Kinetics and mechanism of inhibition of serine esterases by fluorinated aminophosphonates.

This paper reviews previously published data and presents new results to address the hypothesis that fluorinated aminophosphonates (FAPs), (RO)(2)P(O)C(CF(3))(2)NHS(O)(2)C(6)H(5), R=alkyl, inhibit serine esterases by scission of the P-C bond. Kinetics studies demonstrated that FAPs are progressive irreversible inhibitors of acetylcholinesterase (AChE, EC 3.1.1.7.), butyrylcholinesterase (BChE, EC 3.1.1.8.), carboxylesterase (CaE, EC 3.1.1.1.), and neuropathy target esterase (NTE, EC 3.1.1.5.), consistent with P-C bond breakage. Chemical reactivity experiments showed that diMe-FAP and diEt-FAP react with water to yield the corresponding dialkylphosphates and (CF(3))(2)CHNHS(O)(2)C(6)H(5), indicating lability of the P-C bond. X-ray crystallography of diEt-FAP revealed an elongated (and therefore weaker) P-C bond (1.8797 (13)A) compared to P-C bonds in dialkylphosphonates lacking alpha-CF(3) groups (1.805-1.822A). Semi-empirical and non-empirical molecular modeling of diEt-FAP and (EtO)(2)P(O)C(CH(3))(2)NHS(O)(2)C(6)H(5) (diEt-AP), which lacks CF(3) groups, indicated lengthening and destabilization of the P-C bond in diEt-FAP compared to diEt-AP. Active site peptide adducts formed by reacting diEt-FAP with BChE and diBu-FAP with NTE catalytic domain (NEST) were identified using peptide mass mapping with mass spectrometry (MS). Mass shifts (mean+/-SE, average mass) for peaks corresponding to active site peptides with diethylphosphoryl and monoethylphosphoryl adducts on BChE were 136.1+/-0.1 and 108.0+/-0.1Da, respectively. Corresponding mass shifts for dibutylphosphoryl and monobutylphosphoryl adducts on NEST were 191.8+/-0.2 and 135.5+/-0.1Da, respectively. Each of these values was statistically identical to the theoretical mass shift for each dialkylphosphoryl and monoalkylphosphoryl species. The MS results demonstrate that inhibition of BChE and NEST by FAPs yields dialkylphosphoryl and monoalkylphosphoryl adducts, consistent with phosphorylation via P-C bond cleavage and aging by net dealkylation. Taken together, predictions from enzyme kinetics, chemical reactivity, X-ray crystallography, and molecular modeling were confirmed by MS and support the hypothesis that FAPs inhibit serine esterases via scission of the P-C bond.

Individual vitality pattern of in situ dental biofilms at different locations in the oral cavity.

The aim of the study was to examine the three-dimensional vitality structure of dental biofilms grown simultaneously at different locations in the oral cavity over a 48-hour period. Eight healthy volunteers wore special acrylic appliances. On each buccal side of the upper and the lower jaw three glass slabs were inserted, allowing for growth of a biofilm mimicking approximal plaque. After 48 h, the specimens were removed and biofilms were stained using two fluorescent dyes which selectively stain vital bacteria green and dead bacteria red. Under the confocal laser scanning microscope optical sections of 1 microm throughout the biofilm were made. To assess the vitality values (proportion of vital bacteria) of the whole biofilm as well as the vitality distribution in the different plaque sections an image analysis program was used. Plaque from the different locations revealed mean vitality values between 64.4 and 75.7% in the upper jaw and between 64.3 and 76.8% in the lower jaw, which were not statistically different. However, a great variation of the vitality values for the different layers and among the 8 subjects was found. Nevertheless, the analysis of the data of each single volunteer revealed a very similar vitality pattern in all twelve locations.

Impact of the intraoral location on the rate of biofilm growth.

The aims of the present study were: a) to assess the impact of the intraoral location on the rate of biofilm growth, and b) to establish an in vivo biofilm model to examine intraoral biofilm growth. Eight healthy volunteers wore acrylic splints with 15 glass slabs each in the upper and lower jaws to build up plaque. After 48 h, the specimens were removed and stained using the vital fluorescence technique. Biofilm thickness was evaluated by confocal laser scanning microscopy (CLSM). The mean plaque thickness amounted to 77.6 +/- 29.1 microm on the buccal sites of the upper jaw and 71.9 +/- 26.3 microm on the buccal sites of lower jaw. On the palatal site a biofilm of 52.1 +/- 26.2 microm thickness was grown, which was significantly less compared with the other locations evaluated (p < 0.001). The results demonstrate that the in situ biofilm thickness on the buccal sites was similar irrespective of the location in the oral cavity. The new splint system described may be a useful tool for further standardised experimental studies regarding influences on growth and structure of intraoral biofilms.

Immune response to respiratory syncytial virus in young Brazilian children.

We have evaluated the cellular and humoral immune response to primary respiratory syncytial virus (RSV) infection in young infants. Serum specimens from 65 patients <=12 months of age (39 males and 26 females, 28 cases <3 months and 37 cases > or = 3 months; median 3 3.9 months) were tested for anti-RSV IgG and IgG subclass antibodies by EIA. Flow cytometry was used to characterize cell surface markers expressed on peripheral blood mononuclear cells (PBMC) from 29 RSV-infected children. There was a low rate of seroconversion in children <3 months of age, whose acute-phase PBMC were mostly T lymphocytes (63.0 +/- 9.0%). In contrast, a higher rate of seroconversion was observed in children >3 months of age, with predominance of B lymphocytes (71.0 +/- 17.7%). Stimulation of PBMC with RSV (2 x 10(5) TCID50) for 48 h did not induce a detectable increase in intracellular cytokines and only a few showed a detectable increase in RSV-specific secreted cytokines. These data suggest that age is an important factor affecting the infants' ability to develop an immune response to RSV.

Immune response to respiratory syncytial virus in young Brazilian children

We have evaluated the cellular and humoral immune response to primary respiratory syncytial virus (RSV) infection in young infants. Serum specimens from 65 patients <=12 months of age (39 males and 26 females, 28 cases <3 months and 37 cases > or = 3 months; median 3 ± 3.9 months) were tested for anti-RSV IgG and IgG subclass antibodies by EIA. Flow cytometry was used to characterize cell surface markers expressed on peripheral blood mononuclear cells (PBMC) from 29 RSV-infected children. There was a low rate of seroconversion in children <3 months of age, whose acute-phase PBMC were mostly T lymphocytes (63.0 ± 9.0 percent). In contrast, a higher rate of seroconversion was observed in children >3 months of age, with predominance of B lymphocytes (71.0 ± 17.7 percent). Stimulation of PBMC with RSV (2 x 10(5) TCID50) for 48 h did not induce a detectable increase in intracellular cytokines and only a few showed a detectable increase in RSV-specific secreted cytokines. These data suggest that age is an important factor affecting the infants' ability to develop an immune response to RSV

Chloroephedrine: contaminant of methamphetamine synthesis with cardiovascular activity.

Chloroephedrine is an intermediate and possible contaminant formed when methamphetamine is manufactured using ephedrine or pseudoephedrine as precursors. The purpose of this study was to determine whether this contaminant has biological activity and might contribute to methamphetamine-induced cardiovascular toxicity. In conscious rats, the (-) and (+) isomers of chloroephedrine (0.1 and 1.0 mg/kg, i.v.) dose-dependently increased mean arterial pressure (MAP) and heart rate (HR). The potency of the pressor effects of (-) and (+)-chloroephedrine was between that of ephedrine and pseudoephedrine. The increases in HR elicited by the four stimulants were similar except that the tachycardia elicited by all doses of ephedrine and pseudoephedrine were preceded by a brief decrease in HR. The i.v. administration of 10 mg/kg of (+) or (-)-chloroephedrine produced biphasic (decrease followed by increase) the MAP and HR responses. Ephedrine and pseudoephedrine did not decrease MAP at any dose tested. The initial decrease in HR elicited by (-)-chloroephedrine was significantly reduced and the hypotensive response abolished by atropine, indicating that these components of the MAP and HR responses resulted from vagal activation. The secondary pressor response elicited by (-)-chloroephedrine was significantly reduced and the tachycardia significantly increased by pretreatment with phentolamine (3 mg/kg, i.v.). The increase in HR was reversed by propranolol. These results indicate that (-) and (+)-chloroephedrine have sympathomimetic properties similar to other known sympathomimetic stimulants. In addition, larger doses of chloroephedrine can activate the vagus nerve. The combination of (+)-methamphetamine and (-)-chloroephedrine did not markedly alter the magnitude of the MAP and HR responses of (+)-methamphetamine alone except at high doses of (-)-chloroephedrine (10 mg/kg). Contamination of illicit methamphetamine with chloroephedrine may have toxic consequences.

Clinical patterns and seasonal trends in respiratory syncytial virus hospitalizations in São Paulo, Brazil.

The respiratory viruses are recognized as the most frequent lower respiratory tract pathogens for infants and young children in developed countries but less is known for developing populations. The authors conducted a prospective study to evaluate the occurrence, clinical patterns, and seasonal trends of viral infections among hospitalized children with lower respiratory tract disease (Group A). The presence of respiratory viruses in children's nasopharyngeal was assessed at admission in a pediatric ward. Cell cultures and immunofluorescence assays were used for viral identification. Complementary tests included blood and pleural cultures conducted for bacterial investigation. Clinical data and radiological exams were recorded at admission and throughout the hospitalization period. To better evaluate the results, a non- respiratory group of patients (Group B) was also constituted for comparison. Starting in February 1995, during a period of 18 months, 414 children were included- 239 in Group A and 175 in Group B. In Group A, 111 children (46.4%) had 114 viruses detected while only 5 children (2.9%) presented viruses in Group B. Respiratory Syncytial Virus was detected in 100 children from Group A (41.8%), Adenovirus in 11 (4.6%), Influenza A virus in 2 (0.8%), and Parainfluenza virus in one child (0.4%). In Group A, aerobic bacteria were found in 14 cases (5.8%). Respiratory Syncytial Virus was associated to other viruses and/or bacteria in six cases. There were two seasonal trends for Respiratory Syncytial Virus cases, which peaked in May and June. All children affected by the virus were younger than 3 years of age, mostly less than one year old. Episodic diffuse bronchial commitment and/or focal alveolar condensation were the clinical patterns more often associated to Respiratory Syncytial Virus cases. All children from Group A survived. In conclusion, it was observed that Respiratory Syncytial Virus was the most frequent pathogen found in hospitalized children admitted for severe respiratory diseases. Affected children were predominantly infants and boys presenting bronchiolitis and focal pneumonias. Similarly to what occurs in other subtropical regions, the virus outbreaks peak in the fall and their occurrence extends to the winter, which parallels an increase in hospital admissions due to respiratory diseases.

[Viral infections in hospitalized children affected by acute lower respiratory tract disease] / Infecções virais em crianças internadas por doença aguda do trato respiratório inferior.

OBJECTIVE: To evaluate the frequency of the main respiratory viruses in hospitalized children affected by acute lower respiratory tract disease at a university hospital. METHODS: This is a prospective trial that included two cohorts of hospitalized children in the period from April to July 1996. The groups were selected according to the presence of lower respiratory tract disease on admission: Group A- with acute disease (history of less than 7 days) and B- without present or recent respiratory disease. The parameters for defining lower respiratory tract disease included physical and/or radiological pulmonary changes. Clinical and radiological criteria were established for the classification of lower respiratory tract diseases in group A. Nasopharyngeal swab was collected from all children on admission for viral detection by cellular cultures and direct immunofluorescence. RESULTS: 201 cases were selected, 126 in group A and 75 in group B. Viruses were identified in 71 children from groupA(56.4%) and only in 3 from group B (4.0%). The predominant agent in group A was respiratory syncytial virus, identified in 66 cases; adenovirus (4) and influenza (1) were detected in other patients. In group B two patients with respiratory syncytial virus and one with adenovirus were identified. The patients from group A affected by respiratory syncytial virus were younger (median age 3 months versus 13 months) and more wheezy on physical examination (78.7%) than the other patients of the group (33.3%). This virus was associated to most of the bronchiolitis cases (84%) and to half of the pneumonia cases (46.4%). CONCLUSION: The authors found a significant presence of viruses in the majority of children hospitalized with acute lower respiratory tract disease. The respiratory syncytial virus was the predominant agent identified. These results are similar to others previously reported both in developed and some developing countries. The authors emphasize that the present study evaluated only partially the possibility of simultaneous infection by other pathogens and that the present protocol was conducted during the season with the highest incidence of respiratory syncytial virus.

[Idiopathic primary pulmonary hemosiderosis: treatment with cyclophosphamide and prednisone]. / Hemossiderose pulonmar idiopática tratada com ciclofosfamida e prednisona.

The authors present the case of a child diagnosed as having idiopathic pulmonary hemosiderosis at five years of age who had a good clinical outcome at the age of ten years. Initially the patient was treated with prednisone and chloroquine with poor results. When cyclophosphamide was added to prednisone, the patient demonstrated clinical and radiological remission. To date, the patient has been followed for one year without any medication, and has had only one limited episode of hemosiderosis. The authors also suggest that the therapeutic regimen with cyclophosphamide and prednisone may be useful for some selected cases.

[Serotypes of pneumococci isolated from children with pneumonia: implication of pneumococcal specific immunization]. / Sorotipos de pneumococos isolados de crianças com pneumonia: implicação com a imunização específica anti-pneumocócica.

For the 15 years from 1978 to 1992 serologic typing was performed on 124 pneumococcus isolates from children with acute pneumonia. The source of bacteria was material obtained by aspirative pulmonary punction, pleural fluid or blood; 122 capsular antigens representing groups and types could be determined. Of the 122 isolates serogrouped 14, 1, 6, 5, 4, 7, 23, 19 and 4, accounted for 25.4%; 23.8%; 13.1%; 9.0%; 4.9%; 4.9%; 4.1%; 4.1%; 4.1%; respectively, of cases. The currently available 23-valent vaccine would provide protection against 89.3% of identified pneumococci in our study, but because of its poor immunogenicity in children less than 2 years old (73.0%) they would have received reduced protection by the use of this vaccine. The distribution of pneumococci serogroups found in our study has an intermediary pattern in relation to those found at develop countries (6, 14, 18, 19, 23) and developing ones (1, 2, 3, 5, 7, 12, 46). The new conjugate vaccines, with limited number of pneumococcal groups/types, should be analysed before the introduction in different geographic areas.

The pursuit of equity: a health sector case study from Vietnam.

The health care system in Vietnam has long been cited as an example of primary health care that has worked well. The achievements of the system during the past decades have indeed been impressive, but the changing economic situation in Vietnam has consequences for all public sector activities, including health care. Liberalization of economic policies has encouraged private medical practice and free trade in medicines and drugs, while financial support for the state health system is decreasing. Equity has always been an important goal for the vietnamese health system, but it becomes harder and harder to realize under the new conditions of financing. The restrictions in centralized planning and funding brought about by recent changes also reveal weak points in the system, from planning to training to management at the different levels. This situation is discussed and issues concerning policy, legislation and human resources are highlighted in terms of their effect on equity.

Falha na terapeutica inalatoria domiciliar para asma: analise e proposta / Therapeutic failure in the domiciliary management of asthmatic children: analysis and proposal

Os autores analisaram 19 casos de criancas asmaticas que apresentavam crises espaticas severas apesar da utilizacao domiciliar de aerosol de salbutamol. Os pacientes eram seguidos em ambulatorio especializado. Foram avaliadas as tecnicas de utilizacao do inalador, constatando-se 32 erros. Na continuidade do trabalho foi acoplado espacador ao aerosol e realizado treinamento breve. A avaliacao subsequente da tecnica inalatoria observou apenas 1 erro em todo o grupo.

Recovery of Staub Effect and amelioration of insulin secretion defects after glyburide treatment in non-insulin-dependent diabetes.

Staub Effect or improved glucose disposal after repetitive glucose loads does not occur in untreated diabetes. In non-insulin dependent diabetes (NIDDM) there is impaired insulin response to intravenous (i.v.) glucose injection, especially in early insulin release (EIR) and the lesser known post EIR suppression of insulin levels below basal, or acute insulin decrement (AID). To test the ability of a second generation sulfonylurea, glyburide, to affect glucose primed glucose disposal and insulin secretory patterns, sixteen NIDDM male subjects received three hourly intravenous glucose loads while untreated and again after six months of glyburide therapy. After treatment there was a fall of fasting glycemia from 204 +/- 11 to 147 +/- 8 mg/dl (p less than 0.001), of all glucose levels during the i.v. glucose tolerance tests (p less than 0.025) and glycosylated hemoglobin from 8 +/- 0.3% to 7.6 +/- 0.3% (p less than 0.005). Before treatment i.v. glucose disposal (K value) changed very little after successive glucose challenges, but after glyburide all mean K values were higher, and glucose primed glucose disposals were faster after the second (K2) and third (K3) glucose injection than after the first (K1) (p less than 0.025 and p less than 0.01 respectively). In the untreated state, there was higher and significant EIR by the third glucose load, (p less than 0.025) while AID was clearly more pronounced after the second load (p less than 0.001). After glyburide treatment EIR was significantly higher than before in all loads, and mean AID was no longer demonstrable. Insulin summation (S) after successive i.v. loads maintained a stepwise increase both before and after treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Pneumonia pneumococica de evolucao atipica em criancas / Pneumococcal pneumonia with atypical evolution in children

Os autores apresentam 13 casos de pneumonia pneumococica aguda em crianca cujo padrao de apresentacao e evolucao mimetizava pneumonia de outras etiologias, em especial, a pneumonia estafilococica. Tecem comentarios sobre a provavel multiplicidade de fatores envolvidos no comportamento anomalo da infeccao pulmomar nestas criancas e sugerem que, em pelo menos sete delas, os pneumococos eram moderadamente resistentes a penicilina. O Streptococcus pneumoniae tem sido considerado, ao longo do tempo, o agente principal das pneumonias bacterianas agudas, de origem comunitaria, fora do periodo neonatal. Em nosso meio, o pneumococo foi identificado em 18 a 35 por cento das pneumonias bacterianas em criancas, em uma serie de trabalhos , sendo o agente etiologico principal em todas as faixas etarias, com excecao do primeiro mes de vida. Antes do advento da antibioticoterapia, a mortalidade de criancas com pneumonia oscilava de 20 a 50 por cento. Esta alta taxa decaiu, drasticamente, para menos de 1 por cento na era antibiotica. A alta eficacia dos antimicrobianos contra este tipo de infeccao, traduzida por rapida melhora clinica e radiologica dos pacientes e a raridade das complicacoes, nao estimularam muitos estudos sobre o assunto....