ASCITIC AND SERUM LEVELS OF TUMOR BIOMARKERS (CA 72-4, CA 19-9, CEA AND CA 125) IN DISCRIMINATION OF CAUSE OF ASCITES: A PROSPECTIVE STUDY.

BACKGROUND: The role of ascitic and serum levels of various tumour biomarkers in the discrimination of cause of ascites is not well established. OBJECTIVE: To evaluate the role of serum and ascitic levels of tumor biomarkers (CA 72-4, CA 19-9, CEA and CA 125) in discrimination of cause of ascites. METHODS: A prospective study was conducted in consecutive patients presenting with ascites. Serum and ascitic levels of CA 19-9, CA 125, CA 72-4 and carcinoembryonic antigen (CEA) were determined at the presentation. The patients with cirrhotic ascites, tuberculous peritonitis (TBP) and peritoneal carcinomatosis (PC) were eventually included in analysis. RESULTS: Of the 93 patients (58 males, mean age 47 years) included, the underlying cause was cirrhosis in 31, PC in 42 and peritoneal tuberculosis in 20. The best cutoff for discriminating benign and malignant ascites for serum CEA, CA 19-9 and CA 72-4 were 6.7 ng/mL, 108 IU/mL and 8.9 IU/mL, respectively. The best cutoff for discriminating benign and malignant ascites for ascitic CA 125, CEA, CA 19-9 and CA 72-4 were 623 IU/mL, 8.7 ng/mL, 33.2 IU/mL and 7 IU/mL, respectively. CONCLUSION: The performance of single biomarker for the prediction of underlying PC is low but a combination of serum CA 19-9 and CA 72-4 best predicted the presence of peritoneal carcinomatosis.

Linoleic and oleic acids enhance cell migration by altering the dynamics of microtubules and the remodeling of the actin cytoskeleton at the leading edge.

Fatty acids (FA) have a multitude of biological actions on living cells. A target of their action is cell motility, a process of critical importance during cancer cell dissemination. Here, we studied the effect of unsaturated FA on ovarian cancer cell migration in vitro and its role in regulating cytoskeleton structures that are essential for cell motility. Scratch wound assays on human ovary cancer SKOV-3 cell monolayers revealed that low doses (16 µM) of linoleic acid (LA, 18:2 ω6) and oleic acid (OA; 18:1 ω9) promoted migration, while α-linolenic acid (ALA, 18:3 ω3), showed a migration rate similar to that of the control group. Single cell tracking demonstrated that LA and OA-treated cells migrated faster and were more orientated towards the wound closure than control. In vitro addition of those FA resulted in an increased number, length and protrusion speed of filopodia and also in a prominent and dynamic lamellipodia at the cell leading edge. Using time-lapse video-microscopy and FRAP we observed an increase in both the speed and frequency of actin waves associated with more mobile actin and augmented Rac1 activity. We also observed that FA induced microtubule-organizing center (MTOC)-orientation towards the cell front and affected the dynamics of microtubules (MT) in the direction of cell migration. We propose that environmental cues such as OA and LA present in ascitic fluid, should be taken into account as key factors for the regulation of cell migration.

P16 as a marker of carcinoma in effusions and peritoneal washing.

BACKGROUND: Considering the potential of p16 as a marker for diagnosis, prognosis and therapeutic response, the aim of this study was to assess its presence, via immunocytochemistry, in metastatic carcinoma of different primary sites and histological types obtained from effusions and peritoneal washings. A total of 118 samples including 85 of metastatic carcinoma and 33 samples of benign effusion/peritoneal washing were prepared by the plasma/thromboplastin method. Immunocytochemistry reactions were performed on cell block sections using antibodies against p16, claudin-4, MOC-31, calretinin, HBME and CD68. RESULTS: P16 overexpression was observed in 88.23% of all carcinoma samples. All cervix adenocarcinoma samples showed p16 overexpression. Overexpression in adenocarcinomas of ovary, lung and breast was observed in 93.75, 93.10 and 75% of the samples, respectively. Overexpression was observed in all different histological types analyzed: small cell carcinoma (lung), squamous cell carcinoma (cervical) and urothelial carcinoma (bladder). The specificity of p16 for carcinoma detection was of 96.96%. CONCLUSION: Overexpression of p16 was observed in most metastatic carcinoma, from different primary sites and histological types, obtained from effusions and peritoneal washings. Due to its high frequency of overexpression in metastatic carcinoma, p16 may play a possible role in tumor progression and it may be considered as a complementary diagnostic marker depending on histological type and primary site of carcinoma.

Peritoneal fluid collection in healthy cattle and evaluation of changes in cell morphology during storage in refrigerated and non-refrigerated samples / Colheita do fluido peritoneal de bovinos sadios e verificação das alterações na morfologia celular quando mantido refrigerado ou à temperatura ambiente

This study aimed to evaluate the appropriate sites of abdominocentesis for peritoneal fluid collection in cattle and to investigate the time of cell viability in vitro, comparing three methods of sample conservation. Twenty-one healthy cattle (19 females and 2 males) were subjected to a laparocentesis procedure to obtain peritoneal fluid, with punctures in three defined sites: left cranial, right cranial, and right caudal. The total peritoneal fluid collected was divided into three aliquots and maintained under three preservation conditions: room temperature (26°C), refrigeration (4°C), and room temperature (26°C) with the addition of 1µL of 10% formaldehyde per 1mL of peritoneal fluid. The peritoneal fluid analysis performed immediately after collection consisted of: physical examination (color, appearance, volume, and specific gravity), biochemical measures (pH, total protein, fibrinogen, creatinine, and glucose), and cellularity (total and differential counts). The determination of proteins and the examination of cells were repeated in each separate aliquot at two, four, six, and eight hours after harvest. Data were analyzed through repeated measures ANOVA or Friedman test. The harvest was productive in 67% of cattle. The left cranial and the right cranial puncture sites were the most appropriate. Peritoneal fluid analyzed after collection, the total protein concentration ranged from 1.4 to 3.6g/dL, and number of leukocytes ranged from 54 to 1,322 cells/µL; 60 to 95% of leukocytes were lymphocytes. The protein concentration decreased, but the absolute values of leukocytes, lymphocytes, and segmented neutrophils did not change up to eight hours after collection, independent of the maintenance method. Cell lysis was delayed by cooling, and the addition of formaldehyde did not help preserve the integrity of cellular morphology...(AU)

O estudo teve como objetivo avaliar os locais adequados de laparocentese para a colheita de fluido peritoneal de bovinos e estabelecer o tempo de viabilidade celular in vitro, comparando três métodos de conservação. Vinte e um bovinos hígidos (19 fêmeas e 2 machos) foram submetidos ao procedimento de laparocentese para obtenção de fluido peritoneal, com punção em três pontos definidos: cranial esquerdo, cranial direito e caudal direito. O volume total do líquido peritoneal foi dividido em três alíquotas mantidas sob três métodos de conservação: temperatura ambiente (26°C); refrigeração (4°C); e temperatura ambiente (26°C) com adição de 1µL de formol 10% para cada 1mL de líquido peritonial. A análise do líquido peritoneal realizada imediatamente após sua obtenção consistiu em: exames físico (cor, aspecto, volume e densidade); bioquímicos (pH, proteína total, fibrinogênio, creatinina e glicose); e da celularidade (contagens total e diferencial). A determinação de proteínas e o exame da celularidade foram repetidos, em cada alíquota separada, as duas, quatro, seis e oito horas após a colheita. Análise de variâncias de medidas repetidas ou teste de Friedman foram empregados para avaliação ao longo do tempo. A colheita foi produtiva em 67% dos bovinos e os locais de punção craniais esquerdo e direito foram os mais adequados. A concentração de proteína total variou de 1,4 a 3,6g/dL e o número de leucócitos de 54 a 1.322 células/µL, com predomínio de linfócitos (60 a 95% das células) no fluido peritoneal analisado logo após a colheita. A concentração de proteínas diminuiu, mas os valores absolutos de leucócitos, de linfócitos e de neutrófilos segmentados não se modificaram até oito horas após a colheita, independente do método de manutenção das amostras. A lise celular foi retardada pela refrigeração e a adição de formol não contribuiu para preservar a integridade da morfologia celular...(AU)

Peritoneal fluid collection in healthy cattle and evaluation of changes in cell morphology during storage in refrigerated and non-refrigerated samples / Colheita do fluido peritoneal de bovinos sadios e verificação das alterações na morfologia celular quando mantido refrigerado ou à temperatura ambiente

This study aimed to evaluate the appropriate sites of abdominocentesis for peritoneal fluid collection in cattle and to investigate the time of cell viability in vitro, comparing three methods of sample conservation. Twenty-one healthy cattle (19 females and 2 males) were subjected to a laparocentesis procedure to obtain peritoneal fluid, with punctures in three defined sites: left cranial, right cranial, and right caudal. The total peritoneal fluid collected was divided into three aliquots and maintained under three preservation conditions: room temperature (26°C), refrigeration (4°C), and room temperature (26°C) with the addition of 1µL of 10% formaldehyde per 1mL of peritoneal fluid. The peritoneal fluid analysis performed immediately after collection consisted of: physical examination (color, appearance, volume, and specific gravity), biochemical measures (pH, total protein, fibrinogen, creatinine, and glucose), and cellularity (total and differential counts). The determination of proteins and the examination of cells were repeated in each separate aliquot at two, four, six, and eight hours after harvest. Data were analyzed through repeated measures ANOVA or Friedman test. The harvest was productive in 67% of cattle. The left cranial and the right cranial puncture sites were the most appropriate. Peritoneal fluid analyzed after collection, the total protein concentration ranged from 1.4 to 3.6g/dL, and number of leukocytes ranged from 54 to 1,322 cells/µL; 60 to 95% of leukocytes were lymphocytes. The protein concentration decreased, but the absolute values of leukocytes, lymphocytes, and segmented neutrophils did not change up to eight hours after collection, independent of the maintenance method. Cell lysis was delayed by cooling, and the addition of formaldehyde did not help preserve the integrity of cellular morphology. Laparocentesis is a safe and secure procedure in cattle and maybe more productive when performed in specific sites on the left or right sides of the cranial abdominal wall. Peritoneal fluid samples may be analyzed with reliable results for up to eight hours after collection when kept refrigerated and for up to six hours when kept at room temperature.(AU)

O estudo teve como objetivo avaliar os locais adequados de laparocentese para a colheita de fluido peritoneal de bovinos e estabelecer o tempo de viabilidade celular in vitro, comparando três métodos de conservação. Vinte e um bovinos hígidos (19 fêmeas e 2 machos) foram submetidos ao procedimento de laparocentese para obtenção de fluido peritoneal, com punção em três pontos definidos: cranial esquerdo, cranial direito e caudal direito. O volume total do líquido peritoneal foi dividido em três alíquotas mantidas sob três métodos de conservação: temperatura ambiente (26°C); refrigeração (4°C); e temperatura ambiente (26°C) com adição de 1µL de formol 10% para cada 1mL de líquido peritonial. A análise do líquido peritoneal realizada imediatamente após sua obtenção consistiu em: exames físico (cor, aspecto, volume e densidade); bioquímicos (pH, proteína total, fibrinogênio, creatinina e glicose); e da celularidade (contagens total e diferencial). A determinação de proteínas e o exame da celularidade foram repetidos, em cada alíquota separada, as duas, quatro, seis e oito horas após a colheita. Análise de variâncias de medidas repetidas ou teste de Friedman foram empregados para avaliação ao longo do tempo. A colheita foi produtiva em 67% dos bovinos e os locais de punção craniais esquerdo e direito foram os mais adequados. A concentração de proteína total variou de 1,4 a 3,6g/dL e o número de leucócitos de 54 a 1.322 células/µL, com predomínio de linfócitos (60 a 95% das células) no fluido peritoneal analisado logo após a colheita. A concentração de proteínas diminuiu, mas os valores absolutos de leucócitos, de linfócitos e de neutrófilos segmentados não se modificaram até oito horas após a colheita, independente do método de manutenção das amostras. A lise celular foi retardada pela refrigeração e a adição de formol não contribuiu para preservar a integridade da morfologia celular. A laparocentese é um procedimento seguro e de execução fácil em bovinos sendo mais produtiva quando realizada em locais específicos à esquerda ou à direita craniais da parede abdominal. Amostras de fluido peritoneal podem ser analisadas com resultados confiáveis quando mantidas refrigeradas por até oito horas após a colheita e quando mantidas à temperatura ambiente por até seis horas.(AU)

Cytokines in peritoneal fluid of ovarian neoplasms.

The aim of this study was to evaluate cytokine levels (IL-2, IL-8, TNF-α, IL-5, IL-6 and IL-10) in the peritoneal fluid in non-neoplastic tumours, benign ovarian neoplasms and malignant ovarian neoplasms. Peritoneal fluid or ascites was collected from 117 patients with neoplastic and non-neoplastic ovarian tumours. Cytokine levels were assessed by ELISA. The unpaired groups were compared by the Kruskal-Wallis test with Dunn's post-test. Higher IL-6 levels were found in malignant neoplasms when compared to non-neoplastic tumours (p=.0241). There was no significant difference in the evaluation of other cytokines. Therefore, higher IL-6 levels in peritoneal fluid are related to the diagnosis of ovarian cancer. Further studies should be performed to evaluate the profile of cytokines in the peritoneal fluid of patients with ovarian tumours, and may be a new diagnostic strategy and a future target for treatment.Impact statementWhat is already known on this subject? Cytokines can be dosed in both the serum and peritoneal fluid, and in the ascitic fluid of women with ovarian neoplasia. Elevated levels of IL-6 were found in the ascitic fluid of patients with malignant ovarian tumours.What the results of this study add? To our knowledge, this is the first study in the literature that evaluates a large panel of cytokines in the peritoneal fluid (and not only in ascites), comparing non-neoplastic tumours, benign neoplasms and malignant ovarian neoplasms.What the implications are of these findings for clinical practice and/or further research? The cytokine dosage in the peritoneal fluid should be considered to map a profile of inflammatory cytokines that permeate the peritoneal cavity of patients with ovarian cancer. The dosage of these cytokines can be a potential pre-surgical tumour marker. In addition, a better understanding of the pattern of cytokines around ovarian neoplasia may be targeted for further studies in the development of new therapies for ovarian cancer.

Tuberculosis abdominal: patología infrecuente en un paciente joven: reporte de un caso / Abdominal tuberculosis: an unusual disease in a young patient: a case report

Resumen Introducción: La tuberculosis abdominal es un problema reemergente, y es una de las enfermedades transmisibles más importante en todo el mundo. A pesar de las expectativas acerca de su erradicación en países en desarrollo, ha sido recientemente declarada de nuevo como una patología de emergencia mundial. Con el aumento de su incidencia y prevalencia, su forma abdominal es una de las presentaciones de afectación extrapulmonar más comunes. Objetivo: Dado que la tuberculosis puede afectar diversos órganos, tiene una amplia gama y gran espectro de signos y síntomas que dificultan su diagnóstico y retrasan el tratamiento. Por esto, se realiza esta revisión de tema, concentrándonos en que el alto índice de sospecha debe ser un factor importante en el diagnóstico precoz, para que una vez establecido, se pueda iniciar el tratamiento ayudando a prevenir y disminuir las altas tasas de morbilidad y mortalidad evidenciadas en la actualidad. Caso Clínico: Paciente joven con presencia de ascitis secundaria a tuberculosis abdominal confirmada por una biopsia y el aumento de la adenosin deaminasa en el líquido peritoneal. Se describen los principales hallazgos clínicos, paraclínicos, estudios imagenológicos y tratamiento.

Introduction: Abdominal tuberculosis is a reemerging problem and is one of the most important communicable diseases in the world. Despite expectations about the eradication in developing countries, it has recently been re-declared as a global emergency pathology. The increased incidence and prevalence shows an abdominal shape as one of the most common extrapulmonary involvement presentations. Objective: Since tuberculosis can affect various organs, it has a wide range and spectrum of signs and symptoms that make diagnosis difficult and delay treatment. Therefore, this review of the topic is done, concentrating on the fact that the high suspicion index should be an important factor in the early diagnosis. Treatment can be initiated helping to prevent and reduce high morbidity and mortality rates. Case Report: We present a case of a young patient with ascites secondary to abdominal tuberculosis confirmed by biopsy and increased adenosine deaminase in the peritoneal fluid. The main clinical findings, paraclinic, imaging studies and treatment are described.

Plasma and peritoneal fluid concentrations of ceftriaxone after intravenous and intraperitoneal administration in horses.

Intraperitoneal (IP) use of antimicrobial agents may lead to therapeutic effects with better clinical results than intravenous (IV) administration. The aim of this study was to compare plasma and peritoneal fluid concentrations of ceftriaxone after IP and IV administration in horses, and to evaluate possible adverse effects. One group of five horses received 25mg/kg ceftriaxone diluted in 1L saline solution by IP catheter once daily for 5 days, while a second group of five horses received 25mg/kg ceftriaxone diluted in 250mL saline solution by IV injection once daily for 5days and 1L saline solution by IP catheter once daily for 5 days. Peritoneal fluid and plasma were collected to determine ceftriaxone concentrations after the first and fifth administration. IP administration of ceftriaxone resulted in concentrations above a minimum inhibitory concentration (MIC) of 1µg/mL for 24h in peritoneal fluid and for 12h in plasma, while IV administration of ceftriaxone resulted in lower peritoneal fluid concentrations, which remained above a MIC of 1µg/mL for 12h in peritoneal fluid and 10h in plasma. No adverse effects were observed. Comparisons of ceftriaxone concentrations, time of occurrence of the maximum (Tmax) and minimum (Tmin) concentrations, and the mean residence time (MRT), between the two groups showed that IP administration provided greater availability of cephalosporin in peritoneal fluid. The IP use of ceftriaxone (25mg/kg diluted in 1L saline solution once daily) may be useful for the prophylaxis and/or treatment of peritonitis in horses.

Ceftriaxone pharmacokinetics by new simple and sensitive ultra-high-performance liquid chromatography method.

Ceftriaxone is a cephalosporin antibiotic with a potent antimicrobial activity and excellent penetration in most body fluids such as pleural, peritoneal, spinal and brain. These facts contribute to the application of ceftriaxone in the treatment of bacterial peritonitis, an abdominal disorder in veterinary medicine, with potential risk of death. The determination of ceftriaxone levels in plasma and peritoneal fluid may be used to assess the pharmacokinetic profile at various instances of administration and allows observing if the concentrations needed are being achieved. Therefore a method was developed and validated for the determination of ceftriaxone in plasma and peritoneal fluid which after was applied in a pharmacokinetic profile study. The bioanalytical method validation was performed according to widely acceptable experiments. Two horses were used as a model of the method applicability; ceftriaxone was intraperitoneally administered to these animals as a single dose. The plasma and peritoneal fluid analysis were performed using an UHPLC system in reverse phase chromatography mode in fully validated conditions. The methods have shown linearity between 0.49 and 500µg/mL for plasma, and between 0.24 and 500µg/mL for peritoneal fluid. The quantitative analysis of ceftriaxone in these matrices allows monitoring of the therapy. This method showed improved sensitivity as well as the quantitation in peritoneal fluid.

Refractory ascites in systemic lupus erythematosus: further biological support of intraperitoneal steroid treatment as a suitable therapeutical option.

The objective of this report was to evaluate the ascitic fluid of a patient with refractory lupus ascites (proband) at different time points-pre- and post-intraperitoneal treatment with dexamethasone-using a multiparametric approach which included the presence of autoantibodies and pro- and anti-inflammatory cytokines and chemokines, and a proteomic analysis. As controls, we studied two additional patients also with lupus ascites (only at basal evaluation) and two patients with ascites due to alcoholic liver cirrhosis. High levels of anti-dsDNA and anti-nucleosomes autoantibodies were detected in the ascitic fluid of all lupus patients and remained elevated in the proband throughout the follow-up. All lupus patients have detectable ascitic high levels of IL-6, IL-8, IL-10, TNF-α, MCP-1, and IGF-1 which diminished gradually in the proband after intraperitoneal dexamethasone. In the proteomic analysis of the ascitic fluid, a marked increment of apolipoprotein A1 was observed and again, it diminished gradually after intraperitoneal treatment. Our findings further support the use of intraperitoneal steroids as an effective therapeutic option for refractory ascites in systemic lupus erythematosus.

Evaluation of data from 35 dogs pertaining to dehiscence following intestinal resection and anastomosis.

The objectives of this study were to evaluate blood and abdominal fluid lactate and glucose, fluid cytology, culture, and volume 24 and 48 hr following intestinal resection and anastomosis in dogs with and without closed-suction drains and to correlate findings with survival. Thirty-five client-owned dogs that underwent intestinal resection and anastomosis were prospectively enrolled in the study. Abdominal fluid was submitted for culture at surgery and again 24 hr postoperatively. Twenty-four and 48 hr postoperatively, blood and abdominal fluid glucose and lactate were measured and fluid was submitted for cytology. Abdominal fluid was collected either from a closed-suction drain or by abdominocentesis. Patients were followed either for 14 days or until death. Comparisons were made based on development of dehiscence and presence or absence of a drain. Patients with dehiscence were more likely to have positive cultures at 24 hr and to have had more bowel resected. Surviving patients without drains had significantly smaller differences in blood and fluid glucose and lactate both 24 and 48 hr postoperatively than surviving patients with drains. The significant differences identified between patients with and without drains suggests a need for further research into the effect of drains on abdominal fluid values.

[Diagnostic of ascites due to portal hypertension: accuracy of the serum-ascites albumin gradient and protein analises in ascitic fluid]. / Eficacia de la gradiente de albúmina sangre-ascític y los análisis de proteínas en líquido ascítico en el diagnóstico de ascitis hipertensiva portal.

OBJECTIVE: To evaluate the diagnostic accuracy of the Serum-Ascites Albumin Gradient (GASA), Protein Concentration in the Ascitic Fluid (PTLA), Albumin Concentration in the ascitic fluid (CAA) and the Protein Ascites/Serum Ratio (IPAS) for the diagnosis of ascites due to portal hypertension. MATERIALS AND METHODS: it was an observational and retrospective study of validation of diagnostic tests. The study population was patients from a National Public Health Hospital Daniel Alcides Carrion of Callao, Peru, during the period January to December of 2012, patients over 15 years old with a diagnosis of ascites which samples were taken for study by paracentesis with an standard technique, it was analyzed total protein and albumin, as well as study of total protein and albumin in blood. We obtained the diagnostic accuracy, sensitivity, specificity, PPV and NPV of the Serum-Ascites Albumin Gradient (GASA), Protein Concentration in the Ascitic Fluid (PTLA), Albumin Concentration in the ascitic fluid (CAA) and the Protein Ascites/Serum Ratio (IPAS) for the diagnosis of ascites due to portal hypertension. To determine ascites by HTP as diagnostic tests we took into account: GASA >= 1.1, PTLA <2.5, CAA <1.1 or IPAS< 0.5. RESULTS: There were 126 patients diagnosed with ascites, 10 patients was excluded for having incomplete data. Of the 116 patients, the average age was 53.03 +/- 15.73 years old, male 65 (56%) and female 51 (44%). 61 (52%) had ascites due to portal hypertension from liver cirrhosis, and 55 (48%) of ascites due to NO HTP. The sensitivity and specificity for GASA was 93% and 47% respectively, for PTLA was 80% and 89% respectively, for CAA was 85% and 87% respectively and for the IPAS was 83% and 80% respectively. The área under the ROC curve for GASA was 0.70, ATPL was 0.84, IPAS was 0.81 and CAA was 0.86, we found statistically significant differences between GASA compared to the other three parameters (p<0.01 ). CONCLUSION: The diagnostic accuracy of CAA, ATPL and IPAS is higher than the GASA to discriminate between ascites due to HTP or NO HTP, so that they could be used in clinical practice alone or together to achieve a diagnostic approach more successful.

Evaluation of tumor markers for the differential diagnosis of benign and malignant ascites.

INTRODUCTION: The diagnosis of malignant ascites is a challenging problem in clinical practice, non-invasive techniques should be developed to improve diagnostic accuracy. The diagnostic performances of tumor markers in malignant ascites remained unsettled. Our aim was to evaluate diagnostic performance of tumor markers in differential diagnosis of benign and malignant ascites. MATERIAL AND METHODS: A total of 437 patients were enrolled, and the relevant parameters of the patients were analyzed for the differentiation of benign ascites from malignant ascites. RESULTS: At the predetermined cutoff values of tumor makers, tumor markers in ascitic fluid showed better diagnostic performance than those in serum. Combined use of tumor markers and the cytology increased the diagnostic yield of the latter by 37%. In cytologically negative malignant ascites, tumor markers provided assistance in differentiating malignant ascites from benign ascites, and the combination of ascitic tumor markers yielded 86% sensitivity, 97% specificity. CONCLUSION: Use of a panel of tumor markers exhibited excellent diagnostic performance in diagnosing malignant ascites, which indicated the detection of tumor markers may represent a beneficial adjunct to cytology, thus guiding the selection of patients who might benefit from further invasive procedures.

Proteomic identification of fucosylated haptoglobin alpha isoforms in ascitic fluids and its localization in ovarian carcinoma tissues from Mexican patients.

BACKGROUND: Ovarian cancer is the most lethal gynecologic disease due to delayed diagnosis, and ascites production is a characteristic of patients in advanced stages. The aim of this study was to perform the proteomic analysis of ascitic fluids of Mexican patients with ovarian carcinoma, in order to detect proteins with a differential expression pattern in the continuing search to identify biomarkers for this disease. METHODS: Samples were collected from 50 patients from the Instituto Nacional de Cancerología of México under informed consent and with approval of the bioethics and scientific committees. After elimination of abundant proteins (Albumin/IgGs) samples were processed for 2D electrophoresis and further protein identification by Mass Spectrometry (MALDI-TOF). Molecules of interest were followed by western blot and lectin binding assays, and their tissue location by histo-immunofluorescence and confocal analysis. RESULTS AND DISCUSSION: An area with a differential expression pattern among samples was located in the 2D gels. Identified proteins were 6 alpha 1 isoforms and 1 alpha 2 isoform of Haptoglobin, and 2 isoforms of Transthyretin. While Transthyretin isoforms were constitutively expressed in all samples, clear differences in the expression pattern of Haptoglobin alpha isoforms were found. Moreover, increased levels of fucosylation of Haptoglobin alpha isoforms analyzed in 40 samples by Aleuria aurantia lectin binding by 1D overlay assay showed a positive correlation with advanced stages of the disease. Tissue detection of Haptoglobin and its fucosylated form, by histo-immunofluorescence in biopsies of ovarian cancer, also showed a correlation with ovarian cancer progression. Moreover, results show that fucosylated Haptoglobin is produced by tumor cells. CONCLUSIONS: Increased numbers of highly fucosylated Haptoglobin alpha isoforms in ascitic fluids and the presence of fucosylated Haptoglobin in tumor tissues of ovarian cancer Mexican patients associated with advanced stages of the disease, reinforce the potential of fucosylated Haptoglobin alpha isoforms to be characterized as biomarkers for disease progression.

Eficacia de la gradiente de albúmina sangre-ascitis y los análisis de proteínas en líquido ascítico en el diagnóstico de ascitis hipertensiva portal: accuracy of the serum-ascites albumin gradient and protein analises in ascitic fluid / Diagnostic of ascites due to portal hypertension

Objetivo: Evaluar la exactitud diagnóstica de la gradiente albúmina sangre/ascitis (GASA), proteínas totales en líquido ascítico (PTLA), albúmina en líquido ascítico (CAA) e índice de proteínas ascitis/suero (IPAS) para el diagnóstico de ascitis por hipertensión portal. Materiales y métodos: Se realizó un estudio, observacional, retrospectivo, de validez de pruebas diagnósticas. La población estudiada fueron pacientes mayores de 15 años con diagnóstico de ascitis a los cuales se les tomó una muestra para estudio del líquido ascítico mediante la técnica estándar de paracentesis, analizando proteínas totales y albúmina, además de estudio de proteínas totales y albúmina en sangre en el Hospital de Salud Pública Nacional Daniel Alcides Carrión del Callao, Perú (HNDCA), durante el periodo de enero a diciembre del 2012. Se obtuvo la exactitud diagnóstica, sensibilidad, especificidad, VPP y VPN de la gradiente albumina sangre/ascitis (GASA), proteínas totales en líquido ascítico (PTLA), albúmina en líquido ascítico (CAA) e índice de proteínas ascitis/suero (IPAS) para el diagnóstico de ascitis por hipertensión portal o no HTP. Para determinar ascitis por HTP según las pruebas diagnósticas se tomo en cuentas: GASA≥1,1, PTLA<2,5, CAA<1,1 o IPAS<0,5. Resultados: se obtuvieron 126 pacientes con diagnóstico de ascitis a los cuales se excluyó 10 pacientes por tener datos incompletos. De los 116 pacientes finales la edad promedio fue de 53,03 ± 15,73 años, pacientes de sexo masculino fueron 65 (56%) y femenino 51 (44%). Se encontró 61 (52%) líquidos ascíticos debido a HTP por cirrosis hepática, y 55 (48%) de ascitis por NO HTP. La sensibilidad y especificidad para el GASA fue de 93% y 47% respectivamente, para PTLA fue de 80% y 89% respectivamente, para CAA fue de 85% y 87% respectivamente y para el IPAS fue de 83% y 80% respectivamente. El área bajo la curva ROC para el GASA fue de 0,70, de las PTLA fue de 0,84, del IPAS fue de 0,81 y de la CAA fue de 0,86; encontrándose diferencia estadísticamente significativa entre el GASA comparado con los otros tres parámetros (p<0,01). Conclusión: La exactitud diagnóstica de la CAA, PTLA y IPAS es superior a la del GASA para discriminar entre ascitis por HTP o NO HTP, por lo que podrían ser usados en la práctica clínica de forma aislada, o en conjunto para lograr una aproximación diagnóstica más acertada.

Objective: To evaluate the diagnostic accuracy of the Serum-Ascites Albumin Gradient (GASA), Protein Concentration in the Ascitic Fluid (PTLA), Albumin Concentration in the ascitic fluid (CAA) and the Protein Ascites/Serum Ratio (IPAS) for the diagnosis of ascites due to portal hypertension. Materials and methods: it was an observational and retrospective study of validation of diagnostic tests. The study population was patients from a National Public Health Hospital Daniel Alcides Carrion of Callao, Peru, during the period January to December of 2012, patients over 15 years old with a diagnosis of ascites which samples were taken for study by paracentesis with an standard technique, it was analyzed total protein and albumin, as well as study of total protein and albumin in blood. We obtained the diagnostic accuracy, sensitivity, specificity, PPV and NPV of the Serum-Ascites Albumin Gradient (GASA), Protein Concentration in the Ascitic Fluid (PTLA), Albumin Concentration in the ascitic fluid (CAA) and the Protein Ascites/Serum Ratio (IPAS) for the diagnosis of ascites due to portal hypertension. To determine ascites by HTP as diagnostic tests we took into account: GASA ≥ 1.1, PTLA <2.5, CAA <1.1 or IPAS< 0.5. Results: There were 126 patients diagnosed with ascites, 10 patients was excluded for having incomplete data. Of the 116 patients, the average age was 53.03 +/- 15.73 years old, male 65 (56%) and female 51 (44%). 61 (52%) had ascites due to portal hypertension from liver cirrhosis, and 55 (48%) of ascites due to NO HTP. The sensitivity and specificity for GASA was 93% and 47% respectively, for PTLA was 80% and 89% respectively, for CAA was 85% and 87% respectively and for the IPAS was 83% and 80% respectively. The area under the ROC curve for GASA was 0.70, ATPL was 0.84, IPAS was 0.81 and CAA was 0.86, we found statistically significant differences between GASA compared to the other three parameters (p<0.01 ). Conclusion: The diagnostic accuracy of CAA, ATPL and IPAS is higher than the GASA to discriminate between ascites due to HTP or NO HTP, so that they could be used in clinical practice alone or together to achieve a diagnostic approach more successful.

Neospora caninum and Toxoplasma gondii: relationship between hepatic lesions, cytological and biochemical analysis of the cavitary liquid during the acute phase of the diseases in experimental models.

The objective of this study was to evaluate the pathogenesis of ascites in mice infected with Toxoplasma gondii and gerbils infected with Neospora caninum during the acute phase disease. For that, 12 gerbils [Experiment I: not infected/control (n=6) and infected (n=6)] and 12 mice [Experiment II: control (n=6) and infected (n=6)] were used. Infected gerbils and mice showed marked ascites on days 5-7 post-infection (PI), while the not-infected animals had not ascites. Peritoneal liquid was collected from the all mice with uninfected animals receiving 1.5mL of saline solution into their abdominal cavity, allowing the recovery of cavity liquid. As a result, it was possible to observe differences in physics, chemistry and cytological analysis of the fluid cavity of animals infected with N. caninum and T. gondii, when they were compared with uninfected animals, as well as between animals experimentally infected. Additionally both, N. caninum and T gondii, caused an increase in the levels of nitric oxide (NOx-nitrate/nitrite), protein oxidation (AOPP) and lipid peroxidation (TBARS), while serum total protein and albumin were reduced in infected gerbils and mice. Gerbils infected with N. caninum showed multiple large cells with multilobulated nucleus, lytic necrosis and abundant amount of eosinophilic cytoplasm into the hepatic parenchyma. By the other hand, mice infected with T. gondii developed myriad foci of lytic necrosis combined with tachyzoites and cysts containing bradyzoites in liver. Both experimental models for N. caninum and T. gondii showed inflammatory foci and tachyzoites the peritoneum, which could be a major cause of ascites. Toxoplasmosis and neosporosis were able to cause clinical signs in experimental models with similar alterations in peritoneal fluid; however the toxoplasmosis histological changes were much more evident. Therefore, the pathogenesis of ascites appears to be directly related to liver damage, which strongly suggests alteration in the normal production of proteins as observed in this study, along with peritonitis.

Measurement of serum and peritoneal levels of amyloid protein A and their importance in the diagnosis of pelvic endometriosis.

OBJECTIVE: To evaluate serum and peritoneal concentrations of amyloid protein A in women with endometriosis and to compare them with those of women without endometriosis. STUDY DESIGN: A prospective study evaluated 76 women suspected of having pelvic endometriosis. Fifty-seven women (group A) were confirmed by videolaparoscopy and had their serum and peritoneal amyloid A concentrations measured by ELISA. The average levels from group A were compared to those obtained in group B. Group B was composed of 13 women without endometriosis, submitted to elective laparoscopy for tubal ligation. RESULTS: Peritoneal amyloid A concentrations in group A (310.3 +/- 97.8 ng/mL) were higher than those of group B (53.4 +/- 58.2 ng/mL); p = 0.0. However, serum concentrations in groups A (14.01 +/- 32.3 ng/mL) and B (9.5 +/- 15.9 ng/mL) did not differ significantly; p = 0.35. CONCLUSION: The peritoneal amyloid A protein concentration in pelvic endometriosis was higher when compared to normal controls, corroborating the inflammatory nature of the disease. This finding suggests that the procedure of evaluating the peritoneal amyloid A concentration in endometriosis merits further investigation.

Characteristics of ascitic fluid from patients with suspected spontaneous bacterial peritonitis in emergency units at a tertiary hospital.

CONTEXT AND OBJECTIVE: Spontaneous bacterial peritonitis (SBP) is a complication of ascites, especially in cirrhosis. Ascitic fluid with 250 or more neutrophils/mm³ is an acceptable criterion for diagnosis, even when bacterial fluid cultures are negative. The aims here were to estimate SBP frequency among emergency room patients based on cellular criteria and evaluate the biochemical profile of these fluids. DESIGN AND SETTING: Retrospective study at a public tertiary hospital. METHODS: Laboratory records of patients with ascites attended in emergency rooms between November 2001 and November 2006, from whom ascitic fluid samples were sent to the laboratory due to suspected SBP, were evaluated. The 691 samples included were divided into group A (presumed SBP: ≥ 250 neutrophils/mm³; n = 219; 31.7%) and group B (no presumed SBP: < 250 neutrophils/mm³; n = 472; 68.3%). Patients' sex and age; ascitic fluid characteristics (numbers of neutrophils, leukocytes and nucleated cells); bacteriological characteristics; and protein, lactate dehydrogenase, adenosine deaminase and glucose concentrations were evaluated. RESULTS: Among group A cultured samples, 63 (33.8%) had positive bacterial cultures with growth of pathogens commonly associated with SBP. In total, the group A samples showed higher lactate dehydrogenase levels than seen in the group B samples. The latter presented predominance of lymphocytes and macrophages. CONCLUSION: Among the ascitic fluid samples with clinically suspected SBP, 31.7% fulfilled the cellular diagnostic criteria. Positive bacterial isolation was found in 33.8% of the cultured samples from the presumed SBP group.

Taenia crassiceps: A secretion-substance of low molecular weight leads to disruption and apoptosis of seminiferous epithelium cells in male mice.

The present research was performed to isolate and study the effects of a low molecular weight (<1300Da) parasite-associated substance, obtained from peritoneal fluids of female mice infected with Taenia crassiceps cysticerci, on seminiferous epithelium cells of male mice testis. The results showed an intense disruption of Sertoli cells and germ cells within the seminiferous tubules of experimental mice, along with the destruction of their gap junction (GJ). Significant generalized apoptosis of germ cells within seminiferous tubules was determined by TUNEL staining (P=0.0159). In addition, a significant number of infiltrating macrophages were found in the luminal space of these seminiferous tubules (P<0.0001). Finally, electron microscopy studies revealed structural and morphological abnormalities in the somatic cells (Sertoli and Leydig cells) and in the germ cells, primarily in the round and elongate spermatids.

Characteristics of ascitic fluid from patients with suspected spontaneous bacterial peritonitis in emergency units at a tertiary hospital / Características do líquido ascítico de pacientes com suspeita de peritonite bacteriana espontânea nas unidades de emergência de um hospital terciário

CONTEXT AND OBJECTIVE: Spontaneous bacterial peritonitis (SBP) is a complication of ascites, especially in cirrhosis. Ascitic fluid with 250 or more neutrophils/mm³ is an acceptable criterion for diagnosis, even when bacterial fluid cultures are negative. The aims here were to estimate SBP frequency among emergency room patients based on cellular criteria and evaluate the biochemical profile of these fluids. DESIGN AND SETTING: Retrospective study at a public tertiary hospital. METHODS: Laboratory records of patients with ascites attended in emergency rooms between November 2001 and November 2006, from whom ascitic fluid samples were sent to the laboratory due to suspected SBP, were evaluated. The 691 samples included were divided into group A (presumed SBP: > 250 neutrophils/mm³; n = 219; 31.7 percent) and group B (no presumed SBP: < 250 neutrophils/mm3; n = 472; 68.3 percent). Patients' sex and age; ascitic fluid characteristics (numbers of neutrophils, leukocytes and nucleated cells); bacteriological characteristics; and protein, lactate dehydrogenase, adenosine deaminase and glucose concentrations were evaluated. RESULTS: Among group A cultured samples, 63 (33.8 percent) had positive bacterial cultures with growth of pathogens commonly associated with SBP. In total, the group A samples showed higher lactate dehydrogenase levels than seen in the group B samples. The latter presented predominance of lymphocytes and macrophages. CONCLUSION: Among the ascitic fluid samples with clinically suspected SBP, 31.7 percent fulfilled the cellular diagnostic criteria. Positive bacterial isolation was found in 33.8 percent of the cultured samples from the presumed SBP group.

CONTEXTO E OBJETIVO: Peritonite bacteriana espontânea (PBE) é uma complicação da ascite, especialmente na cirrose. Líquido ascítico com 250 ou mais neutrófilos/mm³ é um critério aceitável para o diagnóstico, mesmo com cultura bacteriana negativa. Os objetivos foram estimar a frequência de PBE em pacientes atendidos na sala de emergência, baseando-se no critério celular e avaliar o perfil bioquímico desses líquidos peritoneais. TIPO DE ESTUDO E LOCAL: Estudo retrospectivo em hospital público terciário. MÉTODOS: Foram avaliados registros laboratoriais de pacientes com ascite atendidos no setor de emergência entre novembro de 2001 e novembro de 2006, cujas amostras de líquido ascítico foram encaminhadas ao laboratório por suspeita de PBE. As 691 amostras incluídas foram divididas em grupo A (PBE presumida: > 250 neutrófilos/mm³; n = 219; 31.7 por cento) e grupo B (Ausência de PBE presumida: < 250 neutrófilos/mm3; n = 472; 68.3 por cento). Também foram avaliados sexo e idade dos pacientes além de características dos líquidos ascíticos: número de neutrófilos, leucócitos e células nucleadas; bacteriologia; e concentrações de proteínas, desidrogenase láctica, adenosina deaminase e glicose. RESULTADOS: Das amostras cultivadas do grupo A, 63 (33,8 por cento) tiveram cultura bacteriana positiva com crescimento de patógenos comumente associados à PBE. O total de amostras do grupo A exibiu maiores níveis de desidrogenase lática que as do grupo B. Este último demonstrou predomínio de linfócitos e macrófagos. CONCLUSÃO: Dos líquidos ascíticos com suspeita clínica de PBE, 31.7 por cento preencheram o critério diagnóstico celular. O isolamento bacteriano foi positivo em 33.8 por cento das amostras cultivadas no grupo PBE presumida.