Localization of Entamoeba histolytica amebopore in amebic liver abscesses in hamsters.

Amebopore was purified from axenically grown trophozoites of the Entamoeba histolytica strain HM1:IMSS. The purification procedure involved Mono Q anion-exchange chromatography and electroelution. Sequence analysis of the final product revealed that amebopore A was completely pure. Polyclonal antibodies against the purified amebopore were obtained from rabbits, and Western blot studies demonstrated their specificity. Sections of experimental, acute (1, 2, 3, and 4 days), amebic liver abscesses produced in hamsters were stained with the anti-amebopore antibody; in all the analyzed stages, amebopore appeared as a constitutively expressed cytoplasmic molecule in trophozoites. No extracellular or hepatocyte-membrane amebopore was found. This study is the first to trace amebopore in an in vivo model of amebic liver abscesses.

Ex vivo evaluation of PBMNCs collected with a new cell separator.

BACKGROUND: This study reports on an evaluation of the ability of a cell separator (Amicus, Baxter Healthcare) and the integral MNC computer software program to collect a variety of MNC subsets. The collection efficiency (CE) of the Amicus for these MNC subsets was compared to that of another cell separator (CS-3000 Plus, Baxter). The collected MNCs were also assayed ex vivo to determine if these cells remained functional. STUDY DESIGN AND METHODS: Healthy volunteer blood donors were recruited to provide PBMNCs for the isolation of CD3+, CD4+, CD8+, CD19+, NK, and gammadelta+ cells and monocytes. Cells were collected with an Amicus (test arm; n = 16) or a CS-3000 Plus (control arm; n = 11) cell separator. Cells were counted on a flow cytometer and CEs were calculated. For functional studies, the Amicus-collected MNC data were compared to CS-3000 Plus historical data. Functional studies performed included surface antigen expression assays (CD8+), proliferation assays (CD4+ and CD8+ cells), NK cytotoxicity assays for K562 and HUVE cells, and E-selectin induction on endothelial cells through NK+ contact dependency. Dendritic cells (DCs) were generated from CD34+ cells collected on the Amicus, positively selected by the use of antibody-bound, magnetic bead technology, and then cultured ex vivo with a combination of growth factors to generate the DCs. RESULTS: CEs were higher on the Amicus than on the CS-3000 Plus for CD3+ (68 vs. 54%), CD4+ (70 vs. 56%), CD8+ (68 vs. 52%), and CD19+ (60 vs. 48%) cells (p<0.05). For the two separators, CEs were equivalent for monocytes, NK+, and gammadelta+ cells. The Amicus separator collected significantly fewer platelets than did the CS-3000 Plus (p<0.00001). CD4+, CD8+, and NK cells proliferated normally. NK cells appropriately stimulated E-selectin expression on endothelial cells. Culture-generated DCs obtained by using Amicus-collected CD34+ cells expressed appropriate cell surface markers. CONCLUSION: The Amicus separator is acceptable for the collection of PBMNC subsets. The device collects CD3+, CD4+, CD8+, and CD19+ T- and B-cell subsets with greater efficiency and collects MNCs with significantly fewer contaminating platelets than does the CS-3000 Plus. Cells collected on the Amicus are suitable for use in a variety of research and clinical immunobiologic studies.

[Cerebellar hemorrhage in full-term newborn babies]. / Hemorragia cerebelosa en recién nacidos a término.

INTRODUCTION: Cerebellar hemorrhage (CH) has been observed in 5 to 10% of the autopsies done on newborn babies. Since neuroimaging techniques have become available it is easier to diagnose the condition. In this paper we report on a series of cases of CH in full-term newborn babies. OBJECTIVES: To determine the number of patients with CH diagnosed by neuroimaging, make a descriptive study and analyze their progress. RESULTS: Between 1984 and 1999 six patients had CH, three boys and three girls, five born after their mother's first pregnancy. Four were vaginal births; in two forceps were used and in one a vacuum extractor; two were born by cesarean section. Four showed symptoms within the first 48 hours of life, one on the fourth day and one on the twenty fifth day. The latter had hemorrhagic disease of the newborn. In five patients transfontanellar ultrasound was useful in diagnosis. In all six cases computerized axial tomography scan confirmed the diagnosis. Cerebral magnetic resonance (MR) was done in three cases. No arteriovenous malformations were shown on angio-MR. Two patients had hydrocephalus and both were treated by ventriculo-peritoneal shunts. Three cases had transient ventricular dilatation which improved with medical treatment. The patient with hemorrhagic disease of the newborn had alterations in blood clotting. In three patients metabolic studies were normal. Five patients were treated conservatively and only one neurosurgically. Subsequent evolution was characterized by the presence of psychomotor retardation with mild cerebellar signs. At school age, only observed in two cases, there were learning difficulties with a low intellectual coefficient and problems with reading and writing. In one case there was epilepsy, controlled by use of two antiepileptic drugs. CONCLUSIONS: Half the cases of CH transfontanellar ultrasound is useful in diagnosis. Most patients were managed conservatively. During the clinical course there were psychomotor retardation, cerebellar signs, cognitive deficits with learning problems and epilepsy.

Hemorragia cerebelosa en recién nacidos a término / Cerebellar hemorrhage in full-term newborn babies

Introducción. Las hemorragias cerebelosas (HC) se han observado en el 5-10 por ciento de las autopsias neonatales. Con el advenimiento de las técnicas de neuroimagen pueden diagnosticarse con mayor facilidad. En el presente trabajo presentamos nuestra casuística de HC en recién nacidos a término (RNAT). Objetivos. Determinar el número de pacientes con HC diagnosticados por neuroimagen, realizar un estudio descriptivo y analizar su evolución. Resultados. Seis pacientes entre 1984 y 1999 padecieron una HC en el período neonatal, tres niños y tres niñas, cinco producto del primer embarazo; cuatro nacieron vía vaginal -en dos se utilizaron fórceps y en uno vacuum extractor-, y dos fueron cesáreas. Cuatro presentaron sintomatología durante las primeras 48 horas de vida, uno al cuarto día y uno a los 25 días; éste presentó enfermedad hemorrágica del recién nacido (EHRN).En cinco la ecografía tranfontanelar (ECOTF) fue útil en el diagnóstico. En seis la TAC confirmó el diagnóstico y en tres se realizó resonancia magnética (RM) cerebral, la angio-RM no detectó malformaciones arteriovenosas. Dos presentaron hidrocefalia, en ambos se colocó derivación ventriculoperitoneal, y tres presentaron dilatación ventricular transitoria que mejoró con el tratamiento médico. El paciente con EHRN mostró alteraciones de coagulación, en tres los estudios metabólicos fueron negativos. Cinco recibieron tratamiento conservador y solamente uno neuroquirúrgico. La evolución se caracteriza por la presencia de retraso psicomotor con signos cerebelosos leves. En la edad escolar, evaluable sólo en dos casos, se constata la presencia de trastornos de aprendizaje con cociente intelectual bajo y problemas de lectoescritura, un caso manifiesta epilepsia controlada con dos fármacos. Conclusiones. El 50 por ciento de las HC que se presentaron en neonatos a término fueron producto de partos distócicos. Un caso fue secuendaria a EHRN. La ECOTF es útil en el diagnóstico. La mayoría fueron tratados con medidas conservadoras. La evolución posterior viene marcada por retraso psicomotor, signos cerebelosos, déficit cognitivos con problemas escolares y epilepsia (AU)

In vitro collection and posttransfusion engraftment characteristics of MNCs obtained by using a new separator for autologous PBPC transplantation.

BACKGROUND: A clinical study was performed to evaluate the peripheral blood progenitor cell (PBPC) collection, transfusion, and engraftment characteristics associated with use of a blood cell separator (Amicus, Baxter Healthcare). STUDY DESIGN AND METHODS: Oncology patients (n = 31) scheduled for an autologous PBPC transplant following myeloablative therapy were studied. PBPCs were mobilized by a variety of chemotherapeutic regimens and the use of G-CSF. As no prior studies evaluated whether PBPCs collected on the Amicus separator would be viable after transfusion, to ensure patient safety, PBPCs were first collected on another cell separator (CS-3000 Plus, Baxter) and stored as backup. The day after the CS-3000 Plus collections were completed, PBPC collections intended for transfusion were performed using the Amicus instrument. For each transplant, >2.5 x 10(6) CD34+ PBPCs per kg of body weight were transfused. RESULTS: Clinical data collected on the donors immediately before and after PBPC collection with the Amicus device were comparable to donor data similarly obtained for the CS-3000 Plus collections. While the number of CD34+ cells and the RBC volume in the collected products were equivalent for the two devices, the platelet content of the Amicus collections was significantly lower than that of the CS-3000 Plus collections (4.35 x 10(10) platelets/bag vs. 6.61 x 10(10) platelets/bag, p<0.05). Collection efficiencies for CD34+ cells were 64 +/- 23 percent for the Amicus device and 43 +/- 14 percent for the CS-3000 Plus device (p<0.05). The mean time to engraftment for cells collected via the Amicus device was 8.7 +/- 0.7 days for >500 PMNs per microL and 9.7 +/- 1.5 days to attain a platelet count of >20,000 per microL-equivalent to data in the literature. No CS-3000 Plus backup cells were transfused and no serious adverse events attributable to the Amicus device were encountered. CONCLUSIONS: The mean Amicus CD34+ cell collection efficiency was better (p<0.05) than that of the CS-3000 Plus collection. Short-term engraftment was durable. The PBPCs collected with the Amicus separator are safe and effective for use for autologous transplant patients requiring PBPC rescue from high-dose myeloablative chemotherapy.

Tissue transplant-transmitted infections.

Tissue bankers, as well as those transplanting tissues, have been sensitized to the possibility of transmission of fatal infection via tissue transplants, particularly following recent reports of a few cases of AIDS or HIV infection from bone, semen, and skin grafts. It is beyond the scope of this review to describe the steps taken by tissue banks to enhance the safety of tissue transplants. Of note is the fact that a number of new donor screening tests, such as those for antibody to HIV, HBcAg, hepatitis C virus, and human T cell lymphotropic virus type I, have recently been implemented. In addition, rapid advances in the medical history screening of tissue donors and tissue procurement, processing, and preservation continue. Viral inactivation studies are also being undertaken. All these measures are being introduced to increase the safety of tissue transplants.

Four crossmatch methods to select platelet donors.

We employed four crossmatch techniques to select platelet donors for refractory patients. Forty-four donor-recipient pairs were studied in 32 patients. Analysis of effectiveness of platelet transfusions revealed that only 18 percent of transfusions gave a borderline response; the remainder were either effective or not effective at all. The corrected predictive values of three crossmatch tests were as follows: enzyme-linked immuno-specific assay, 81 percent; platelet immunofluorescence test, 73 percent; and lymphocytotoxicity, 70 percent (p greater than 0.05). The predictive value of these tests did not differ in HLA-matched versus unmatched platelet transfusions. Donor selection by lymphocytotoxicity compatibility did not appear to be useful if donors were selected by either of the other two methods. The fourth test, antiglobulin-modified lymphocytotoxicity, offered no advantage over lymphocytotoxicity. Our data suggest that platelet crossmatching assays are a useful adjunct to the selection process for the platelet donor in addition to ABO, Rh, and HLA matching.

Application of the enzyme linked immunospecific assay (ELISA) for the detection of platelet antibodies.

Many methods have been described to identify platelet antibody, but they are either not very sensitive or too complex for general use. Therefore, we have developed an enzyme immunoassay for the detection of platelet antibodies in serum. The method involves incubating platelets with serum antibody; any attached antibody is shown by the addition of an enzyme (alkaline phosphatase) labeled anti-human IgG, followed by assay of the enzyme reaction with its substrate. The reaction product is indicated by a color change, which is proportional to the antibody concentration. Assay conditions such as the use of paraformaldehyde fixed versus unfixed platelets, conjugate dilutions, and substrate concentration and incubation time were investigated. Positive results were obtained in 16 of 19 sera of patients with various diseases including 2 of 4 patients with idiopathic thrombocytopenic purpura, 2 of 2 with post-transfusion purpura, 2 of 3 with neonatal purpura, and all 9 polytransfused patients. Sensitivity and specificity were 84% and 98%, respectively. Also, enzyme linked immunospecific assay (ELISA) was found to be superior to the lymphocytotoxicity (LCT) and platelet immunofluorescence test (PIIFT) for platelet antibody identification.