Evaluation of Cytopathological Techniques for the Diagnosis of Canine Visceral Leishmaniosis with Lymph Node Samples.

The identification of the parasite in cytological smears of lymph node aspirates is a widely applied technique for the direct diagnosis of Leishmania spp. infection, especially in endemic areas. Although very specific, this method has limited sensitivity, and improving the technique would be highly desirable. This study aimed to evaluate the efficacy of conventional smear cytology (SC), liquid-based cytology (LBC), cell block (CB) stained with haematoxylin and eosin (HE) and immunocytochemistry (ICC), and formalin-fixed paraffin wax-embedded tissue immunohistochemistry (FFPE-IHC) compared with serology and polymerase chain reaction for the diagnosis of canine visceral leishmaniosis (CVL) in lymphoid tissue. The use of a preservative medium and centrifugation for cytological samples reduced the number of unsatisfactory artefacts/background. Moreover, LBC allowed excellent cellular preservation and the application of ancillary techniques, such as CB and ICC. SC was the most accurate morphological diagnostic method (45.0%). CB-ICC alone or associated with SC demonstrated significantly higher sensitivity (70.0% and 72.0%, respectively) when compared with SC alone (34.00%). CB-ICC was found to be more effective in the detection of infected animals with mild clinical signs, similar to FFPE-IHC. The specificity and positive predictive value were similar between all methods. Finally, the detection limit for CB-ICC and SC + CB-ICC was identical (18.46 amastigotes/mm2). Our study suggests that CB-ICC is a promising tool for improvement of the cytopathological diagnosis of CVL and may be applied in routine epidemiological screening.

Potamotrygon motoro stingray venom induces both neurogenic and inflammatory pain behavior in rodents.

Freshwater stingray accidents cause an immediate, intense, and unrelieved pain which is followed by edema, erythema and necrosis formation. Treatment for stingray envenomation is based on administration of analgesic, antipyretic and anti-inflammatory drugs. Concerning pain control, it is prescribed to immerse punctured limb on hot water to alleviate pain. There are no studies demonstrating specific targets on which stingray venom acts to promote pain. Therefore, the aim of this work was to investigate some mechanisms of Potamotrygon motoro venom (PmV) that contribute to nociception induction. Evaluating spontaneous pain behavior in mice injected i.pl. with PmV, it was seen that PmV induced both neurogenic and inflammatory pain. PmV also induced hyperalgesia in both mice and rats, evaluated through electronic von Frey and rat paw pressure test, respectively. Partial inhibition of hyperalgesia was observed in mice treated with cromolyn or promethazine, which indicated that mast cell and histamine via H1 receptor participate in the inflammatory pain. To search for some targets involved in PmVinduced hyperalgesia, the participation of TRPV1, calcium channels, neurokinins, CGRP, and norepinephrine, was evaluated in rats. It was seen that PmV-induced hyperalgesia occurs with the participation of neurokinins, mainly via NK1 receptor, CGRP, and calcium influx, through both P/Q and L-type voltage-dependent calcium channels, besides TRPV1 activation. The data presented herein indicate that PmV causes hyperalgesia in rodents which is dependent on the participation of several neuroinflammatory mediators.

Multilocus family-based association analysis of seven candidate polymorphisms with essential hypertension in an african-derived semi-isolated brazilian population.

Background. It has been widely suggested that analyses considering multilocus effects would be crucial to characterize the relationship between gene variability and essential hypertension (EH). Objective. To test for the presence of multilocus effects between/among seven polymorphisms (six genes) on blood pressure-related traits in African-derived semi-isolated Brazilian populations (quilombos). Methods. Analyses were carried out using a family-based design in a sample of 652 participants (97 families). Seven variants were investigated: ACE (rs1799752), AGT (rs669), ADD2 (rs3755351), NOS3 (rs1799983), GNB3 (rs5441 and rs5443), and GRK4 (rs1801058). Sensitivity analyses were further performed under a case-control design with unrelated participants only. Results. None of the investigated variants were associated individually with both systolic and diastolic BP levels (SBP and DBP, respectively) or EH (as a binary outcome). Multifactor dimensionality reduction-based techniques revealed a marginal association of the combined effect of both GNB3 variants on DBP levels in a family-based design (P = 0.040), whereas a putative NOS3-GRK4 interaction also in relation to DBP levels was observed in the case-control design only (P = 0.004). Conclusion. Our results provide limited support for the hypothesis of multilocus effects between/among the studied variants on blood pressure in quilombos. Further larger studies are needed to validate our findings.

A novel missense mutation p.L76P in the GJB2 gene causing nonsyndromic recessive deafness in a Brazilian family.

Mutations in the GJB2 gene, encoding connexin 26 (Cx26), are a major cause of nonsyndromic recessive hearing loss in many countries. We report here on a novel point mutation in GJB2, p.L76P (c.227C>T), in compound heterozygosity with a c.35delG mutation, in two Brazilian sibs, one presenting mild and the other profound nonsyndromic neurosensorial hearing impairment. Their father, who carried a wild-type allele and a p.L76P mutation, had normal hearing. The mutation leads to the substitution of leucine (L) by proline (P) at residue 76, an evolutionarily conserved position in Cx26 as well as in other connexins. This mutation is predicted to affect the first extracellular domain (EC1) or the second transmembrane domain (TM2). EC1 is important for connexon-connexon interaction and for the control of channel voltage gating. The segregation of the c.227C>T (p.L76P) mutation together with c.35delG in this family indicates a recessive mode of inheritance. The association between the p.L76P mutation and hearing impairment is further supported by its absence in a normal hearing control group of 100 individuals, 50 European-Brazilians and 50 African-Brazilians.

A novel missense mutation p. L76P in the GJB2 gene causing nonsyndromic recessive deafness in a Brazilian family

Mutations in the GJB2 gene, encoding connexin 26 (Cx26), are a major cause of nonsyndromic recessive hearing loss in many countries. We report here on a novel point mutation in GJB2, p.L76P (c.227C>T), in compound heterozygosity with a c.35delG mutation, in two Brazilian sibs, one presenting mild and the other profound nonsyndromic neurosensorial hearing impairment. Their father, who carried a wild-type allele and a p.L76P mutation, had normal hearing. The mutation leads to the substitution of leucine (L) by proline (P) at residue 76, an evolutionarily conserved position in Cx26 as well as in other connexins. This mutation is predicted to affect the first extracellular domain (EC1) or the second transmembrane domain (TM2). EC1 is important for connexon-connexon interaction and for the control of channel voltage gating. The segregation of the c.227C>T (p.L76P) mutation together with c.35delG in this family indicates a recessive mode of inheritance. The association between the p.L76P mutation and hearing impairment is further supported by its absence in a normal hearing control group of 100 individuals, 50 European-Brazilians and 50 African-Brazilians.

Association of polymorphisms at the ADIPOR1 regulatory region with type 2 diabetes and body mass index in a Brazilian population with European or African ancestry.

Association studies between ADIPOR1 genetic variants and predisposition to type 2 diabetes (DM2) have provided contradictory results. We determined if two single nucleotide polymorphisms (SNP c.-8503G>A and SNP c.10225C>G) in regulatory regions of ADIPOR1 in 567 Brazilian individuals of European (EA; N = 443) or African (AfA; N = 124) ancestry from rural (quilombo remnants; N = 439) and urban (N = 567) areas. We detected a significant effect of ethnicity on the distribution of the allelic frequencies of both SNPs in these populations (EA: -8503A = 0.27; AfA: -8503A = 0.16; P = 0.001 and EA: 10225G = 0.35; AfA: 10225G = 0.51; P < 0.001). Neither of the polymorphisms were associated with DM2 in the case-control study in EA (SNP c.-8503G>A: DM2 group -8503A = 0.26; control group -8503A = 0.30; P = 0.14/SNP 10225C>G: DM2 group 10225G = 0.37; control group 10225G = 0.32; P = 0.40) and AfA populations (SNP c.-8503G>A: DM2 group -8503A = 0.16; control group -8503A = 0.15; P = 0.34/SNP 10225C>G: DM2 group 10225G = 0.51; control group 10225G = 0.52; P = 0.50). Similarly, none of the polymorphisms were associated with metabolic/anthropometric risk factors for DM2 in any of the three populations, except for HDL cholesterol, which was significantly higher in AfA heterozygotes (GC = 53.75 +/- 17.26 mg/dL) than in homozygotes. We conclude that ADIPOR1 polymorphisms are unlikely to be major risk factors for DM2 or for metabolic/anthropometric measurements that represent risk factors for DM2 in populations of European and African ancestries.

Association of polymorphisms at the ADIPOR1 regulatory region with type 2 diabetes and body mass index in a Brazilian population with European or African ancestry

Association studies between ADIPOR1 genetic variants and predisposition to type 2 diabetes (DM2) have provided contradictory results. We determined if two single nucleotide polymorphisms (SNP c.-8503G>A and SNP c.10225C>G) in regulatory regions of ADIPOR1 in 567 Brazilian individuals of European (EA; N = 443) or African (AfA; N = 124) ancestry from rural (quilombo remnants; N = 439) and urban (N = 567) areas. We detected a significant effect of ethnicity on the distribution of the allelic frequencies of both SNPs in these populations (EA: -8503A = 0.27; AfA: -8503A = 0.16; P = 0.001 and EA: 10225G = 0.35; AfA: 10225G = 0.51; P < 0.001). Neither of the polymorphisms were associated with DM2 in the case-control study in EA (SNP c.-8503G>A: DM2 group -8503A = 0.26; control group -8503A = 0.30; P = 0.14/SNP 10225C>G: DM2 group 10225G = 0.37; control group 10225G = 0.32; P = 0.40) and AfA populations (SNP c.-8503G>A: DM2 group -8503A = 0.16; control group -8503A = 0.15; P = 0.34/SNP 10225C>G: DM2 group 10225G = 0.51; control group 10225G = 0.52; P = 0.50). Similarly, none of the polymorphisms were associated with metabolic/anthropometric risk factors for DM2 in any of the three populations, except for HDL cholesterol, which was significantly higher in AfA heterozygotes (GC = 53.75 ± 17.26 mg/dL) than in homozygotes. We conclude that ADIPOR1 polymorphisms are unlikely to be major risk factors for DM2 or for metabolic/anthropometric measurements that represent risk factors for DM2 in populations of European and African ancestries.

Reverse transcription-polymerase chain reaction assay for rabies virus detection

Este estudo teve por objetivo implantar um protocolo de amplificação genômica, precedida de transcrição reversa (RT-PCR) para o gene da nucleoproteína do vírus da raiva, para a utilização dessa metodologia em laboratórios onde são realizadas investigações para a detecção do vírus rábico. Foram utilizadas 50 amostras de tecido encefálico de animais (44 bovinos, 5 eqüinos e 1 quiróptero) oriundos do Estado do Rio de Janeiro, positivos por imunofluorescência direta e/ou prova biológica para o vírus rábico. A extração do RNA foi feita a partir da suspensão a 10 por cento em PBS pH7,2 do tecido encefálico utilizando-se a metodologia de TRIzolTM (Life Technologies) e o protocolo de RT-PCR descrito por Heaton et al. (1997), incluindo algumas modificações. Dentre as 50 amostras analisadas, 50 foram positivas pela prova biológica e pela RT-PCR e destas, 49 foram positivas pela imunofluorescência direta. Estes resultados demonstram ser este protocolo de RT-PCR uma metodologia sensível, específica, rápida e extremamente valiosa, podendo ser utilizada como rotina em laboratórios que trabalham no diagnóstico de vírus rábico.

Cisto testicular em carpa comum

São descritas duas formações císticas no testículo direito de um exemplar de carpa comum ( Cyprinus carpio). O cisto maiormedia Sem de diâmetro e o menor 0,5cm, de paredes finas, sendo preenchidos por um líquido amarelo-claro transparente.Microscopicamente, o testículo mostrava um severo processo atrófico ·relacionado com extensa proliferação conjuntiva.

Do urinary mononuclear cells reflect disease activity in lupus nephritis?

Glomerular inflammation is associated with urinary mononuclear cells (UMC) in a number of diseases including IgA nephropathy and glomerulonephritis. We examined UMC from children with lupus nephritis for a number of years to characterize the types of mononuclear cells found in urine and to determine if they were associated with active lupus nephritis. Detailed analysis of UMC by cell counts and by flow cytometry showed that monocytes were the clearly dominant cell type. Evaluation of the smaller number of lymphocytes found in the urine of patients with active lupus nephritis demonstrated a strong predominance of CD8+ lymphocytes, in contrast to the normal CD4+/CD8+ ratio that is found in peripheral blood. The degree of proteinuria strongly correlated with the presence of UMC. The UMC counts decreased as their clinical condition improved as indicated by lower indices of flare. These observations suggest that UMC may be a valuable tool in detecting and monitoring disease activity in patients with severe lupus nephritis. More importantly, this study indicated that both monocytes and cytotoxic CD8+ T cells may play a role in pathogenesis of lupus nephritis.

Differential effects of obesity with and without hyperinsulinemia on plasma lipoprotein(a) concentrations in men.

To determine the association of in vivo concentrations of insulin, obesity, and gender with lipoprotein(a) [Lp(a)] levels, we used a cross-sectional population-based survey of a multistage random sample of the Mexico City adult population. We studied 423 normoglycemic, normotensive subjects from an original sample of 825, comprised of 239 men and 189 women with a mean age of 38.6 years (range, 17 to 90). All subjects were divided into 8 groups according to body mass index, fasting insulin, and gender. Lp(a) concentrations (mg/dL) were similar in obese women with and without high insulin levels (19.9 v 18.6), but hyperinsulinemic obese men had significantly lower Lp(a) levels than normoinsulinemic obese men (7.9 v 29.4). In addition, the proportion of obese men with Lp(a) concentrations of > or = 30 mg/dL was significantly higher in the normoinsulinemic than in the hyperinsulinemic (29.2% v 0.0%). The frequency distribution of Lp(a) levels was shifted to a lower range in hyperinsulinemic men compared with normoinsulinemic men. Our results show that in men, hyperinsulinemic obesity is associated with low Lp(a) levels, while obesity with normoinsulinemia is related to increased Lp(a) concentration. These observations were not found in women. These findings may explain the conflicting results reported by several studies.

[Immune response produced by anti-rabies vaccines in marmosets (Callithrix sp)]. / Resposta imune produzida por vacinas anti-rábicas em sagüis (Callithrix sp).

In order to evaluate the immune response produced by rabies vaccines in new world nonhuman primates, thirty marmosets (Callithrix sp) were divided into five groups of six individuals and submitted to five different antirabies vaccination schemes using two distinct commercially available animal vaccines. The first was produced in suckling mouse brain (Fuenzalida and Palacios), and the second in NIL-2 cell culture. Post-vaccine serological monitoring was carried out periodically. The results showed that the Fuenzalida and Palacios vaccine was not able to protect the animals when using a single dose or even with a booster. But when submitted to a vaccination routine similar to that used for humans, the marmosets showed detectable antibodies, and only one succumbed to rabies after being challenged. In addition, the vaccine produced in NIL-2 cell culture induced high antibody levels in all vaccinated animals and all animals survived the viral challenge.

Effect of the new diagnostic criteria for diabetes in the Mexico City study.

OBJECTIVE: To assess the effect of the new American Diabetes Association (ADA) diagnostic criteria for diabetes in the Mexico City survey. METHODS: The data available from the Mexico City study were included. This cross-sectional survey was conducted from January 1991 to March 1992 and involved 805 adults (20 years of age or older; mean age, 41 years). Survey procedures and results were previously published. Of the residents with diabetes, 74.2% had a previous diagnosis, and 25.8% were diagnosed on the basis of fasting plasma glucose (FPG) levels obtained during the survey. For the current report, these data were reanalyzed on the basis of the new ADA diagnostic criteria. RESULTS: The prevalence of newly diagnosed diabetes with use of the old and the new criteria was 23.1% versus 32.3% in men (P = 0.31) and 27.5% versus 36.3% in women (P = 0.32), respectively. The crude prevalence rate of diabetes increased from 8.7% to 9.3% (P = 0.42), and the age-adjusted rate increased from 10.6% to 11.2% for women (P = 0.64) and from 6.0% to 6.5% for men (P = 0.49). The prevalence of impaired FPG was 4.8%. Those patients with impaired FPG or newly diagnosed diabetes with FPG levels between 126 and 140 mg/dL had a more atherogenic risk profile than did those with normal carbohydrate metabolism. CONCLUSION: On the basis of the 1990 population census in Mexico City, the new ADA criteria will add 28,331 patients with diabetes (6.9%) to an already insufficient medical system. For those patients with impaired FPG or newly diagnosed diabetes on the basis of the new criteria, management should be focused on the control of the diverse and highly prevalent coronary risk factors.

[Risk factors for the development of germ cell tumors in children]. / Factores de riesgo para el desarrollo de tumores de células germinales en niños.

The search for risk factors for development of germ cell tumors (GCT) in children who lived in Mexico City (MC). A protective, observational, case-control study was conducted in children under 15 years of age resident in MC, insurer by the Mexican Institute of Social Security. The study population was selected between January 1st, 1990 and December 31st, 1994. Parents of the children were interviewed with a 230-items precoded questionnaire, validated previously with a pilot study. For analysis were obtained simple frequencies and odds ratios (OR) and 95% confidence interval (95%CI). There were 21 cases and 105 controls. The most significant risk factors were winter conception (OR = 7.6, 95% CI 1.5-39.3; P = 0.007); low parental education level (OR = 2.9, 95% CI 1.1-7.5; P = 0.026); and parental combined dust and electricity exposure before pregnancy (OR = 26, 95% CI 2.28-1291.86; P = 0.0007). during (OR8.58, 95% CI 0.89-106.55; P = 0.041) and after pregnancy (OR = 9.66, 95% CI 0.99-120.22; P = 0.027). There was a protective effect with repetitive infections during infancy. In conclusion, Winter conception is in accordance with infectious etiology theory of GCT development. The low parental education level and the combined exposure to dust and electricity are very important. The protective effect of repetitive infections and other factors make necessary more epidemiologic studies in this field.

Prevalence of high blood pressure and associated coronary risk factors in an adult population of Mexico City.

BACKGROUND: Several studies have addressed arterial hypertension prevalence in Mexico. However, few include an analysis of other types of hypertension and their associated risk factors. The present work describes the prevalence of high blood pressure (HBP), isolated systolic hypertension (ISH) and diastolic hypertension (DH) and their association to certain risk factors of cardiovascular disease in an adult population of Mexico City. METHODS: A cross-sectional study was performed on 825 subjects aged between 20 and 90 years, selected by multistage cluster sampling. HBP was diagnosed by previous history if systolic blood pressure was > or = 140 mmHg and/or diastolic blood pressure > or = 90 mmHg. The measurements taken included body mass index, waist-to-hip ratio, systolic and diastolic blood pressure, levels of insulin, glucose, trigylycerides, total cholesterol, high and low density lipoprotein cholesterol, and lipoprotein(a). RESULTS: Prevalence adjusted by age for HBP was 19.4%, for ISH, 4.7%, and for DH, 4.1%. Age had an important influence on HBP and ISH with a highly significant X2t. The profile of metabolic variables was modified according to sex and type of hypertension. Thus, in DH, metabolic variables were more affected than in other types of hypertension. CONCLUSIONS: Results in HBP prevalence in the present study were lower than in other surveys performed in Mexico. It must be noted, however, that much care should be taken to choose the strategy of subject selection, since results of the prevalence of a disease depend on it to a great extent. The ISH and DH and their association to risk factors must be studied thoroughly because they constitute different clinical entities.

Incidence of malignant neoplasms in children attending Social Security Hospitals in Mexico City.

An increase in neoplasms in Mexican children has been reported. In 1991, the incidence in children from Mexico City (MC) was 70 (x 10(6) child/year), although this rate might be underestimated. The aim of the present study was to estimate the incidence of malignant neoplasms in children resident in MC attending Social Security (SS) hospitals. This study was a retrospective hospital survey. All records of childhood malignant neoplasms diagnosed between 1992 and 1993 in the two SS hospitals which attend childhood neoplasms in MC were reviewed. Histopathological diagnoses were reevaluated and incidence rates (x 10(6) child/ year) in terms of age, sex, and place of residence were estimated. A total of 667 cases were found for the period of study, of which 199 corresponded to residents of MC. The neoplasms with highest prevalence were leukemias (39.2%), lymphomas (17.6%), and central nervous system tumors (12.6%). A general incidence of 94.3 was found, which was highest in children under 5 years of age. Leukemias had an incidence of 36.4, lymphomas of 15.2, and central nervous system tumors of 12.0. Prevalence was higher in boys (male/female ratio of 1.6). As for the place of residence, the highest incidence corresponded to children living in the southern areas of MC. Eighty percent of the leukemias were acute lymphoblastic, while 54% of solid neoplasms were classified as stages III and IV. In conclusion, the incidence of malignant neoplasms in children resident in MC treated at SS hospitals is consistent with that found worldwide, and also with the Latin American pattern.

Expansion of CD3+CD56+ lymphocytes correlates with induction of cytotoxicity by interleukin-2 gene transfer in human breast tumor cultures.

BACKGROUND: Mice immunized with murine mammary carcinoma cells genetically engineered to secrete interleukin-2 (IL-2) are rendered resistant to subsequent challenge with unmodified tumor cells, and in the case of mice bearing established tumors, the rate of development of pulmonary metastases is reduced. Despite these encouraging animal results, little is known about the induction of antitumor immunity by IL-2 gene transfer in human breast cancer. METHODS: Adenovirally mediated IL-2 gene transfer was performed in 12 tumor fragment cultures established from seven primary breast cancers. Autologous tumor infiltrating lymphocytes (TILs) or peripheral blood mononuclear cells (PBMCs) were cocultured with transduced tumor fragments, and changes in phenotype and cytotoxicity were measured. RESULTS: IL-2 was never detectable in the untransduced cultures, but it peaked at 5.0-1,324.8 ng/ml in the transduced cultures. Lymphocyte counts declined in all untransduced cultures, but they increased two- to sevenfold in four transduced cultures. CD4:CD8 ratios decreased from a mean of 2.11 at baseline to 1.27 after stimulation in coculture (p = 0.03). Expansion of lymphocytes expressing the natural killer cell phenotype (CD3-CD56+) occurred in only one culture, but the CD3+CD56+ population increased in four of six cultures. Lymphocytes from four of 10 cocultures generated significant cytotoxicity against allogeneic breast cancer cells. Induction of cytotoxicity correlated with expansion of the CD3+CD56+ phenotype (R2 = 0.805, p = 0.02). CONCLUSIONS: IL-2 gene expression by human breast cancer causes expansion of CD3+CD56+ cytotoxic-lymphocytes. This phenotype is consistent with that of a non-major histocompatibility complex (MHC)-restricted cytokine induced killer cell population previously described.

[Contribution to the anatomical study of medial arm flap]. / Contribuição ao estudo anatômico do retalho medial do braço.

The reports of transfer of the medial arm flap have been inconsistent, and confusion remains about its anatomy and use. The advantages of this donor site, however, justify new researches about the vascular anatomy of the flap. In this study, the vascular supply to the medial side of the arm was analysed in fifteen fresh cadaver dissections. The methods, the anatomical bases, and the technique of elevation of the flap were detailed. Methylene blue injection studies showed the extent of the medial arm flap. The authors found that, in most cases, the superior ulnar collateral artery was the main vessel of this flap, giving cutaneous branches to medial arm skin. The diameter of the superior ulnar collateral artery was measured and considered adequate for microsurgical transfer. If no major cutaneous branches are noted from superior ulnar collateral artery, then a significant direct cutaneous branch from the brachial artery will be always present. With this in mind, the dissection must initially proceed carefully, preserving all vessels that may contribute to cutaneous perfusion. The authors conclude that with knowledge of its anatomical variations, the medial arm flap is a plausible reconstructive option.

Lipoprotein(a) and cardiovascular risk in adult Mexicans.

A cross-epidemiologic study concerning cardiovascular risk factors in a random sample population of Mexico City was carried out in 1991 and 1992. Lipoprotein(a) (Lp(a)) levels in 404 men and 311 women aged 20-90 years were determined by enzyme-linked immunoassay on fasting plasma. Men and women showed similar age-adjusted Lp(a) values. For the whole population the median Lp(a) was 6.9 mg/dL and the prevalence of high Lp(a) (> 30 mg/dL) was 14%. Small positive correlations between Lp(a) and plasma cholesterol (rs = 0.16) and low density lipoprotein cholesterol (LDL-C) (rs = 0.21), and a negative one with insulin (rs = -0.13) were found. In a multiple regression analysis, insulin and LDL-C were the variables that best explained the variation of Lp(a) in our sample. Our data show that Lp(a) in our population is similar to that found in other populations. An association of Lp(a) with myocardial infarction was observed (high Lp(a) was seen in 33% of atherosclerotic individuals versus 14% in healthy subjects) but did not reach statistical significance.