Valdênia Maria Oliveira Souza

Interference of methysergide, a specific 5‐hydroxytryptamine receptor antagonist, with airway chronic allergic inflammation and remodelling in a murine model of asthma

Background Airway remodelling encompasses the structural changes observed in asthmatic airways. Mast cells, through the release of histamine and 5‐hydroxytryptamine (serotonin), are implicated in early asthmatic reactions, bronchoconstriction and mucosal oedema, and in the development of bronchial hyperresponsiveness. However, the association between serotonin and remodelling processes in murine model of airways inflammation remains to be elucidated.

Modulation of the Induction of Lung and Airway Allergy in the Offspring of IFN-γ-Treated Mother Mice

Recent studies have highlighted the influence of fetal/maternal interactions on the development of asthma. Because IFN-γ reduces Th2-mediated allergic responses, we assessed its capacity to modulate asthma in the offspring when injected into mothers during pregnancy. IFN-γ was injected in CD1 female mice on day 6.5 of gestation. Immediately after birth, male newborns were housed in cages with interchanged mothers: the offspring from IFN-γ-treated mothers were breastfed by normal mothers (IFN/nor), and those from normal mothers were breastfed by IFN-γ-treated (Nor/IFN) or normal mothers (Nor/nor). Immediately after weaning, the spleen cells from IFN/nor and Nor/IFN mice produced less IL-4 and more IFN-γ than Nor/nor mice when stimulated with Con A. At the age of 6–7 wk, mice were immunized with OVA on days 0 and 7. From day 14 to 16, they were exposed to aerosolized OVA. The bronchoalveolar lavage fluid from Nor/nor mice showed eosinophilia, a large number of these cells being present in perivascular and peribronchial regions of lung tissues. IFN/nor or Nor/IFN mice showed greatly reduced eosinophil numbers in bronchoalveolar lavage fluid. In addition, lung sections from IFN/nor, but not Nor/IFN mice showed almost normal histology. In OVA-sensitized IFN/nor and Nor/IFN mice, the production of IFN-γ, IL-4, and IL-5 by spleen cells was significantly reduced as compared with cells from the OVA-sensitized Nor/nor group. IgE and anaphylactic IgG1 were also reduced in plasma of IFN/nor mice. In conclusion, the presence of IFN-γ during pregnancy confers to the fetus a protection against allergenic provocations in the adult life.

Discovery of Phthalimides as Immunomodulatory and Antitumor Drug Prototypes

Modulation of the immune system is an emerging concept in the control of tumor growth. While there are many mechanisms that underlie the role the immune system plays in tumor cells, minimizing metastasis by attenuating the expression of pro-angiogenic cytokines, or up-regulating the expression of endothelial factors that are crucial for the angiogenic process in metastasis and alternatively enhancing the antitumor immunity mediated by interferon-g and interleukin-2 are the most significant features identified to date. 3] In view of this, the discovery of small immunomodulating agents is a task that is currently receiving much attention. Among the anticancer and immunomodulatory drug candidates that have entered into clinical trials, the majority are analogues of thalidomide (Thl, 1), such as lenalidomide (Revlimid, CC-5013) and ACTIMID (CC-4047). Studies of structure–activity relationships (SARs) in the analogues and metabolites of Thl have shown that phthalimide is an essential pharmacophoric fragment. Following this line of research, phthalimide has commonly been employed in the design of potential anti-inflammatory, immunomodulatory, antiangiogenic, and antitumor drug candidates. Given this promising outlook, the strategy of molecular hybridization using phthalimide as a pharmacophoric fragment has figured prominently in recent research and has given rise to many successful outcomes. As an example, potent and selective histone deacetylase (HDAC) inhibitors have been designed by hybridizing two distinct structural domains: phthalimide and the hydroxamic acid subunit. This has led to the identification of various SARs and the discovery of a new potent hybrid lead compound, (E)-3-(1oxo-2-(1,2-diphenylethyl)isoindolin-6-yl)-N-hydroxyacrylamide, which has been described as having selective toxicity against tumor cells. On the other hand, the thiosemicarbazones have figured prominently in the vast number of structural subunits used to design anticancer agents. Some well-known mechanisms involving thiosemicarbazones involve the inhibition of ribonucleotide reductase, alteration of DNA structure, and the chelation of endogenous metals. An example of this versatility was recently reported by Gottesman and co-workers, who used the molecular hybridization of the b-isatin scaffold and thiosemicarbazones. It was clearly demonstrated that the insertion of a thiosemicarbazone subunit into optimal templates leads to an improvement in the anticancer properties of b-isatins, paving the way for the discovery of potent and selective anticancer compounds such as the lead compound 1-(5’-fluoroisatin)-4-(4’-methoxyphenyl)-3thiosemicarbazone (2 ; IC50 = 5.2 mm against multidrug-resistant cells that express P-glycoprotein). Because of this unique pharmacological profile, the attachment of thiosemicarbazones has been employed both in the design of ligands for further complexation with transition metals and during the processes of hit-to-lead or lead-todrug conversions. Bearing in mind the molecular pharmacophores outlined above and structural requirements, we describe herein the design, synthesis, and pharmacological evaluation of 11 new potential antitumor and immunomodulatory agents. To establish an appropriate set of SARs, we first prepared phthalimides containing the thiosemicarbazide 2 b or thiosemicarbazone 4 subunits. An attempt was then made to synthesize two bioisosteres of 2 b : the semicarbazide 2 a and aminoguanidine 2 c derivatives, in addition to the analogues of 2 b containing N-methyl or N-phenyl substituents. Subsequently, a short series of phthalimides bearing the thiazolin-4-one ring (compounds 3 a–d) was also investigated, for reasons of the bioisosteric relationship present between thiazolin-4-ones and thiosemicarbazones and the significant number of thiazolin-4-ones that are active against multidrug-resistant cancer cells, as in the case of the lead compound 3. Our design incorporated the molecular hybridization approach suggested by the structural features of prototypes 1–3 in addition to molecular bioisosterism (Figure 1). The compound series 2 a–f was first prepared by using microwave irradiation, in view of a recent report in which the reaction of N-(hydroxymethyl)phthalimide with arylamines using microwave heating under normal conditions rapidly furnishes [a] S. M. T. Cavalcanti, L. C. D. CoÞlho, Dr. M. Z. Hernandes, Prof. A. C. L. Leite, Dr. V. M. O. Souza Department of Pharmaceutical Sciences Centre for Health Science, Federal University of Pernambuco (UFPE) 50740-520 Recife, PE (Brazil) Fax: (+ 55) 081-2126-8510 E-mail : ana.leite@pq.cnpq.br [b] Dr. C. Pessoa, P. M. P. Ferreira, Dr. L. V. C. Lotufo, Prof. M. O. de Moraes Department of Physiology and Pharmacology Centre for Health Science, Federal University of Cear 60430-270 Fortaleza, CE (Brazil) [c] Dr. C. A. De Simone Department of Physics and Informatics Institute of Physics, University of S1⁄4o Paulo 13560-970 S1⁄4o Carlos, SP (Brazil) [d] Dr. V. M. A. Costa, Dr. V. M. O. Souza Laboratory of Immunopathology Keiso-Asami (LIKA) Department of Parasitology, UFPE 50740-520 Recife, PE (Brazil) Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/cmdc.200900525.

β-Lapachone: A naphthoquinone with promising antischistosomal properties in mice

The activity of β-lapachone (3,4-dihydro-2,2-dimethyl-2H-naphthol[1,2-b]pyran-5,6-dione, β-lap) against different stages of Schistosoma mansoni was investigated in mice. Mice infected with 50 cercariae (BH strain) were intraperitoneally treated at a dose of 50 mg/kg for 5 consecutive days, starting on the 1st, 14th, 28th and 45th days after infection, to evaluate the effect of β-lap on skin schistosomula, lung schistosomula, young worms (before oviposition) and adult worms (after oviposition), respectively. All animals were euthanized 60 days after infection. β-Lap significantly reduced (p<0.001) the number of worms in 29.78%, 37.2%, 24.2% and 40.22% when administered during the phases of skin schistosomula, lung schistosomula, young worms and adult worms, respectively. Significant reduction was also achieved in terms of female burden. In all groups, there was significant reduction in the number of eggs and granulomas in the hepatic tissue. When the intervention was performed during the phase of adult worms, β-lap reduced the size of hepatic granulomas and changed the oogram pattern, lowering the percentage of immature eggs and increasing the percentage of mature and dead eggs. Our data indicate that β-lap has moderate antischistosomal properties. Its molecule may also be used as a prototype for synthesis of new naphthoquinone derivatives with potential schistosomicidal properties. Further studies with different formulations containing β-lap are needed to clearly establish the best dose and route of administration and its mechanism of action against schistosomes.

Delayed local inflammatory response induced by Thalassophryne nattereri venom is related to extracellular matrix degradation

Symptoms evoked by Thalassophryne nattereri fish envenomation include local oedema, severe pain and intense necrosis with strikingly inefficient healing, continuing for several weeks or months. Investigations carried out in our laboratory showed that, in the venom‐induced acute inflammation, thrombosis in venules and constrictions in arterioles were highly visible, in contrast to a notable lack of inflammatory cell. Nevertheless, the reason that the venom toxins favour delayed local inflammatory response is poorly defined. In this study, we analysed the movement of leucocytes after T. nattereri venom injection in the intraplantar region of Swiss mice, the production of pro‐inflammatory mediators and the venom potential to elicit matrix metalloproteinase production and extracellular matrix degradation. Total absence of mononuclear and neutrophil influx was observed until 14 days, but the venom stimulates pro‐inflammatory mediator secretion. Matrix metalloproteinases (MMP)‐2 and MMP‐9 were detected in greater quantities, accompanied by tissue degradation of collagenous fibre. An influx of mononuclear cells was noted very late and at this time the levels of IL‐6, IL‐1β and MMP‐2 remained high. Additionally, the action of venom on the cytoskeletal organization was assessed in vitro. Swift F‐actin disruption and subsequent loss of focal adhesion was noted. Collectively these findings show that the altered specific interaction cell‐matrix during the inflammatory process creates an inadequate environment for infiltration of inflammatory cells.

Influence of maternal schistosomiasis on the immunity of adult offspring mice

Schistosoma mansoni infection modulates the immunity to unrelated antigens in the host. In this study, we have investigated the effect of pregnancy and nursing from schistosomotic mother mice on the immune response to ovalbumin (OA), in adult offspring. Then, newborn mice were divided into four groups: animals born from infected mothers (BIM) suckled by non-infected mothers; animals from non-infected mothers suckled by infected mothers (SIM); and two other groups that were mice born and suckled in infected mothers (BSIM) or non-infected (control) mothers. The adult offspring were immunized with OA plus adjuvant. We compared the OA-specific hypersensitivity reactions (HR), antibodies levels (IgG, IgG2a) and the cytokine production in splenocyte cultures. Remarkable interleukin (IL)-10 synthesis was observed in mice BIM; while the anti-OA antibodies levels and immediate HR were impaired. IL-10 neutralization recovered this suppression. Differently, in mice SIM and BSIM there was an enhancement in the anti-OA humoral response and high IL-2 production, however low level of the IL-10 was detected in mice BSIM. In conclusion, schistosomotic pregnancy provides an immunosuppressive potential, IL-10 dependent, which was sustained throughout adult life. Regardless, suckling by infected mothers induces great responsiveness to an unrelated antigen and repairs the inhibitory potential acquired during prenatal stage.

Phthaloyl amino acids as anti-inflammatory and immunomodulatory prototypes

A series of phthalimide analogs were synthesized by derivatization of phthalic anhydride, a highly toxic substance, using a “one pot” condensation reaction to α-amino acids. All phthaloyl amino acid derivatives presented anti-oral inflammatory activity, but compounds 2e and 2g were found to possess the best activities comparable to thalidomide. Most of the compounds effectively suppressed nitric oxide production in murine cells stimulated with lipopolysaccharide. N-phthaloyl amino acids did not exhibit any significant cytotoxicity in vitro when tested against tumor cells as well as a spleen cell culture of BALB/c mice. Compounds 2a, 2g, and 2h were able to inhibit TNF-α and IL-1β production by macrophages. At the same concentration, thalidomide did not exhibit significant inhibitory activity.Graphical Abstract[The pharmacological evaluation of eight N-phthaloyl amino acids 2a–h, including their anti-inflammatory and cytotoxic effects and nitric oxide, TNF-α, and IL-1β production, is described.]

Ultrastructural analysis of β-lapachone-induced surface membrane damage in male adult Schistosoma mansoni BH strain worms.

The present study provides, for the first time, conclusions on the in vitro schistosomicidal properties of β-lap. Adult male Schistosoma mansoni worms of the BH strain were used for the study. Motility, mortality, cell viability and alterations in the tegument were employed as schistosomicidal parameters. Alterations in motility were observed 6h after incubation in concentrations of 50 and 100 μM. β-lap decreased significantly the worm viability, reducing the formation of formazan in 17.7%, 27.4% and 54.8% at concentrations of 25, 50 and 100 μM, respectively. Mortality in concentrations of 50 and 100 μM was of 67% and 100%, respectively, after 24h. The death of the parasite was preceded by progressive surface membrane damage, characterized by tegument peeling, spine reduction and erosion, blister formation and rupture, and the emergence of holes. In addition to this, in the anterior portion, intense general edema, areas of cracking with a wrinkled surface, furrows and a fibrous appearance were also observed. The results of the present study thus provide a sound basis for further in-depth studies of the schistosomicidal properties of β-lap, both in the laboratory and in the field.

Giardia lamblia e alergia respiratória: estudo em uma amostra de crianças de área urbana com frequência elevada da protozoose

OBJECTIVES: There is a high incidence of intestinal parasite infection in urban areas in the Northeast of Brazil. Giardia lamblia infections have been associated with increased prevalence of cutaneous allergies and gastrointestinal disorders. However, little is known about the relationship between giardiasis and allergic diseases of the airways. The present study aimed to investigate the possible association between respiratory allergic diseases and infections by G. lamblia in children from urban areas. METHODS: This study recruited 110 patients of both sexes aged 5-15 years. Patients were administered a questionnaire evaluating clinical symptoms and were given skin tests, parasite tests and serum tests. RESULTS: A high incidence of G. lamblia was observed (45%, 50/110). Infections by this protozoan were not associated with increased risk of respiratory allergy (p = 0.075), high total IgE levels (p = 0.701), positive specific IgE tests (p = 0.250), or positive skin tests for a range of environmental allergens (p = 0.239). CONCLUSION: This study demonstrated that symptoms of asthma, skin allergy and serum markers were not associated with G. lamblia infections in this sample of children from urban areas.OBJECTIVES There is a high incidence of intestinal parasite infection in urban areas in the Northeast of Brazil. Giardia lamblia infections have been associated with increased prevalence of cutaneous allergies and gastrointestinal disorders. However, little is known about the relationship between giardiasis and allergic diseases of the airways. The present study aimed to investigate the possible association between respiratory allergic diseases and infections by Giardia lamblia in children from urban areas. METHODS This study recruited 110 patients of both sexes aged 5-15 years. Patients were administered a questionnaire evaluating clinical symptoms and were given skin tests, parasite tests and serum tests. RESULTS A high incidence of Giardia lamblia was observed (45%, 50/110). Infections by this protozoan were not associated with increased risk of respiratory allergy (p = 0.075), high total IgE levels (p = 0.701), positive specific IgE tests (p = 0.250), or positive skin tests for a range of environmental allergens (p = 0.239). CONCLUSION This study demonstrated that symptoms of asthma, skin allergy and serum markers were not associated with Giardia lamblia infections in this sample of children from urban areas.

Ascaris lumbricoides infection in urban schoolchildren: Specific IgE and IL-10 production ,

BACKGROUND Helminth infections and allergies are diseases with intense Th2 lymphocytes participation and characterised by a high IgE and Interleukin-(IL) IL-4, IL-5 production and eosinophilia. However, helminths also induce IL-10 production, which may alter the outcome of allergic diseases in infected patients. OBJECTIVE This experimental study analyses the relationship between IL-10 production by cell culture from geohelminth infected and non-infected children and specific IgE to Ascaris lumbricoides (Asc) or Blomia tropicalis (BT). METHODS IL-10 content in supernatant from peripheral blood mononuclear cell culture from nine helminth infected and eleven non-infected patients was determined by ELISA after in vitro stimulation with Asc or BT extracts. RESULTS A positive association was observed between total IgE levels and anti-Ascaris and anti-Blomia tropicalis specific IgE, independent of infection status. For both helminth-infected and non-infected groups, there was no difference in IL-10 production in response to Asc extract, even though anti-Ascaris IgE levels were higher in the latter group. In response to BT stimulus, a lower production of IL-10 by the geohelminth-infected group was observed, but with no relationship between IL-10 production and specific IgE to BT. CONCLUSION The results suggest that anti-Ascaris IgE in non-infected patients may be associated to a resistance to parasites. Levels of specific IgE to parasite antigens or B. tropicalis allergen were not impaired by IL-10 production in children from an urban area in which geohelminthiasis is endemic.