A semi-automated microscopic image analysis method for scoring Ki-67 nuclear immunostaining.

Nuclear proliferation marker MIB-1 (Ki-67) immunohistochemistry (IHC) is used to examine tumor cell proliferation. However, the diagnostic or prognostic value of the Ki-67 nuclear staining intensity and location, defined as nuclear gradient (NG), has not been assessed. This study examined the potential association between Ki-67 NG and cell cycle phases and its effect on the prognosis of pulmonary typical carcinoid (PTC) tumors. We propose a method for classifying the NG of Ki-67 during the cell cycle and compare the results between PTC, pulmonary adenocarcinoma (PAD), and breast ductal carcinoma (BDC). A literature review and objective analysis of IHC-stained paraffin sections were used to determine the Ki-67 labeling index and composed a stratification of the NG into NG1, NG2, and NG3/4 categories. A semi-automated image analysis protocol was established to determine the Ki-67 NG in PTC, PAD, and BDC. High intraobserver consistency and moderate interobserver agreement were achieved in the determination of Ki-67 NG in tumor specimens. NG1 and NG2 were lower in PTC than in PAD and BDC. Cox multivariate analysis of PTC after adjusting for age and number of metastatic lymph nodes showed that Ki-67 NG1 and NG2 significantly predicted clinical outcomes. The semi-automated method for quantification of Ki-67 nuclear immunostaining proposed in this study could become a valuable diagnostic and prognostic tool in PTC.

Assessing the impact of the presence of pharmacists in a clinical department of a Lebanese hospital.

BACKGROUND: Clinical pharmacy services are still in their very early implementation stages in Lebanon. The objective of this pilot study was to evaluate the impact of clinical pharmacist's presence at the infectious diseases department of Hôtel-Dieu de France University Hospital of Beirut (HDF) and to evaluate the acceptance of pharmacist's interventions by healthcare providers. MATERIAL AND METHODS: A 21-month prospective analysis was conducted, including 240 hospitalized patients in the infectious diseases department of HDF and 475 interventions performed by the pharmacist. A clinical pharmacist and pharmacy residents were present for 1 to 2hours/day in the ward. A pharmaceutical care plan was established and used to document patients' problems and pharmacist's interventions. Main criteria analyzed were: types and frequencies of pharmaceutical problems detected, types of pharmaceutical interventions performed, their acceptance by the prescribers and/or nurses, and factors affecting the interventions and their acceptance. RESULTS: The most frequent pharmaceutical problem detected was incorrect dosage and the three most frequent interventions performed by the pharmacist were stop/start/substitute a drug, change drug dosage/or daily distribution, and change administration time. The acceptance was the highest for I.TIM (change drug administration time) and the lowest for I.FOL (request a lab test/exam/clinical follow-up). DISCUSSION AND CONCLUSION: Even a short daily pharmacist's presence is an added value in inpatient care at the infectious diseases department of Hôtel-Dieu de France University Hospital. Areas of improvement are a better communication between the pharmacist and the prescribers, a direct contact between pharmacist and patient and a longer presence of the clinical pharmacist in the clinical department.

Monitoring non-small cell lung cancer progression and treatment response through hyaluronic acid in sputum.

We evaluated whether hyaluronan (HA) levels in the sputum could be used as a noninvasive tool to predict progressive disease and treatment response, as detected in a computed tomography scan in non-small cell lung cancer (NSCLC) patients. Sputum samples were collected from 84 patients with histological confirmation of NSCLC, 33 of which were in early-stage and 51 in advanced-stage disease. Patients received systemic chemotherapy (CT) after surgery (n=36), combined CT and immunotherapy (IO) (n=15), or targeted therapy for driver mutation and disease relapse (N=4). The primary end-point was to compare sputum HA levels in two different concentrations of hypertonic saline solution with overall survival (OS) and the secondary and exploratory end-points were radiologic responses to treatment and patient outcome. Higher concentrations of HA in the sputum were significantly associated to factors related to tumor stage, phenotype, response to treatment, and outcome. In the early stage, patients with lower sputum HA levels before treatment achieved a complete tumor response after systemic CT with better progression-free survival (PFS) than those with high HA levels. We also examined the importance of the sputum HA concentration and tumor response in the 51 patients who developed metastatic disease and received CT+IO. Patients with low levels of sputum HA showed a complete tumor response in the computed tomography scan and stable disease after CT+IO treatment, as well as a better PFS than those receiving CT alone. HA levels in sputum of NSCLC patients may serve as a candidate biomarker to detect progressive disease and monitor treatment response in computed tomography scans.

Determinants of treatment of mental disorders in Lebanon: barriers to treatment and changing patterns of service use.

AIMS: To investigate for the first time the determinants and barriers of seeking help for mental disorders in the Arab world based on a national study: Lebanese Evaluation of the Burden of Ailments and Needs Of the Nation (L.E.B.A.N.O.N). METHODS: A nationally representative (n = 2857) and multistage clustered area probability household sample of adults ≥18 years and older was assessed for lifetime and 12 months mental disorders using the Composite International Diagnostic Interview. In addition, detailed information was obtained on help- seeking behaviour and barriers to treatment. RESULTS: In total, 19.7% of the Lebanese with mental disorders sought any type of treatment: 91% of those who sought treatment did so within the health sector. Severity and perceived severity of disorders predicted seeking help, the highest being for panic disorder. The greatest barrier to seek help was low perceived need for treatment (73.9%). Stigma was reported to be a factor only in 5.9% of those who thought about seeking treatment. Eighty per cent of the Lebanese reported they would not be embarrassed if friends knew they were seeking help from a professional. CONCLUSIONS: A small fraction of Lebanese seek help for their mental health problems: female gender, higher education and income are predictors of positive attitudes to help seeking. Severity and recognition of disorders, more than stigma, to get treatment seem to be the most important factors in determining help seeking. The findings underscore the importance of helping the public recognise mental health disorders.

Detection of sputum cofilin-1 as indicator of malignancy.

Cofilin-1 (CFL1), a small protein of 18 kDa, has been studied as a biomarker due to its involvement in tumor cell migration and invasion. Our aim was to evaluate CFL1 as an indicator of malignancy and aggressiveness in sputum samples. CFL1 was analyzed by ELISA immunoassay in the sputum of 73 lung cancer patients, 13 cancer-free patients, and 6 healthy volunteers. Statistical analyses included ANOVA, ROC curves, Spearman correlation, and logistic regression. Sputum CFL1 levels were increased in cancer patients compared to cancer-free patients and volunteers (P<0.05). High expression of sputum CFL1 was correlated to T4 stage (P=0.01) and N stage (P=0.03), tobacco history (P=0.01), and squamous cell carcinoma histologic type (P=0.04). The accuracy of sputum CFL1 in discriminating cancer patients from cancer-free patients and healthy volunteers were 0.78 and 0.69, respectively. CFL1 at a cut-off value of 415.25 pg/mL showed sensitivity/specificity of 0.80/0.70 in differentiating between healthy volunteers and cancer patients. Sputum CFL1 was also able to identify cancer-free patients from patients with lung cancer. The AUC was 0.70 and, at a cut-off point ≥662.63 pg/mL, we obtained 60% sensitivity and 54% specificity. Logistic regression analysis controlled for tobacco history, histologic types, and N stage showed that cancer cell-associated CFL1 was an independent predictor of death. Smoker patients with squamous cell carcinoma, lymph node metastasis and sputum CFL1>1.475 pg/mL showed augmented chance of death, suggesting lung cancer aggressiveness. CFL1 presented diagnostic value in detecting lung cancer and was associated to tumor aggressiveness.

A semi-automated microscopic image analysis method for scoring Ki-67 nuclear immunostaining

Nuclear proliferation marker MIB-1 (Ki-67) immunohistochemistry (IHC) is used to examine tumor cell proliferation. However, the diagnostic or prognostic value of the Ki-67 nuclear staining intensity and location, defined as nuclear gradient (NG), has not been assessed. This study examined the potential association between Ki-67 NG and cell cycle phases and its effect on the prognosis of pulmonary typical carcinoid (PTC) tumors. We propose a method for classifying the NG of Ki-67 during the cell cycle and compare the results between PTC, pulmonary adenocarcinoma (PAD), and breast ductal carcinoma (BDC). A literature review and objective analysis of IHC-stained paraffin sections were used to determine the Ki-67 labeling index and composed a stratification of the NG into NG1, NG2, and NG3/4 categories. A semi-automated image analysis protocol was established to determine the Ki-67 NG in PTC, PAD, and BDC. High intraobserver consistency and moderate interobserver agreement were achieved in the determination of Ki-67 NG in tumor specimens. NG1 and NG2 were lower in PTC than in PAD and BDC. Cox multivariate analysis of PTC after adjusting for age and number of metastatic lymph nodes showed that Ki-67 NG1 and NG2 significantly predicted clinical outcomes. The semi-automated method for quantification of Ki-67 nuclear immunostaining proposed in this study could become a valuable diagnostic and prognostic tool in PTC.

Monitoring non-small cell lung cancer progression and treatment response through hyaluronic acid in sputum

We evaluated whether hyaluronan (HA) levels in the sputum could be used as a noninvasive tool to predict progressive disease and treatment response, as detected in a computed tomography scan in non-small cell lung cancer (NSCLC) patients. Sputum samples were collected from 84 patients with histological confirmation of NSCLC, 33 of which were in early-stage and 51 in advanced-stage disease. Patients received systemic chemotherapy (CT) after surgery (n=36), combined CT and immunotherapy (IO) (n=15), or targeted therapy for driver mutation and disease relapse (N=4). The primary end-point was to compare sputum HA levels in two different concentrations of hypertonic saline solution with overall survival (OS) and the secondary and exploratory end-points were radiologic responses to treatment and patient outcome. Higher concentrations of HA in the sputum were significantly associated to factors related to tumor stage, phenotype, response to treatment, and outcome. In the early stage, patients with lower sputum HA levels before treatment achieved a complete tumor response after systemic CT with better progression-free survival (PFS) than those with high HA levels. We also examined the importance of the sputum HA concentration and tumor response in the 51 patients who developed metastatic disease and received CT+IO. Patients with low levels of sputum HA showed a complete tumor response in the computed tomography scan and stable disease after CT+IO treatment, as well as a better PFS than those receiving CT alone. HA levels in sputum of NSCLC patients may serve as a candidate biomarker to detect progressive disease and monitor treatment response in computed tomography scans.

Ceftriaxone therapy of bacterial meningitis: cerebrospinal fluid concentrations and bactericidal activity after intramuscular injection in children treated with dexamethasone.

Antibiotic therapy is administered intravenously to children with bacterial meningitis to achieve the highest possible blood and cerebrospinal fluid (CSF) concentrations. However, intravenous access for the entire duration of therapy may be difficult in some children. Intramuscular therapy offers a more versatile option; however, CSF concentrations and bactericidal activity following im injection in children concurrently treated with dexamethasone have not been studied. We prospectively evaluated 37 children given an im dose of ceftriaxone on either the 3rd, 6th or 9th day of antibiotic therapy while receiving dexamethasone for the first 4 days of treatment. All children were required to have normal peripheral perfusion at the time of im injection. Four to 6 hours after im injection CSF was obtained. The average age of study patients was 28 months; Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae type b were responsible for 95% of all infections. All children studied had detectable CSF ceftriaxone concentrations, with mean (+/- SD) concentrations (microgram/ml) on Days 3, 6 and 9 of therapy of 5.7 +/- 5.5 (n = 12), 5.2 +/- 5.0 (n = 14) and 2.0 +/- 2.6 (n = 10), respectively. All CSF bactericidal titers for N. meningitidis, S. pneumoniae and H. influenzae type b, regardless of day of im injection, were > or = 1:64. Intramuscular ceftriaxone therapy of bacterial meningitis may be a reasonable therapeutic option for the convalescing child with good peripheral perfusion.

The antimeningococcal vaccine VAMENGOC B-C induced poor serum and salivary antibody response in young Brazilian children.

In 1989 about 2.3 million Brazilian children received the antimeningococcal vaccine VAMENGOC B-C (Havana, Cuba). We evaluated the serum and secretory immune response of vaccinated children by enzyme-linked immunosorbent assay with outer membrane complex antigens. Western blotting and bacterial adherence inhibition assays with human buccal epithelial cells were performed with some of the samples. Serum and salivary antibody concentrations to Neisseria meningitidis Group B of vaccinated children < 4 years old were not significantly higher than those of nonvaccinated children, as observed in convalescing patients used as positive controls. Older children (4 to 6 years old) presented a slight increase in antibody OD indexes. Sera and saliva from vaccinated children showed a weak reaction with meningococcal antigen by Western blotting and were unable to inhibit significantly the adherence of N. meningitidis B to buccal epithelial cells. These data suggest that this vaccine induced a poor serum and salivary antibody response in the population studied.

Human serum beta2-microglobulin levels: correlation with total serum IgG and placental IgG transfer in HIV-infected and non-HIV infected individuals.

The acquisition of maternal immunoglobulin G (IgG) is fundamental to the immune defence of the neonate. The receptor responsible for IgG transfer across the human placenta has also been implicated in the maintenance of IgG levels in the circulation. beta2-microglobulin is part of the Fc receptor (FcR) that has recently been purified from the human placenta. In HIV infection, increasing serum levels of total IgG and beta2-microglobulin are observed as the disease progresses. Herein, we have investigated the correlation between beta2-microglobulin and total serum IgG levels in HIV-seropositive mothers and their term neonates (HIV group, n = 37), as well as in HIV-seronegative mothers and their term neonates (control group, n = 50). Serum maternal beta2-microglobulin was directly correlated with total serum IgG levels in HIV-infected mothers (r = 0.58; P = 0.0002), but not in healthy HIV-seronegative mothers (r = -0.20; P = 0.16). Maternal serum beta2-microglobulin was also inversely correlated with placental antibody transfer of total IgG in mother-newborn pairs from the HIV group (r = 0.38; P = 0.02), but not from the control group (r = 0.15. P = 0.31). These results seem to indicate that, in HIV infection, elevated serum beta2-microglobulin levels could be involved in maintenance of abnormally high total serum IgG concentrations; by interfering with the binding of IgG to Fc receptors at the maternal-fetal interface, they might also reduce IgG transfer. By contrast, in normal non-HIV infected individuals, serum beta2-microglobulin levels do not appear implicated in regulation of these two phenomena.

[Prevalence of Haemophilus influenzae resistant to ampicillin, cefaclor, cefotaxime, chloramphenicol and cotrimoxazol isolated from laboratories in the city of São Paulo, Brazil]. / Prevalência de Haemophilus influenzae resistentes a ampicilina, cefaclor, cefotaxime, cloranfenicol e cotrimoxazol isolados de laboratórios na cidade de São Paulo.

OBJECTIVE: To determine Haemophilus influenzae resistant to ampicillin and other antibiotics isolated from different clinical specimens. METHODS: Isolates of H. influenzae were identified by culture with V and X factors and the aminolevulinic test. Nitrocefin was used to detect beta-lactamase (betaLac) production isolates were tested for antimicrobial susceptibility by disc diffusion and agar dilution methods. Serotype b was assessed by slide co-agglutination. RESULTS: From 245 H. influenzae identified, 155 were tested for serotype b, 28% (43/155) of which were positive. The global rate of beta-lactamase-positive isolates was 9% (22/245). Resistance was similar among serotype b (11.6%) and non-type b H. influenzae (9.8%) (p>0.05). No difference on betaLac production was found according to specimen's origin or the patients' age. Resistances to other antibiotics (by agar dilution and disc diffusion method, respectively) were: chloramphenicol 3.3 to 7.1%; cefaclor: 1.6 to 3.9% and cotrimoxazol: 9.1 to 10.5%. No resistance to cefotaxime has been detected; 63% (5/8) beta-Lac-positive isolates by agar dilution showed also resistance to chloramphenicol, compared to 3% (4/118) in the ss-Lac-negative group (p<0.001). CONCLUSIONS: H. influenzae ampicillin-resistance has shown to be lower than other hospital-based-studies in São Paulo, and comparable to rates found in healthy carriers. The association between ampicillin and chloramphenicol resistance was significant: where this pattern is frequently found, the initial therapy for severe H. influenzae infections - like meningitis - should include a third generation cephalosporin.

[BCG test reaction in previously BCG Moreaux-Rio de Janeiro-vaccinated healthy children]. / BCG-teste em crianças sadias previamente vacinadas com a BCG Moreaux-Rio de Janeiro.

BCG-test reaction was carried out in 91 non-tuberculous children between 2 and 11 years old, who had received BCG vaccination during the first six months of life to critically evaluate the usefulness of this test in Pediatric practice. The BCG-test profile of the vaccinated group was shown to have a unimodal distribution, varying from 3 to 17 millimeters, with a median of 6 millimeters. It was demonstrated that the degree of reaction decreases with time. Nevertheless, false-positives could be expected in a significant percentage when reactions larger than 10 millimeters are considered as a cutoff. The specificity can be increased using only reactions with diameter equal or greater than 15 millimeters as an indicator of tuberculous disease.

[Comparison of seroconversion rates between well-nourished and malnourished children after measles immunization at nine months of age]. / Viragem sorológica após a vacinação contra o sarampo, realizada em crianças eutróficas e desnutridas, aos nove meses de idade.

The study was based on activities developed at Embu, SP, between October 1989 and June 1990. Its purpose was to study serological turning after child vaccination against measles at the age of nine months. Two groups were compared, both within the same age limits. Group number I included eutrophic children and group number II included undernourished children. Gomes criteria was used to evaluate the childrens nutritional state. Antibodies (AB) dosage was done through hemagglutination inhibition (HI) and ELISA. These two laboratory methods were also checked regarding its sensibility. Out of 130 children studied, 80 could be evaluated. From this total, 56 (70%) belonged to group I and 24 (30%) belonged to group II. When the ELISA method was used, a significantly higher seroconversion percentage (P < 0.05 or 5%) was found among children belonging to group II. This percentage was not detected when the HI method was used.

Detection of sputum cofilin-1 as indicator of malignancy

Cofilin-1 (CFL1), a small protein of 18 kDa, has been studied as a biomarker due to its involvement in tumor cell migration and invasion. Our aim was to evaluate CFL1 as an indicator of malignancy and aggressiveness in sputum samples. CFL1 was analyzed by ELISA immunoassay in the sputum of 73 lung cancer patients, 13 cancer-free patients, and 6 healthy volunteers. Statistical analyses included ANOVA, ROC curves, Spearman correlation, and logistic regression. Sputum CFL1 levels were increased in cancer patients compared to cancer-free patients and volunteers (P<0.05). High expression of sputum CFL1 was correlated to T4 stage (P=0.01) and N stage (P=0.03), tobacco history (P=0.01), and squamous cell carcinoma histologic type (P=0.04). The accuracy of sputum CFL1 in discriminating cancer patients from cancer-free patients and healthy volunteers were 0.78 and 0.69, respectively. CFL1 at a cut-off value of 415.25 pg/mL showed sensitivity/specificity of 0.80/0.70 in differentiating between healthy volunteers and cancer patients. Sputum CFL1 was also able to identify cancer-free patients from patients with lung cancer. The AUC was 0.70 and, at a cut-off point ≥662.63 pg/mL, we obtained 60% sensitivity and 54% specificity. Logistic regression analysis controlled for tobacco history, histologic types, and N stage showed that cancer cell-associated CFL1 was an independent predictor of death. Smoker patients with squamous cell carcinoma, lymph node metastasis and sputum CFL1>1.475 pg/mL showed augmented chance of death, suggesting lung cancer aggressiveness. CFL1 presented diagnostic value in detecting lung cancer and was associated to tumor aggressiveness.

[Bacterial meningitis - diagnosis and treatment] / Meningites bacterianas - diagnóstico e conduta.

OBJECTIVE: To present new concepts on diagnosis and treatment of bacterial meningitis regarding etiologic agents, pathophysiology and options about antimicrobial, antiinflammatory and supportive therapy. METHODS: Bibliographic review from MEDLINE data including articles published during the last ten years. One classic article published before this period and chapters of textbooks on infectious diseases were also included. RESULTS: Initial empirical antibiotic therapy is chosen according to probable etiologic agents for the age group. In the CSF microbiological analysis, the gram stains can reveal bacteria in 50% to 80% of the cases and the culture in nearly 85%. The tests for detection of bacterial antigens are useful for the diagnosis but they present low sensitivity. The most common agents during the neonatal period continue to be E. coli, Streptococcus B and L. monocytogenes. Beyond this period, the incidence of meningitis by Haemophilus influenzae b had a significant decrease after the introduction of conjugate vaccines. However, S. pneumoniae and N. meningitidis continue to be frequent agents. Currently, the thirdgeneration cephalosporins, ceftriaxone or cefotaxime, are the antibiotic therapy of choice. They are used with ampicillin up to two months of life and alone beyond this age. Dexamethasone has showed to be effective in reducing the inflammatory response and the sequelae, mainly the auditory sequelae. The fluid restriction doesn't offer advantages in the supportive care. CONCLUSIONS: Early diagnosis and prompt treatment are related to good outcome. The new insights on pathophysiology, the new antibiotics and the increasing bacterial resistance have determined changes in treatment.

[Acquired toxoplasmosis] / Toxoplamose adquirida.

OBJECTIVE: The aim of this review is to present some important aspects of acquired toxoplasmosis to the pediatric practitioner. METHOD: All the articles about acquired toxoplasmosis published during the last decade and indexed in the Index Medicus were revised. From each one, interesting aspects were critically selected. RESULTS: We describe aspects of the epidemiology, pathogenesis, clinical manifestations, diagnosis, treatment and prevention related to acquired toxoplasmosis. CONCLUSION: The content of this article may facilitate the management of patients suspected of having acquired toxoplasmosis.