Myelodysplastic syndrome progression to acute myeloid leukemia in a cat FeLV seroreactive

Background: Acute myeloid leukemia (AML) is a group of non-lymphoid hematological tumors characterized by aberrant proliferation and/or decreased apoptosis of a clone of non-mature cells, resulting in the accumulation of immature blast cells in the bone marrow and peripheral blood. It is considered rare, as it represents 10% of neoplasms of hematopoietic origin. However, it is known that felines seroreactive for FIV and FeLV are more predisposed and reports of this type of leukemia in cats in the literature are scarce. Thus, the objective of this study was to evaluate the blood and bone marrow of a cat seroreactive for FeLV that presented with myelodysplastic syndrome that progressed to acute myeloid leukemia. Case: A 6-year-old male mixed-breed cat, neutered, seroreactive for FeLV, showed apathy, weight loss, and pale mucous membranes. Initial peripheral blood smear evaluation revealed hypochromic normocytic anemia, leukopenia, neutropenia, lymphopenia, and thrombocytosis with many macropackets and giant platelets. Based on this blood picture, a long-spectrum antimicrobial therapy with amoxicillin and clavulanate [Clavulin® BD - 25 mg/kg, every 12 h] was started. Granulocyte colony stimulating factor used filgrastim (rHu G-CSF) [Fiprina® - 5 µg/kg, SC, every 48 h] and appetite stimulant mirtazapine [Mirtz® - 2 mg/cat, orally, every 48 h] were used to correct leukopenia and nutritional status, respectively. Follow-up blood smear evaluation on the 30th day showed persistence of the hematological changes noticed earlier. A bone marrow puncture was performed, and immunosuppressive therapy with prednisolone [Predsim® - 4 mg/kg, orally, every 24 h] was initiated. The aspirated material showed increased cellularity for age, decreased myeloid:erythroid ratio, and 39.8% of blasts of myeloid origin. An average of 17.7 megakaryocytes were observed per field (10x magnification). Bone marrow cytological evaluation suggested acute myeloid leukemia with dysmegakaryocytopoiesis. After the diagnosis, the examinations were repeated monthly, and there was still intense leukopenia. However, in view of the stable clinical status and leukopenia with neutropenia, treatment for leukemia was not instituted and only supportive treatment was administered when necessary. Eight months after the diagnosis, clinical status had worsened, and unlike the earlier hemograms, global leukocyte count had increased with predominant lymphocytosis (95% of the total leukocytes) with atypical lymphocytes. The cat died a few days later. Discussion: Bone marrow evaluation is indicated when peripheral blood cell abnormalities are present and cannot be explained in the context of the clinical history. In the present report, the bone marrow aspirate was hypercellular (cellularity above 75%); however, intense leukopenia was observed in the peripheral blood. In myelodysplastic syndromes, it is common for the bone marrow to be normal to hypercellular, which occurs when there is a greater production of myeloid or erythroid cell lines in response to the loss, destruction, or consumption of cells. Despite this, cytopenias may be present in the peripheral blood, since the defective cells undergo apoptosis and die before being released into the circulation, characterizing inefficient hematopoiesis. The diagnosis of acute leukemia comprises a variety of hematopoietic neoplasms that are complex and unique. Each acute leukemia subtype has defining characteristics that affect the prognosis and treatment of each animal.

The porcine biomodel in translational medical research: From biomodel to human lung transplantation / El biomodelo porcino en la investigación médica traslacional: del biomodelo al humano en trasplante pulmonar.

Introduction: Human and porcine anatomy are comparable. In consequence, the porcine biomodel has the potential to be implemented in the training of surgical professionals in areas such as solid organ transplantation. Objectives: We described the procedures and findings obtained in the experiments of translational respiratory medicine with the porcine biomodel, within an experimentation animal laboratory, and we present a comparative review between human and porcine lung. Materials and methods: The experiment was done in nine pigs of hybrid race within a laboratory of experimental surgery. The anatomy and histology of the respiratory tract were studied with fibrobronchoscopy, bronchial biopsy and bronchoalveolar lavage. The bronchoalveolar lavage was studied with liquid-based cytology and assessed with Papanicolau and hematoxylin-eosin staining. Molecular pathology techniques such as immunohistochemistry, flow cytometry, and electronic microscopy were implemented. The pigs were subjected to left pneumonectomy with posterior implantation of the graft into another experimental pig. Results: Histopathologic and molecular studies evidenced predominance of alveolar macrophages (98%) and T-lymphocytes (2%) in the porcine bronchoalveolar lavage. Studies on the porcine lung parenchyma revealed hyperplasic lymphoid tissue associated with the bronchial walls. Electronic microscopy evidenced the presence of T-lymphocytes within the epithelium and the cilia diameter was similar to the human. Conclusions: The porcine biomodel is a viable tool in translational research applied to the understanding of the respiratory system anatomy and the training in lung transplantation. The implementation of this experimental model has the potential to strength the groups who plan to implement an institutional program of lung transplantation in humans.

Introducción. La anatomía humana y porcina son comparables. En consecuencia, el biomodelo porcino tiene el potencial de ser implementado para entrenar al profesional quirúrgico en áreas como el trasplante de órganos sólidos. Objetivo. Describir los procedimientos y hallazgos obtenidos mediante experimentos de medicina respiratoria traslacional con biomodelos porcinos realizados en un laboratorio de experimentación animal, y hacer una revisión comparativa entre el pulmón humano y el porcino. Materiales y métodos. El experimento se llevó a cabo en nueve cerdos de raza híbrida en un laboratorio de cirugía experimental. Se estudiaron la anatomía y la histología de las vías respiratorias mediante fibrobroncoscopia, biopsia bronquial y lavado broncoalveolar. El lavado broncoalveolar se estudió con citología en base líquida y se evaluó con las coloraciones de Papanicolau y hematoxilina y eosina. Se utilizaron técnicas de patología molecular, como inmunohistoquímica, citometría de flujo y microscopía electrónica. Los cerdos se sometieron a neumonectomía izquierda con posterior implante del injerto en otro cerdo experimental. Resultados. Los estudios histopatológicos y moleculares evidenciaron un predominio de macrófagos alveolares (98 %) y linfocitos T (2 %) en el lavado broncoalveolar porcino. En los estudios del parénquima pulmonar porcino se encontró tejido linfoide hiperplásico asociado a las paredes bronquiales. La microscopía electrónica evidenció linfocitos T dentro del epitelio y el diámetro de las cilias porcinas fue similar al de las humanas. Conclusiones. El biomodelo porcino es viable en la investigación traslacional para el entendimiento de la anatomía del sistema respiratorio y el entrenamiento en trasplante pulmonar. La implementación de este modelo experimental podría fortalecer los grupos que planean implementar un programa institucional de trasplante pulmonar en humanos.

El biomodelo porcino en la investigación médica traslacional: del biomodelo al humano en trasplante pulmonar / The porcine biomodel in translational medical research: From biomodel to human lung transplantation

Resumen Introducción. La anatomía humana y porcina son comparables. En consecuencia, el biomodelo porcino tiene el potencial de ser implementado para entrenar al profesional quirúrgico en áreas como el trasplante de órganos sólidos. Objetivo. Describir los procedimientos y hallazgos obtenidos mediante experimentos de medicina respiratoria traslacional con biomodelos porcinos realizados en un laboratorio de experimentación animal, y hacer una revisión comparativa entre el pulmón humano y el porcino. Materiales y métodos. El experimento se llevó a cabo en nueve cerdos de raza híbrida en un laboratorio de cirugía experimental. Se estudiaron la anatomía y la histología de las vías respiratorias mediante fibrobroncoscopia, biopsia bronquial y lavado broncoalveolar. El lavado broncoalveolar se estudió con citología en base líquida y se evaluó con las coloraciones de Papanicolau y hematoxilina y eosina. Se utilizaron técnicas de patología molecular, como inmunohistoquímica, citometría de flujo y microscopía electrónica. Los cerdos se sometieron a neumonectomía izquierda con posterior implante del injerto en otro cerdo experimental. Resultados. Los estudios histopatológicos y moleculares evidenciaron un predominio de macrófagos alveolares (98 %) y linfocitos T (2 %) en el lavado broncoalveolar porcino. En los estudios del parénquima pulmonar porcino se encontró tejido linfoide hiperplásico asociado a las paredes bronquiales. La microscopía electrónica evidenció linfocitos T dentro del epitelio y el diámetro de las cilias porcinas fue similar al de las humanas. Conclusiones. El biomodelo porcino es viable en la investigación traslacional para el entendimiento de la anatomía del sistema respiratorio y el entrenamiento en trasplante pulmonar. La implementación de este modelo experimental podría fortalecer los grupos que planean implementar un programa institucional de trasplante pulmonar en humanos.

Abstract Introduction: Human and porcine anatomy are comparable. In consequence, the porcine biomodel has the potential to be implemented in the training of surgical professionals in areas such as solid organ transplantation. Objectives: We described the procedures and findings obtained in the experiments of translational respiratory medicine with the porcine biomodel, within an experimentation animal laboratory, and we present a comparative review between human and porcine lung. Materials and methods: The experiment was done in nine pigs of hybrid race within a laboratory of experimental surgery. The anatomy and histology of the respiratory tract were studied with fibrobronchoscopy, bronchial biopsy and bronchoalveolar lavage. The bronchoalveolar lavage was studied with liquid-based cytology and assessed with Papanicolau and hematoxylin-eosin staining. Molecular pathology techniques such as immunohistochemistry, flow cytometry, and electronic microscopy were implemented. The pigs were subjected to left pneumonectomy with posterior implantation of the graft into another experimental pig. Results: Histopathologic and molecular studies evidenced predominance of alveolar macrophages (98%) and T-lymphocytes (2%) in the porcine bronchoalveolar lavage. Studies on the porcine lung parenchyma revealed hyperplasic lymphoid tissue associated with the bronchial walls. Electronic microscopy evidenced the presence of T-lymphocytes within the epithelium and the cilia diameter was similar to the human. Conclusions: The porcine biomodel is a viable tool in translational research applied to the understanding of the respiratory system anatomy and the training in lung transplantation. The implementation of this experimental model has the potential to strength the groups who plan to implement an institutional program of lung transplantation in humans.

Estudio del polimorfismo genético de las células de la médula ósea y del sistema nervioso central de ratas mediante la técnica de RAPD / Study of genetic polymorphism of the bone marrow and the central nervous system of rats cells by RAPD technique

Los modelos experimentales en rata han sido de gran utilidad en las evaluaciones terapéuticas o de reemplazo de células en enfermedades neurodegenerativas. Se ha comprobado que las células de la médula ósea (CMO) de ratas pueden diferenciarse en células que no forman parte de sus linajes normales. Hay evidencias de estos procesos de trans-diferenciación, pero aún no se conocen los mecanismos moleculares que activan estos procesos. El propósito de nuestro trabajo fue estudiar el polimorfismo genético del ADN de los tipos celulares que conforman las CMO y las células del sistema nervioso central (SNC), estríatales y de la corteza de ratas mediante la técnica de RAPD. Las CMO, las células mononucleares (CMMO), las células estromales (CEMO) y las del SNC fueron obtenidas de ratas, y su ADN genómico fue purificado y amplificado mediante la técnica de RAPD, utilizando 15 cebadores al azar. Se construyó un dendograma de las bandas de amplificación generadas utilizando el método de UPGMA. Las células estudiadas según el análisis del RAPD quedaron en 2 grupos bien definidos, pudiéndose diferenciar las CEMO del resto de las células estudiadas. Los cebadores OPA-6, 7 y 12, mostraron el polimorfismo genético de los linajes de células estudiadas. Mediante la técnica de RAPD se demostró la variabilidad genética entre las CEMO y las CMMO, células estriadas y de corteza que mostraron una homogeneidad genética, proponiéndose marcadores específicos de RAPD para cada grupo de células. Este es el primer estudio del polimorfismo genético de las CMO y del SNC de ratas.

Experimental models have been of grate usefulness in the therapeutic or replacement cells in neurodegenerative diseases. It has been demonstrated that bone marrow cells (BMC), can be difefferentiated in cells that do not form part of their normal lineage. There is evidence of these trans-differentiation processes in these cells, but nevertheless, molecular mechanisms that activate these differentiation process still not known. The purpose of our work was to study the genetic polymorphism of those cellular types; that conform the rat bone marrow cells (BMC) as well as those of the central nervous system (CNS), striatum cells and cortex ones, trough RAPD technique. BM, mononuclear cells (BMMC), estromal cells (BMSC) and the CNS cells were obtained from rats and genomic ADN was purified and amplified through RAPD technique, using 15 random primers. A dendogram was constructed according to UPGMA method of the amplifying RAPD bands. Studied cells as- according to the RAPD analysis- were grouped into 2 well- defined groups, as CEMO coud be differentiated from the rest of studied cells. OPA-6, 7 and 12 primers showed the genetic polymorphism of the studied lineages cells. Also will be proposed specific RAPD genetic markers. Through RAPD technique permitted the genetic variability was demonstrated betwen BMEC and BMMC of striated cells and of cortex, which demonstratd a genetic homogeneity through RAPD technique so specific genetic markers of RAPD were thus propose for each group of cells. These constitute the first study on genetic polymorphism of BMC and CNS.

Early alveolar bone regeneration in rats after topical administration of simvastatin.

OBJECTIVES: The aim of this study was to ultrastructurally examine the influence of simvastatin on bone healing in surgically created defects in rat mandibles. STUDY DESIGN: Bone defects 0.8 mm in diameter were created in the buccal aspect of first mandibular molar roots and filled with 2.5% simvastatin gel, while the controls were allowed to heal spontaneously. The rats were humanely killed 7, 9, 11, or 14 days postoperatively, and the specimens were processed for scanning and transmission electron microscopy, as well as for colloidal gold immunolabeling of osteopontin. RESULTS: The regenerated alveolar bone in the simvastatin-treated defects presented smaller marrow spaces, and the collagen fibrils were regularly packed exhibiting a lamellar bone aspect. Osteopontin was present through the bone matrix during the wound healing and alveolar bone regeneration. CONCLUSION: The present study provides evidence that a single topical application of 2.5% simvastatin gel improves the quality of the new bone and decreases bone resorption.

Imaging resin-cast osteocyte lacuno-canalicular system at bone-bioactive glass interface by scanning electron microscopy.

The morphology of the osteocyte lacuno-canalicular system at the bone-biomaterial implant-interface has not been fully investigated. In this study, the resin-cast scanning electron microscopy technique was used, for the first time, to image the lacuno-canalicular network within neoformed bone around bioactive glass (BG) particles implanted in rat tibia bone marrow. The most salient finding was that the osteocyte canaliculi pass through the calcium-phosphorus layer formed at the bone-BG interface and reach the silica-rich layer of the reacted BG.

Protein-energy malnutrition alters histological and ultrastructural characteristics of the bone marrow and decreases haematopoiesis in adult mice.

Protein-energy malnutrition (PEM) decreases resistance to infection by impairing a number of physiological processes, including haematopoiesis. The aim of this study was to evaluate the microanatomical aspects of bone marrow (BM) in mice that were subjected to PEM, in particular, with respect to the components of the local extracellular matrix and the proliferative activity of haematopoietic cells. For this, histological, histochemical, immunohistochemical and ultrastructural techniques were used. Two-month old male Swiss mice were fed with a low-protein diet containing 4% protein and control mice fed a 20% protein diet. When the experimental group had attained a 25% loss of their original body weight, we collected the different biological samples. Malnourished mice had presented severe BM atrophy as well as a reduction in proliferating cell nuclear antigen and gelatinous degeneration. The malnourished mice had more fibronectin accretion in paratrabecular and endosteal regions and more laminin deposition in perisinusal sites than controls. Endosteal cell activation and hyperplasia were found, suggesting their participation in the process. Additionally, we have observed a decrease in the capacity of malnourished haematopoietic stroma to support the growth of haematopoietic stem cells (CD34+) in vitro. These findings point to a structural impairment of the haematopoietic microenvironments in mice with PEM, possibly hampering the interactions between cells and cellular signalling.

Structural aberrations of chromosomes 17 and 12 in chronic B-cell disorders.

OBJECTIVES: Genomic aberrations can now be identified in approximately 80% of chronic lymphocytic leukemia, small lymphocytic lymphoma (CLL/SLL) patients. In the present study, four new structural changes involving chromosomes 17 and 12 in CLL/SLL patients are described. METHODS: Five patients were selected for inclusion in the present report among a total of 92 cases with diagnosis of CLL/SLL. Cytogenetic studies and fluorescence in situ hybridization (FISH) analysis to detect some of the most frequent cryptic aberrations occurring in CLL/SLL patients were performed. Clinical studies are also described. RESULTS: Four cases showed structural rearrangements of chromosome 17. A psu dic(17;2)(p11.2;p21), leading to p53 deletion, was observed in a patient who developed a mixed cellularity Hodgkin's disease coexisting with the CLL/SLL in the same lymph node. Epstein Barr virus was detected in the Reed-Sternberg cells. Two cases had a balanced translocation t(2;17)(p21;q23). Both patients showed trisomy 12 and clonal evolution and one of them also had 11q deletion. In addition, a der(17)t(12;17)(q13;q25) as a part of a complex karyotype, and a complex translocation t(5;12;19) (q15;p11;q13) were also found. Four patients had an adverse clinical outcome and died because of disease progression. CONCLUSIONS: Four unreported nonrandom chromosome aberrations in CLL/SLL patients, one of them who might represent a new recurrent abnormality, are described. These uncommon abnormalities, mostly associated with evolving disease, may have implications for the understanding of genetic events associated with disease progression in this pathology.

Effects of chlorophyllin on acetaldehyde: lack of modulation of the rate of sister-chromatid exchanges in mouse bone marrow, and of complex formation in aqueous solution.

Acetaldehyde (Ace) is a reactive compound widely found in natural and industrialized products. On the other hand, chlorophyllin (Chl) is a chloropyll derivative which has shown DNA modulatory effects in several models. The first aim of the present study was to determine the capacity of Ace to increase the rate of sister-chromatid exchanges (SCEs) in mouse bone marrow cells in vivo, as well as to determine its capacity to modify the mitotic index (MI) and the average generation time (AGT). For this experiment we tested four dosages of Ace by the i.p. route (0.4, 4.0, 40.0 and 400 mg/kg), and found a genotoxic effect with the two highest dosages (more than double the basal level was observed with 400 mg/kg). We also found that none of the doses tested modified the MI or the AGT. A second objective was to explore the potential of Chl to modulate the genotoxicity of Ace in the same model. We evaluated whether an oral administration of Chl (2.0, 6.0 and 10.0 mg/kg), given 1 h before an i.p. administration of Ace (100 mg/kg), could modulate the SCEs produced by the mutagen. The result showed a similar SCE rate in both, the Ace-treated mice and those administered with the two chemicals, indicating that Chl was not a modulatory chemical on the genotoxicity of Ace. No modifications were observed concerning the MI or the AGT either. A third objective was to determine whether the two compounds (Ace and Chl) may form a molecular complex in aqueous solution. In agreement with the lack of modulatory effect by Chl, a reversed HPLC and a spectrophotometric analysis showed that the two compounds were unable to form a complex. This report confirms the importance of the specificity concerning the interaction mutagen/antimutagen.

Characterization of bone around titanium implants and bioactive glass particles: an experimental study in rats.

PURPOSE: Many situations in clinical practice require metallic implants to be combined with bone grafts and/or bone substitutes such as bioactive glass (BG). Upon implantation, silica-based BG particles are transformed into a shell containing calcium and phosphate that loses its inner silicon-rich core. The release of silicon by BG particles and its incorporation by newly formed bone tissue in the peri-implant area had not been studied to date. MATERIALS AND METHODS: Thirty Wistar rats were used throughout. Under anesthesia, a commercially pure titanium (Ti) laminar implant was placed inside the medullary compartment of the tibia (Ti group), while in the contralateral tibia (Ti/BG group) a titanium laminar implant and melt-derived BG 45S5 particles were implanted. The animals were sacrificed 14, 30, and 60 days postimplantation. The tibiae were resected, radiographed, and embedded in methyl methacrylate resin. Sections were stained with toluidine blue and analyzed by light microscopy and energy-dispersive x-ray analysis (EDX). The presence of silicon, calcium, and phosphorus was evaluated in the BG particles and in the peri-implant bone tissue for each of the experimental times. RESULTS: The histomorphometric study revealed an increase in peri-implant bone thickness in the Ti/BG group as compared to the Ti group. EDX of newly formed bone tissue showed a transient appearance of silicon at 14 and 30 days postimplantation and a rise in the calcium:phosphorus ratio in peri-implant bone tissue in the Ti/BG group. DISCUSSION: The present study shows an increase in reactive medullary bone formation when BG particles are implanted around a Ti implant. CONCLUSION: The results described in the present study reveal that the release of Si by BG particles is an important issue that warrants further study.

Haematopoiesis and a new mechanism for the release of mature blood cells from the bone marrow into the circulation in snakes (Ophidia).

This is the first description of haematopoiesis in snakes. Studies were carried out on the following species belonging to Ophidia: Bothrops jararaca, Bothrops jararacusu, Waglerophis merremii, Elaphe taeniura taeniura, Boa constrictor,and Python reticulatus. Smears of the peripheral blood and histological preparations from the vertebrae, ribs, liver, and spleen were studied under a light and electron microscope. Myeloid cells were present in the following locations in the vertebrae: the neural spine, zygoapophysial processes, floor of the neural canal, lacunae in the bodies of vertebrae and also inside the ribs. Although the vascular system was well developed, especially around the ribs, vessels inside the marrow cavities were scarce, both in the ribs and elsewhere where haematopoiesis was found. Venous sinuses were well developed in the vertebrae and in the rib regions from their costal head towards the middle area. They consisted of one layer of fine endothelial cells. Mature cells in the process of migration into the general circulation were only sporadically encountered when venous sinuses were studied on perfusion-fixed specimens. In contrast, almost every sinus venosus contained protrusions directed towards the lumen, filled mostly with mature and immature blood cells. Various stages of their formation were seen in the cross sections of venous sinuses ranging from small, newly formed to large, elongated ones, filled with many fully developed and some maturing blood cells. In many cases the apices of the protrusions were ruptured, and mature blood cells, as well as a few immature ones, were seen in their vicinity. This observation led us to a new hypothesis that blood cells are released from the extravascular space into the lumen of venous sinuses. In snakes, these cells are released into the systemic circulation mainly via the rupture of protrusions filled with mature blood cells and, to a lesser degree, by transcytosis as known in mammals. In the spleens from young specimens, 1-2 foci of haematopoiesis were encountered where lymphopoiesis predominated. Haematopoiesis was not detected in the liver.

Evaluation of the genotoxic potential of the isocoumarin paepalantine in in vivo and in vitro mammalian systems.

Paepalantine is an isocoumarin isolated from Paepalanthus vellozioides which showed antimicrobial activity in in vitro experiments. In the present study, paepalantine was tested for possible clastogenic and cytotoxic action. Cultures from different individuals were treated with paepalantine at concentrations of 20, 40 and 80 microg/ml. The effect of isocoumarin was also tested in an in vivo assay using Wistar rat bone marrow cells. Paepalantine was administered intraperitoneally at concentrations of 6.25, 12.5 and 25 mg/kg body weight. Under these conditions paepalantine did not have a clastogenic effect, but was significantly cytotoxic in the in vitro and in vivo mammalian cell systems tested in the present work.

Cocaine mutagenicity and hepatocarcinogenicity evaluations in rodents.

The mutagenicity (clastogenicity) and the carcinogenicity (promoting potential) of cocaine were evaluated, respectively, by the mouse bone marrow micronucleus test (study I) and by the initiated rat liver bioassay (study II). In study I, two administration routes (i.p. and i.v.) and two sampling times (24 and 48 hours) after cocaine treatment were studied. Swiss male mice were treated with cocaine at doses of 0, 18, 37, and 75 mg/kg and 0, 2, 4, and 8 mg/kg by i.p. and i.v. routes, respectively. No significant differences were observed between treated and negative control groups regarding the frequencies of micronuclei and the polichromatic/normochromatic erythrocyte (PCE/NCE) ratios. In study II, the development of putative preneoplastic foci of hepatocytes expressing the enzyme glutathione S-transferase placental form (GST-P+) was utilized as the end-point marker in a 8-week rat liver bioassay. The animals were initiated for carcinogenesis by a single i.p. sub-carcinogenic dose of diethylnitrosamine (DEN). After a 6-week exposure to 5 or 10 mg/kg of cocaine i.v. twice a week there was no enhancement of GST-P+ foci development above the values of the control DEN-only treated animals. Also, cocaine did not induce any toxicity as evidenced by the absence of alterations of rat body and liver weights and of liver biochemical function and morphology. The results suggest that cocaine does not have a mutagenic effect on the mouse bone marrow cells or promoting activity on the rat hepatocarcinogenesis process.

Genotoxicity of an extract of Calendula officinalis L.

A fluid extract of Calendula officinalis L. displayed genotoxic properties when assayed for mitotic segregation in the heterozygous diploid D-30 of Aspergillus nidulans. The extract of Calendula exhibited dose-dependent toxicity and genotoxicity (both mitotic crossing-over and chromosome malsegregation being observed) to Aspergillus in the range of five plate concentrations from 0.1 to 1.0 mg of solids/ml assayed. Mutagenicity testing with the Salmonella/microsome assay in strains TA 1535, TA 1537, TA 98 and TA 100 was negative in a plate incorporation protocol, with concentrations ranging from 50 to 5000 microg/plate (+/- S9). The mouse bone marrow micronucleus test, where the extract was dosed orally up to 1 g/kg for 2 days, was also negative.

Ultrastructural changes of stromal cells of bone marrow and liver after cyclophosphamide treatment in mice.

The direct effect of cyclophosphamide (CY) on bone marrow and liver stromal cells of DBA/2 and C57BL/6 mice after a single intraperitoneal dose of 300 mg/kg, was assessed. Ultrastructural observations were carried out 2,4,6,12,18, and 24 h and 2,3,5,7,10,15,22 and 30 days after CY in femoral bone marrow and liver fixed by immersion or vascular perfusion. A massive depletion of hemopoietic cells was observed in bone marrow as early as 12 h after CY treatment, and normality was recovered only after 10 days. Among stromal cells, sinus endothelial cells, reticular cells, and macrophages were particularly sensitive to CY and showed severe damage as soon as 2 h after treatment. There was also an important dilatation of sinusoids, and the presence of mature red cells in the hemopoietic parenchyma, and macrophages and immature cells in the lumen and in circulating blood demonstrated the loss of integrity of the endothelium. In the liver, injured cells showing vesiculation and disruption of endothelial and Kupffer cells of sinusoids were evident 6 h after CY. The alterations caused by CY were transient. Although recovery of the hemopoietic cells in bone marrow and liver was achieved by day 10, the stromal cells showed damage even 15 days after CY, and a return to normality was only reached on day 30. Thus, the effect of CY on stromal cells, that was longer lasting than the effect on the hemopoietic compartment, demonstrated a higher recovery capacity of hemopoiesis with respect to stromal cells. These results demonstrate that recovery of hemopoiesis occurred even while the severe damage inflicted by CY to the stromal cells remained unrepaired.

Macrophage activation syndrome in systemic juvenile rheumatoid arthritis successfully treated with cyclosporine.

A macrophage activation syndrome, possibly related to methotrexate toxicity, developed in a boy with systemic juvenile rheumatoid arthritis. Corticosteroid administration was ineffective, whereas a prompt response to cyclosporine was observed. Two months later, Pneumocystis carinii pneumonia developed.

Myelodysplasia and deficiency of uridine diphosphate-galactose 4-epimerase.

A 4-year-old girl known to have peripheral uridine diphosphate-galactose 4-epimerase deficiency was examined for bruising and thrombocytopenia. She had dysplastic peripheral blood and bone marrow changes, with a global platelet function defect. Uridine diphosphate-galactose-4-epimerase participates in a metabolic pathway that provides substrates for posttranslational glycosylation of secreted and membrane glycoproteins, including hematopoietic growth factors and their receptors; there may be a causal relationship between the two disorders.

Estudio citogenético en 22 adultos y tres niños con leucemia linfoblástica aguda / Cytogenetic study in 22 adults and therr children with acute lymphoblastic leukemia

Se presentan los resultados del estudio citogenético realizado en sangre periférica y/o médula ósea de 25 pacientes (22 adultos y 3 niños) con LLA estudiados entre 1987 a 1990. Todos los casos presentaron alteraciones cromosómicas (25/25), aunque en 23 pacientes (23/25) estaba presente una línea celular normal. Entre las aberriguaciones estructurales encontradas están t(17;19) (q11;p13), t(2;9;22)(q34;q34;q11), t(1;7)(p13;q33), t(6;11)(q26;p16), t(3;4)(q24-25;q26), t(1;12)(q23;q34), t(2;18)(q15;p12), t(2;4;)(q23;35) y t(4;11)(q21ñq23). Estos rearreglos fueron indicadores de riesgo y se correlacionaron con la respuesta al tratamiento y la supervivencia de los pacientes. Algunas de estas anomalías no habían sido previamente descritas en LLA; sin embargo, los puntos de ruptura coinciden con los observados en otras neoplasias hematológicas, lo cual suguiere que esas regiones son críticas en la patogénesis de estos desórdenes.

Simplified method for the analysis of cellular karyotype and phenotype in leukemias

The objective of this study was to develop a simplified method for the simultaneous analysis of cellular karyotype and phenotype which would permit the identification of cell origin. We studied 6 patients with AML, 3 with CML (one of which was in blastic transformation) and one ALL. We used a method in which the suspension of bone marrow cells was incubated in TC 199 medium with colchicine and with hypotonic solution formed from glycerol, NaCl, KCl, CaCl2, MgCl2 and sucrose. The slides were prepared from this cell suspension by cytospin and stained for peroxidase, PAS, esterases and iron. The karyotype was studied by direct method and culture. It was possible to relate the cytogenetic marker with cytochemistry characteristics in the same cell in 3 cases, showing the feasibility of cytochemistry techniques in cytogenetical preparations. The best preparations were found through peroxidase. The presence of iron granules allowed identification of erythroblastic lineage in the combined staining. Mitosis with a marker chromosome of leukemic clone in an AML cell with negative peroxidase probably showed a proliferation of more primitive precursor not sufficiently differentiated to show markers