Conformational analysis: A new approach by means of chemometrics

In conformational analysis, the systematic search method completely maps the space but suffers from the combinatorial explosion problem because the number of conformations increases exponentially with the number of free rotation angles. This study introduces a new methodology of conformational analysis that controls the combinatorial explosion. It is based on a dimensional reduction of the system through the use of principal component analysis. The results are exactly the same as those obtained for the complete search but, in this case, the number of conformations increases only quadratically with the number of free rotation angles. The method is applied to a series of three drugs: omeprazole. pantoprazole, lansoprazole-benzimidazoles that suppress gastric-acid secretion by means of H(+), K(+)-ATPase enzyme inhibition. (C) 2002 John Wiley Sons. Inc.

Use of comet assay to assess DNA damage in patients infected by Helicobacter pylori - Comparisons between visual and image analyses

Studies of DNA damage in gastric epithelial cells of Helicobacter pylori (H. pylori)-infected patients are conflicting, possibly due to different methods used for scoring DNA damage by Comet assay. Therefore, we compared the sensitivity of visual microscopic analysis (arbitrary units-scores and comets%) and image analysis system (tail moment), in the gastric epithelial cells from the antrum and corpus of 122 H. pylori-infected and 32 non-infected patients. The feasibility of cryopreserved peripheral blood lymphocytes and whole-blood cells for DNA damage biomonitoring was also investigated. In the antrum, the levels of DNA damage were significantly higher in H. pylori-infected patients with gastritis than in non-infected patients with normal mucosa, when evaluated by image analysis system, arbitrary units and comets%. In the corpus, the comets% was not sufficiently sensitive to detect the difference between H. pylori-infected patients with gastritis and non-infected patients with normal mucosa. The image analysis system was sensitive enough to detect differences between non-infected patients and H. pylori-infected patients with mild gastritis and between infected patients with moderate and severe gastritis, in both antrum, and corpus, while arbitrary units and comets% were unable to detect these differences. In cryopreserved peripheral blood lymphocytes, the levels of DNA damage (tail moment) were significantly higher in H. pylori-infected patients with moderate and severe gastritis than in non-infected patients. Overall, our results indicate that the image analysis system is more sensitive and adequate to measure the levels of DNA damage in gastric epithelial cells than the other methods assayed. (c) 2005 Elsevier B.V. All rights reserved.

Cellular immune responses in mice immunized with Anisakis simplex larval antigens

Cellular immune responses to Anisakis simplex L3 antigens were investigated in BALB/c mice injected subcutaneously with a homologous crude extract (CE). Popliteal lymph nodes (PLN) were found to be increased in size and weight after A. simplex CE footpad injection. The effects of A. simplex CE in vitro proliferation were assayed with non-fractionated PLN cells or nylon-wool purified T cells derived from pooled lymph node cells of mice subcutaneously injected with CE. Spleen cells from immunized animals (antigen alone, or larva alone, or antigen plus larva) were studied by flow cytometry. The immunization induced a high proportion of CD4 + and TCR alpha beta + T cells. The number of B cells (CD45 + and TCR alpha beta-) in pre-immunized and infected mice was lower than that observed in animals subjected to infection only. The number of CD4 + T cells increased in the infected and in the pre-immunized and infected mice. In the latter, a decrease of CD8a + T cells was noted. The greatest increase in CD8a+ and TCR alpha beta- T cells was found in mice that had been subjected to infection only. Histological analysis showed that the most prominent lesions were gastric and intestinal in animals infected orally with one larva.

COORDINATE EXPRESSION OF CYTOKERATIN-7 AND CYTOKERATIN-20 DEFINES UNIQUE SUBSETS OF CARCINOMAS

We tested the hypothesis that the coordinate expression of cytokeratin 7 (CK 7) and cytokeratin 20 (CK 20) could distinguish among carcinomas arising from different primary sites. A total of 384 cases of carcinomas primary to various organs, as well as 16 cases of malignant mesothelioma, were evaluated using commercially available monoclonal antibodies and an avidin-biotin immunoperoxidase technique. The subset of tumors strongly expressing both CK 7 and CK 20 included virtually all bladder transitional cell carcinomas and the majority of pancreatic adenocarcinomas; the tumors negative for both CK 7 and CK 20 were largely restricted to hepatocellular, prostate, and renal cell carcinomas in addition to squamous cell and neuroendocrine carcinomas of lung. The CK 7-/CK 20+ immunophenotype, however, was highly characteristic of adenocarcinomas of colorectal origin, whereas CK 7+/CK 20- immunophenotype was typically seen in the vast majority of carcinomas arising from other sites, including ovary, endometrium, breast, and lung, as well as malignant mesothelioma. Gastric carcinomas were the most heterogeneous subgroup with respect to CK 7/CK 20 immunophenotype. In the subset of mucinous tumors, striking immunophenotypic differences were noted among those primary to the breast (CK 7+/CK 20-), gastrointestinal tract (CK 7-/CK 20+), and ovary (CK 7+/CK 20+). In all cases investigated, this CK immunophenotype was invariant in metastatic vs. primary tumors. It is concluded that, in the appropriate clinical setting, the CK 7/CK 20 immunophenotype of carcinomas is a valuable diagnostic marker in the determination of primary site of origin.

Molecular model for the binary complex of uropepsin and pepstatin

The three-dimensional structure of human uropepsin complexed with pepstatin has been modelled using human pepsin as a template. Uropepsin is an aspartic proteinase from the urine, produced in the form of pepsinogen A in the gastric mucosa. The structure is bilobal, consisting of two predominantly beta -sheet lobes which, as observed in other aspartic proteinases, are related by a pseudo twofold axis. A structural comparison between binary complexes of pepsin:pepstatin and uropepsin:pepstatin is discussed. (C) 2001 Academic Press.

Liquid chromatography/electrospray ionization tandem mass spectrometry profiling of compounds from the infusion of Byrsonima fagifolia Niedenzu

A rapid analytical approach suitable to achieve a comprehensive characterization of the compounds present in the infusion prepared from the leaves of Byrsonima fagifolia Niedenzu (Malpighiaceae), a Brazilian plant used as an infusion to treat gastric disorders, was developed. The method was based on high-performance liquid chromatography coupled to electrospray negative ionisation multistage ion trap mass spectrometry (HPLC/ESI-ITMSn). The main ions in the ESI-ITMS spectra were attributed to a quinic acid core containing from one to five galloyl units. Quercetin derivatives containing one and two sugar moieties as well as galloyl esterification were also detected. These results indicated that HPLC/ESI-ITMSn is easily applicable to infusions of this plant and allows the rapid and direct identification of these compounds in crude plant extracts. Copyright (C) 2007 John Wiley & Sons, Ltd.

Fungemia by yeasts in Brazil: occurrence and phenotypic study of strains isolated at the Public Hospital, Botucatu, São Paulo

Objective. - In this study strains of yeasts isolated from the blood of human patients were analyzed taxonomically, their virulence factors were determined and compared, and phenotypic markers were used to compare the samples with respect to phenotypic differences across the range of patients as well as between samples isotated from the same patient.Methods. - the study involved a total of 75 strains of yeast isolated from the blood of in-patients of the Public Hospital, Botucatu, S (a) over tildeo Paulo, Brazil, with a clinical profile of fungemia. The hospital wards with the largest number of fungemias were neonatal intensive care units (ICUs) (32%) followed by gastric surgery (13.4%) and pediatric wards (10.7%). After identification, the samples were analyzed for the production of phospholipase and proteinase enzymes, and biotyped according to their susceptibility to killer toxins.Results. - the most frequent species found was Candida albicans (38.7%) followed by C. parapsilosis (30.7%). In terms of enzyme production, 98.7% of the 75 samples of yeast presented a strongly positive activity for proteinase; however, 78.7% did not present any phospholipasic activity. Six different biotypes were identified, the most frequent being 511 and 888.Conclusion. In association with phenotypic methods, genetic analyses should also be made of the samples under study to help in the rational development of a wider range of preventive measures and better control of hospital-contracted infections. (c) 2005 Published by Elsevier SAS.

Application of liquid chromatography/electrospray ionization tandem mass spectrometry to the analysis of polyphenolic compounds from an infusion of Byrsonima crassa Niedenzu

A fast and reliable method, based on high-performance liquid chromatography coupled to electrospray ionization ion trap tandem mass spectrometry (HPLC/ESI-ITMS), was developed to investigate the infusion prepared from the leaves of Byrsonima crassa Niedenzu (Malpighiaceae), a native plant used in Brazil against gastric disorders. The use of on-line reverse-phase HPLC/ESI-ITMS allowed separation of three major classes of compounds and identification of over 20 very polar compounds characterized as galloylquinic acids, proanthocyanidins, and flavonoid glycosides, as well as the dimeric flavonoid amentoflavone and minor amounts of galloyl hexose and galloyl saccharose. This approach provided data that will allow establishment of a method for a future standardization of the infusion. Copyright (C) 2005 John Wiley & Sons, Ltd.

Natural trans-crotonin: the antiulcerogenic effect of another diterpene isolated from the bark of Croton cajucara Benth.

The nor-clerodane diterpene trans-crotonin isolated from the bark of Croton cajucara BENTH. was investigated for its ability to prevent the formation of gastric-mucosa ulceration in different experimental models in mice. The results obtained from crotonin were compared with those obtained with another diterpene, DHC (trans-dehydrocrotonin) in the same models. When previously administered (p.o.) at the dose of 100 mg/kg, crotonin, as well as DHC, significantly reduced (p<0.05) gastric injury induced by stress (72, 67%), indomethacin/bethanechol (78, 29%) and pylorus ligature (35, 30%). In the HCl/ethanol-induced gastric ulcer model, at oral doses of 100 and 250 mg/kg, crotonin significantly prevented (p<0.05) the formation of gastric lesions by 51 and 56%, respectively, when compared to the control group. Gastric injury was also of significantly less magnitude in the DHC treatment group (p<0.05). In the pylorus-ligature model, crotonin (p.o.), like cimetidine, increased the volume of gastric juice when compared to the control group (p<0.05). No significant modifications where found in gastric parameters such as pH or total acid content after oral crotonin treatment. However, systemic alterations were observed when crotonin (100 mg/kg) was previously administered intraduodenally to mice. We observed significant changes (p<0.001) in gastric-juice parameters such as an increase in volume and a decrease in gastric acidity. Those pre-treated with crotonin as well as with DHC did not increase free mucus production (p>0.05). The results suggest that crotonin presents a significant anti-ulcer effect when assessed in these ulcer-induced models. As with DHC, the antiulcerogenic effects of crotonin are probably related to anti-secretory or/and gastroprotective properties of this substance. In light of results obtained with DHC and natural trans-crotonin in the present study, we concluded that the A-ring of both diterpenes is not directly involved in the antiulcerogenic activity.

Effect of electrolytic and neurotoxic lesions of the median raphe nucleus on anxiety and stress

To study the role played by 5-HT mechanisms of the MRN, behavioural and physiological parameters were presently measured in rats having either electrolytic or 5,7-dihydroxytryptamine (5,7-DHT) lesion of the MRN made 7 days before testing. Half the animals were submitted to 2-h restraint 24 h before the test. In the elevated plus-maze, the electrolytic lesion increased the percentage of open-arm entries and of time spent on open arms - an anxiolytic effect - in both restrained and nonrestrained rats. The neurotoxic lesion had a similar effect, but only on restrained rats. Restraint had anxiogenic effect. The electrolytic lesion increased transitions between the light and dark compartments and the time spent in the bright compartment of the light-dark box in both restrained and nonrestrained rats. The neurotoxic lesion only increased bright time in restrained rats. The incidence, number and size of gastric ulcers were increased by either the electrolytic or the neurotoxic lesion in both restrained and nonrestrained animals. Both types of lesion depleted 5-HT in the hippocampus in restrained and nonrestrained rats. Restraint increased 5-HT levels. These results implicate 5-HT mechanisms of the median raphe nucleus in the regulation of anxiety and in the genesis of gastric stress ulcers. (C) 2001 Elsevier B.V. All rights reserved.

Epstein-Barr virus (EBV) nuclear antigen (EBNA)-4 mutation in EBV-associated malignancies in three different populations

Different ethnic groups with a high human leukocyte antigen (HLA)-A11 prevalence have been shown to experience a high rate of Epstein-Barr virus (EBV) infection, EBV-associated malignancies, and Epstein-Barr nuclear antigen (EBNA)-4 mutations. The epitopes 393-408 and 416-424 of EBNA-4 are major antigenic epitopes that elicit an HLA-A11 cytotoxic T lymphocyte (CTL) response to EBV infection. Mutations selectively involving one or more nucleotide residues in these epitopes affect the antigenicity of EBNA-4, because the mutant EBV strains are not recognized by the HLA-A11-restricted CTLs. To investigate these mutations in common EBV-associated malignancies occurring in different populations, we studied the mutation rate of epitopes 393-408 and 416-424 of EBNA-4 in 25 cases of EBV-associated Hodgkin's disease (HD), nine cases of AIDS-related non-Hodgkin's lymphoma, and 37 cases of EBV-associated gastric carcinoma (GC) from the United States, Brazil, and Japan. We found one or more mutations in these two epitopes in 50% (6/12) of United States HD, 15% (2/13) of Brazilian HD, 50% (6/12) United States GC and 28% (7/25) Japanese GC, and 22% (2/9) of United States AIDS-lymphoma. Similar mutations were found in 30% (3/10) of United States reactive, 0% (0/6) of Brazilian reactive, and 25% (2/8) Japanese reactive tissues. The most frequent amino acid substitutions were virtually identical to those seen in previously reported isolates from EBV-associated nasopharyngeal carcinomas and Burkitt's lymphomas occurring in high prevalence HLA-A11 regions. However, only 2/28 (7%) mutations occurred in HLA-A11-positive patients. Our studies suggest that: 1) EBNA-4 mutations are a common phenomenon in EBV-associated HD, GC, and AIDS-lymphoma; 2) the mutation rate does not vary in these geographic areas and ethnic groups; 3) EBNA-4 mutations in EBV-associated United States and Brazilian HD, United States and Japanese GC, and United States AIDS lymphomas are not related to patients' HLA-A11 status.

Pharmacological and toxicological studies of Drimys angustifolia Miers. (Winteraceae)

Drimys angustifolia Miers. (Winteraceae) is a Brazilian medicinal plant used as analgesic, antiulcer and anti-inflammatory without studies to assure its efficacy and safety Leaf and stem bark extracts were evaluated to determine the antiulcer, analgesic, antiinflammatory and antioxidant activities. Preliminary toxic effects and qualitative phytochemical profile were also performed. The antiulcer activity was detected in both extracts. Administration of the leaf extract at 250 mg/kg inhibited total lesion area by 76.50% (p < 0.01 in ethanol/HCl method), while carbenoxolone at 250 mg/kg reduced lesions by 69.48%. Stem bark extract (250 mg/kg) inhibited lesion by 81.42%, while carbenoxolone by 74.10%. Similar effects were observed in the ethanol-induced ulcer method, but no activity was observed in piroxican model. The effects involve nitric oxide in gastric protection, since the L-NAME treatment reversed the protection given by the extracts. Antioxidant effects suggest an involvement against oxidative stress. In the pain (writhing, tail-flick and hot-plate tests) and inflammation (carrageenan-induced paw edema) models, the extracts did not present any effect. The phytochemical studies demonstrated that both extracts contain flavonoids, saponins, glycosilated triterpenoids, fixed acids, cyanogenic glycosides, quinones, tannins, xanthone and steroidal aglycones. Toxicological studies showed that the extracts are safe at the effective antiulcer doses. (c) 2006 Elsevier B.V.. All rights reserved.

Effect of continuous and pulsed therapeutic ultrasound in the appearance of local recurrence of mammary cancer in rats

Purpose: Ultrasound (US) therapy is an elect rot hermotherapeutic modality that uses US energy to provoke physical and chemical alterations. US therapy has been widely used in physical therapy. However in clinical practice, it is contra-indicated in cancer patients due to the possibility of exacerbating tumor growth.Methods: Sixty-eight female Sprague-Dawley rats bred in UNIFAE vivarium were studied. At 50 days of age, 7, 12-dimetylbenz(a)anthracene (7, 12-DMBA) was administered to 35 rats by gastric gavage to induce mammary cancer After 90 days the mammary glands of the rats belonging to the group with mammary cancer induction and stimulated by US were removed. Animals received either continuous or pulsed US. US waves were generated at a frequency of 1 MHz during 10 days, with an intensity dose of 0.5 W in the continuous group, and 0.9 W (duty cycle: 20%) in the pulsed group.Results: Among the rats treated with continuous US, 44.4% developed local recurrence, while among the rats treated with pulsed US, 22.2% had local tumor recurrence (p < 0.05). No evidence of distant metastases was shown in any of the rats studied.Conclusion: The use ofcontinuous and pulsed therapeutic US promoted the development of local recurrence of mammary cancer in female Sprague-Dawley rats in the postoperative period.

Effect of passive pneumoperitoneum on oesophageal pressure, cardiovascular parameters and blood gas analysis in horses

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Assessment of Gastrointestinal Motility in Renal Transplant Recipients by Alternate Current Biosusceptometry

Background. Gastrointestinal (GI) complications are common after renal transplantation, mainly owing to immunosuppressive therapy. Assessment of GI transit time can facilitate rational management of these disorders.Objective. We evaluate the GI transit parameters in renal transplant recipients taking tacrolimus, azathioprine, and prednisone with the use of the alternate current biosusceptometry (ACB) technique and compared them with healthy volunteers.Methods. Ten renal transplant recipients and 10 healthy volunteers were enrolled in this study. After an overnight fast, patients and volunteers ingested a standard meal containing magnetic markers. The biomagnetic monitoring was performed at 10-minute intervals for at least 8 hours to obtain gastric emptying as well as the colonic arrival time-intensity curves. Mean gastric emptying time (MGET), mean colon arrival time (MCAT), and mean small intestinal transit time (MSITT) were quantified and compared between control and patient groups with results expressed as mean +/- SD.Results. The MGET measured by the ACB technique was 48 +/- 31 minutes and 197 +/- 50 minutes for patients and healthy subjects, respectively. MSITT and MCAT values calculated for patients versus volunteers were 171 +/- 71 minutes versus 197 +/- 71 minutes and 219 +/- 83 minutes versus 373 +/- 52 minutes, respectively. Renal transplant recipients showed significantly faster; gastric emptying and colon arrival times (P < .001) compared with normal volunteers; however, small intestinal transit time was not significantly different (P = .44).Conclusions. In stable renal transplant recipients, the GI transit parameters were significantly faster than in normal healthy volunteers. ACB sensors are versatile technologies that can be used for clinical research, because they offer an excellent opportunity to evaluate GI transit in a noninvasive manner without the use of ionizing radiation.