Various mechanisms and mediators involved in pregnancy, labor, and delivery, are highly conserved among different mammalian species and mast cells emerge as potential and crucial participants in these processes, as it is discussed in this review. (AU)

Role of mast cells in gastrointestinal mucosal defense

The purpose of this review, based on studies from our laboratory as well as from others, is to summarize salient features of mast cell immunobiology and to describe their associations with gastrointestinal mucosal defense. Gastrointestinal mast cells are involved in many pathologic effects, such as food hypersensitivity. On the other hand, they also play a protective role in defense against parasitic and microbial infections. Thus, they have both positive and negative effects, but presently the mechanisms that control the balance of these various effects are poorly known. It has been suggested that stabilization of mast cells may be a key mechanism to protect the gastrointestinal tract from injury. Few molecules are known to possess both mast cell stabilizing and gastrointestinal cytoprotective activity. These include zinc compounds, sodium cromoglycate, FPL 52694, ketotifen, aloe vera, certain flavonoids such as quercetin, some sulfated proteoglycans such as chondroitin sulfate and dehydroleucodine. Dehydroleucodine, a sesquiterpene lactone isolated from Artemisia douglasiana Besser, exhibits anti-inflammatory and gastrointestinal cytoprotective action. The lactone stimulates mucus production, and inhibits histamine and serotonin release from intestinal mast cells. The lactone could act as a selective mast cell stabilizer by releasing cytoprotective factors and inhibiting pro-inflammatory mast cell mediators.

Role of mast cells in gastrointestinal mucosal defense

The purpose of this review, based on studies from our laboratory as well as from others, is to summarize salient features of mast cell immunobiology and to describe their associations with gastrointestinal mucosal defense. Gastrointestinal mast cells are involved in many pathologic effects, such as food hypersensitivity. On the other hand, they also play a protective role in defense against parasitic and microbial infections. Thus, they have both positive and negative effects, but presently the mechanisms that control the balance of these various effects are poorly known. It has been suggested that stabilization of mast cells may be a key mechanism to protect the gastrointestinal tract from injury. Few molecules are known to possess both mast cell stabilizing and gastrointestinal cytoprotective activity. These include zinc compounds, sodium cromoglycate, FPL 52694, ketotifen, aloe vera, certain flavonoids such as quercetin, some sulfated proteoglycans such as chondroitin sulfate and dehydroleucodine. Dehydroleucodine, a sesquiterpene lactone isolated from Artemisia douglasiana Besser, exhibits anti-inflammatory and gastrointestinal cytoprotective action. The lactone stimulates mucus production, and inhibits histamine and serotonin release from intestinal mast cells. The lactone could act as a selective mast cell stabilizer by releasing cytoprotective factors and inhibiting pro-inflammatory mast cell mediators. (AU)

Effect of dehydroleucodine on histamine and serotonin release from mast cells in the isolated mouse jejunum.

OBJECTIVE AND DESIGN: DhL, a lactone isolated from Artemisia douglasiana, prevents gastrointestinal damage elicited by necrosis-inducing agents and exhibits antiinflammatory action. This work examines the effect of DhL on compound 48/80-induced histamine and serotonin release in the isolated mouse jejunum, to determine whether DhL inhibits mediator release from mast cells at the enteric level. MATERIAL: Thirty jejuna from male Balb-c mice were used for the studies. TREATMENT: Samples were incubated sequentially in 9 test tubes containing RBS or 10 microg/ml compound 48/80 or 1.6 mmol/l + 10 microg/ml compound 48/80 at 37 degrees C for 90 minutes (10 min per tube). METHODS: Histamine and serotonin release studies, quantification of granulated mast cells, and evaluation of mast cell ultrastructure were carried out. Differences between groups were determined using analysis of variance followed by Tukey-Kramer multiple comparisons test. RESULTS: Compound 48/80 increased histamine and serotonin release by the tissue (141.95 +/- 62.58 pg/mg tissue vs basal 5.45 +/- 1.04, P<0.01 and 20.04 +/- 2.81 vs basal 9.24 +/- 1.56 ng/ mg tissue, P<0.01, respectively), decreased the number of granulated submucosal mast cells (0.077 +/- 0.0035 vs basal 0.14 +/- 0.015, P<0.05), and elicited evident granule ultrastructural changes. These effects were reduced by dehydroleucodine (19.51 +/- 7.88, P<0.01; 12.69 +/- 1, P<0.05 and 0.143 +/- 0.014, P<0.05, respectively). CONCLUSION: The lactone inhibits compound 48/80-induced histamine and serotonin release from mast cells in the isolated mouse jejunum.

Changes in gastroduodenal 5-hydroxytryptamine-containing cells induced by dehydroleucodine.

In previous work, we demonstrated that dehydroleucodine (DhL), a lactone isolated from Artemisia douglasiana Besser, prevents gastroduodenal damage induced by absolute ethanol (EtOH). The present study examined the effects of DhL - applied alone or before EtOH - on gastroduodenal cells containing 5-hydroxytryptamine (5-HT), to clarify the mechanism of action of the drug. Mice were divided into four groups: (I) control; (II) DhL; (III) EtOH, and (IV) DhL + EtOH. Stomachs and duodena were studied by immunohistochemistry and image analysis. EtOH decreased the immunopositive cell number and the area occupied by these cells. This effect was prevented by DhL. DhL alone did not affect the gastric immunopositive cell number and area. Duodena treated only with DhL exhibited a reduction of immunopositive cell number, but no change in area was observed. We propose that the drug probably inhibits the release of the inflammatory mediator 5-HT from endocrine cells, acting as a 'cell stabilizer' in response to injury.

Effect of dehydroleucodine on mucus production: a quantitative study.

We have demonstrated that dehydroleucodine (DhL), a lactone isolated from Artemisia douglasiana Besser, prevents gastroduodenal damage induced by necrosis-inducing agents such as absolute ethanol. We have also reported, in a qualitative study, that this effect is related to the ability of the drug to stimulate mucus production. The present study was designed to quantitatively evaluate the effect of DhL on adherent mucus layer thickness, to obtain a more objective approach to the mechanism of action of the drug. Mice were divided into two groups: (I) controls were treated with orally administered carboxymethylcellulose (CMC) and (II) experimental animals received DhL in CMC. The thickness of the mucus gel layer was measured in unfixed stomachs and duodena, using an image analysis system. We observed an increase in the adherent mucus layer thickness in the experimental samples. This confirms that one of the main mechanisms involved in the cytoprotective action of the drug is mucus secretion.

Gastroduodenal mucosal protection induced by dehydroleucodine: mucus secretion and role of monoamines.

In previous work we have demonstrated that dehydroleucodine (DhL) prevents gastric damage induced by necrosis-inducing agents such as absolute ethanol (EtOH). In this study we examine the effects of DhL on gastroduodenal morphology and monoamine levels by histological and biochemical methods, respectively, as an approach to elucidating the cytoprotective mechanism of the drug. Histological evidence shows that DhL prevents formation of gastroduodenal mucosal lesions induced by EtOH and that this protective effect is related to the ability of the drug to stimulate mucus production. DhL itself does not affect the tissue concentration of NE, DA and 5-HT. However, it prevents the depletion of DA and 5-HT provoked by EtOH. We propose that the abundant mucoid blanket secreted after treatment with DhL acts as a diffusion barrier against EtOH. It is also possible that DhL could act as a "cell stabilizer," by inhibiting the degranulation of cells containing monoamines.