Simple template-based optimization for pediatric total lymphoid irradiation (TLI) radiotherapy treatments.

Total lymphoid irradiation (TLI) is used in the management of pediatric allogeneic hematopoietic stem cell transplantation (HSCT. This work aims to simplify the treatment planning process for TLI via a proposed template using the volumetric modulated arc therapy (VMAT) technique. Fifteen pediatric patients were planned, prescribed to 8 Gy in 4 fractions. Cost functions included in the template were the ones for the planning target volume (PTV), and conformality cost function (CCF) for the rest of the patient's volume. Conformity index (CI), homogeneity index (HI), conformation number (CN), gradient index (GI), integral dose, and doses to the organs at risk achieved with the template were reported. Cost function influence over various indexes was studied by Wilcoxon signed ranks test. Same 15 patients were planned with 3-dimensional conventional radiotherapy (3D-CRT) technique for comparison. Mean CI and HI were 1.33 and 0.13, respectively, which indicates good dose conformation and homogeneity. Mean CN and GI values were 0.69 and 4.51, respectively. Mean PTV coverage was reached (V100% > 95%). No correlation between the CCF and indexes values was found (p > 0.05). Doses to organs at risk (OARs) were as low as possible without losing PTV coverage. VMAT plan showed higher levels of conformation and similar homogeneity as 3D-CRT plans. Doses to OARs were inferior with VMAT except for the right kidney. The proposed template simplifies the planning of TLI treatments, and it is able to create acceptable plans with little modification in order to reduce doses to certain organs like the kidneys or the heart. VMAT technique showed higher conformation and lower doses to OAR compared to 3D-CRT.

TLI in pediatric patients.

PURPOSE: Hematopoietic progenitor cell transplantation (HSCT) is a procedure used in different hematological diseases as part of the curative treatment, so the investigators propose a system of conditioning of reduced intensity based on total lymphoid irradiation (TLI) as an alternative to the classic total body irradiation (TBI) followed by haploidentical transplantation in patients compatible with a single HLA haplotype, as an alternative to patients who do not have an HLA compatible donor. MATERIALS AND METHODS: A cohort of 25 patients with hematological disease underwent haploidentical HSCT from February 2015 to May 2018, conditioned with TLI from day - 10 (2-4 days of treatment) followed by thiotepa (5 mg/kg/12 h) and melphalan (70 mg/m2/day) prior to HSCT and prophylaxis with ciclosporin (1.5 mg/kg/12 h). 2 Gy/fraction was administered to complete 8 Gy with IMRT and VMAT technique. RESULTS: 12% rejection of the transplant was obtained with acute GVHD < II (48%) and chronic GVHD 12%. No acute toxicity was recorded in irradiated patients and 56% survival of patients at the end of follow-up. CONCLUSION: Conditioning the haploidentical transplant with TLI, IMRT, and VMAT techniques compared with TBI and RT3D-C techniques is a feasible technique that helps inducing the necessary immunosuppression in patients with a high risk of graft rejection, minimal adverse effects, low incidence of GVHD, and high survival rate.

[Phagocytic function of polymorphonuclear leukocytes and lymphocyte subpopulations in patients with amebiasis]. / Función fagocitaria de leucocitos polimorfonucleares y subpoblaciones linfocitarias en pacientes con amibiasis.

We initiated a pilot study to evaluate the functions of polymorphonuclear leukocytes and also to analyze the behavior of lymphocyte subpopulations in patients with different clinical forms of amoebiasis. We found significant differences in many aspects as NBT reduction, Candida digestion, CD4/CD8 relation, Ia expression. We suggest a wider investigation of the mechanism of cell mediated immunity in patients with amoebiasis.

A novel method for estimating killing ability and digestion of Paracoccidioides brasiliensis by phagocytic cells in vitro.

We describe a novel method by which phagocytosis, digestion and killing of Paracoccidioides brasiliensis yeast cells by polymorphonuclear leukocytes or other phagocytic cells may be estimated simultaneously. Suspensions of P. brasiliensis (yeast-like phase) were sonicated, counted and incubated at 37 degrees C with known numbers of phagocytes. Control preparations contained no phagocytic cells. At given intervals samples were incorporated into gelatin nutrient medium and droplets of the mixtures were incubated at room temperature. Live yeast-like P. brasiliensis germinate in vitro and produce filaments. After incubation, droplets may be melted and examined under phase contrast optics, or the cells may be washed and stained by a variation of Papanicolaou's method. Digested P. brasiliensis, intact but non-germinating yeasts and filamented (viable) yeasts may be identified and counted. Killing and digestive abilities of phagocytes may be estimated by the difference between values obtained from phagocyte-containing and control preparations.

Studies on plating efficiency and estimation of viability of suspensions of Paracoccidioides brasiliensis yeast cells.

Mild sonication was used to obtain single cell suspensions of Paracoccidioides brasiliensis. These cells were intact by microscopic criteria. Direct cell counts in a given inoculum and colony formation on various media were used to determine plating efficiency. Sonicated and nonsonicated cell suspensions were used to study plating efficiency and to estimated viability by means of vital dyes. Methylene blue Erythrosin B, and Janus green were unreliable when used with P. brasiliensis, but vital dyes were accurate when tested with Candida albicans. Acridine orange gave more meaningful results of viability. Estimates of viability, however, changed significantly as a result of relatively minor alterations in the composition of the suspending medium. In initial experiments, the plating efficiency of P. brasiliensis was dismally low. It descended abruptly with increasing dilution of inoculum. Efficiency was much improved if horse serum was added to brain heart infusion plates or if glucose glycine yeast extract (GGY) plates were incubated at room temperature and mycelial colonies were counted. With the technique we report, current plating efficiency of sonicated suspensions is of the order of 25%. Our results and procedures have an important bearing upon those studies concerned with in vitro killing of P. brasiliensis in suspensions or with isolating this fungus from clinical or environmental specimens.

Defect of in vitro digestive ability of polymorphonuclear leukocytes in paracoccidioidomycosis.

Selected functions of polymorphonuclear leukocytes were studied in patients with paracoccidioidomycosis (South American blastomycosis), in healthy control individuals, and in patients with diseases unrelated to paracoccidioidomycosis. Patients with paracoccidioidomycosis were also evaluated by standard immunological techniques. Phagocytosis and digestion of Paracoccidioides brasiliensis yeastlike cells in vitro was estimated by an original method. It was based on the appearance of phagocytosed P. brasiliensis in preparations stained by a modification of the Papanicolaou method and examined with phase-contrast optics. Interpretation of such findings was confirmed by electron microscopy. Two strains of P. brasiliensis were used. Strain 8506 was freshly isolated from a patient. Strain Pb9 was known to be nonpathogenic and to have a peculiar cell wall composition. Yeastlike cells of the Pb9 strain were digested significantly better than those of strain 8506. A higher number of leukocytes per fungus cells led to a higher proportion of digested P. brasiliensis. Leukocytes from patients with paracoccidioidomycosis phagocytosed the fungus in a normal way, but had a significant lower ability to digest it in vitro. When individual cases were analyzed, there was an excellent correlation between clinical evolution and digestive ability of polymorphonuclear leukocytes. There was good correlation between both of these and immunological parameters. Leukocytes from all groups behaved comparably in tests of general leukocyte function and in their abilities to kill and digest Candida albicans. Our results indicate that, as a group, polymorphonuclear leukocytes from patients with paracoccidioidomycosis had a significant, rather specific, defect in their in vitro digestive capacity against phagocytosed P. brasiliensis. There was also an inverse correlation between strain pathogenicity and its susceptibility to in vitro digestion by polymorphonuclear leukocytes. Our findings are concordant with and relevant to clinical reality.

In vitro activation of neutrophils by suspensions of Mycobacterium leprae.

Activation, defined as an increase in the proportion of cells that reduce nitroblue-tetrazolium in vitro, is present in neutrophils from patients with reactional lepromatous leprosy but not in neutrophils from patients with non-reactional lepromatous leprosy. Neutrophils from patients with all forms of leprosy are equally well activated by endotoxin in vitro. We have now shown that in vitro activation induced by Mycobacterium leprae suspensions is of comparable magnitude in neutrophils from patients with all forms of leprosy (including lepromatous and reactional lepromatous leprosy). There is no intrinsic neutrophil anergy in patients with lepromatous leprosy vis-à-vis M. leprae as pertains to activation. Spontaneous activation in reactional lepromatous leprosy is likely due to an indirect mechanism, probably of immunologic nature, and not simply to the presence of circulating Mycobacterium leprae in the blood.

New method for estimating digestion of Paracoccidioides brasiliensis by phagocytic cells in vitro.

We describe a method by which phagocytosis and digestion of Paracoccidioides brasiliensis yeast cells by polymorphonuclear leukocytes or other phagocytic cells may be estimated. Suspensions of P. brasiliensis in its yeastlike phase were sonicated, counted, and incubated with known numbers of peripheral blood polymorphonuclear leukocytes. At given intervals, cytocentrifuge droplets were stained by a variation of Papanicolaou's method. Stained preparations were examined with phase-contrast optics. Digested organisms showed total or partial disappearance of protoplasm. Green-stained cell walls resisted digestion. The proportion of digested cells as a function of time was estimated.