Fine needle aspiration cytology of localized Leishmania lymphadenitis

Leishmania lymphadenitis, an uncommon cause of lymphadenopathy, is usually diagnosed by surgical biopsy performed because of suspicion of lymphoma. The cytopathologic features of this disease do not appear to have been previously described. This paper describes the findings in seven cases of Leishmania lymphadenitis diagnosed by fine needle aspiration (FNA) cytology. The identification of Leishman-Donovan bodies in epithelioid cells in the aspirates led to the cytologic diagnosis of Leishmania lymphadenitis, which was histologically confirmed in all cases. Since this disease is self-limiting and requires no treatment, FNA diagnosis is especially useful in that the patient can be spared more invasive diagnostic techniques.     

Subclassification of localized Leishmania lymphadenitis in fine needle aspiration smears

OBJECTIVE: To describe the cytologic findings of localized Leishmania lymphadenitis and discuss the differential diagnosis. STUDY DESIGN: The study group consisted of 133 cases. All of them were diagnosed by fine needle aspiration (FNA) study. The ages ranged between 3 and 80 years, 102 were male and 31 female. Seventy lymph nodes were excised. RESULTS: The FNA smears revealed a polymorphic population of cells composed of lymphocytes, histiocytes, giant cells, abnormal plasma cells and tingible body macrophages. Leishman-Donovan (LD) bodies were identified in all cases, but their number differed from case to case. Granulomas, dendritic cells, mast cells and lymphoglandular bodies were identified in a substantial number of cases. Depending upon the presence of characteristic cytologic findings, the cases were divided into five major groups: acute inflammation with giant cells, histiocytic granulomas, epithelioid cell granulomas, plasma cell type and mixed histioplasmacytic type. CONCLUSION: Leishmaniasis is an uncommon cause of cervical lymphadenitis but should be considered in the differential diagnosis of unexplained lymphadenopathy in endemic countries. Demonstration of LD bodies is necessary for the diagnosis of this self-limited condition, for which no treatment is required.     

New clues to organ size control in plants

Plant growth has unparalleled importance for human civilization, yet we are only starting to gain an understanding of its mechanisms. The growth rate and final size of plant organs is determined by both genetic constraints and environmental factors. Regulatory inputs act at two control points: on proliferation; and on the transition between proliferation and differentiation. Cell-autonomous and short-range growth signals act within meristematic domains, whereas diffusible signals from differentiated parts to proliferating cells provide measures of geometry and size and channel environmental inputs.     

New clues to understand how CENP-A maintains centromere identity

The centromere is a specialized chromosomal region that directs the formation of the kinetochore, a huge protein assembly that acts as the attachment site for spindle microtubules and carries out chromosome movement during cell division. Centromere loss or the presence of extra centromeres adversely affect chromosome segregation and may result in aneuploidy, a condition found in many human tumors and a major cause of miscarriages and birth defects. Consequently, understanding the basis of centromere determination and propagation is of great relevance to both fundamental and clinical research. In recent years, it has become clear that centromeres are defined by the presence of a histone H3 variant known as Centromere Protein A, CENP-A, or CenH3. Much effort has been devoted to understanding the mechanisms that drive the assembly of CENP-A containing nucleosomes exclusively onto centromeric DNA, as well as the peculiar structure of these nucleosomes. We have recently developed an immunofluorescence-based assay that measures CENP-A incorporation in the centromeres of chromosomes assembled in Xenopus egg extracts. The spatial and temporal specificity of CENP-A deposition observed in human cells can be recapitulated in this in vitro system, making it suitable to dissect the precise role of the different factors that contribute to this pathway. Here, we discuss our results together with other recent advances in our understanding of the mechanisms that mediate centromere inheritance.     

New clues to the molecular pathogenesis of myelodysplastic syndromes

During the past few years our understanding of the genetic basis for the myelodysplastic syndromes (MDS) has improved significantly. A few subgroups have been studied in detail and the genetic alterations are now to a great extent revealed. In 5q- syndrome haploinsufficiency of the ribosomal gene RPS14 appears to cooperate with loss of two micro-RNAs miR-145 and miR-146 to induce key features of the disease. Some mutations are specific for certain categories of MDS while others, such as TET2 seem to occur across the various categories. JAK2 mutations are mainly found in patients with myeloproliferative characteristics. The prognostic implications of most of the novel mutations are not yet fully understood, moreover, functional studies are required in order to understand the interplay between the different lesions; how they give rise to the disease and how some may lead to disease evolution including leukemic transformation. An improved understanding of the pathophysiology of MDS may lead to the identification of suitable targets for future drug development.     

Fine needle aspiration cytologic diagnosis of toxoplasma lymphadenitis. A case report with detection of a Toxoplasma bradycyst in a Papanicolaou-stained smear

BACKGROUND: Fine needle aspiration (FNA) cytologic diagnosis of toxoplasmic lymphadenitis with demonstration of a tissue cyst containing bradyzoites has been very rarely reported. CASE: A 17-year-old female presented with a mobile, painless, 2-cm-diameter swelling over the right suprascapular area. Clinical diagnosis was lipoma. FNA smears showed features of reactive lymphoid hyperplasia, including tingible body macrophages and groups of epithelioid histiocytes. A Toxoplasma cyst with bradyzoites was also demonstrated in a Papanicolaou-stained smear. Following FNA cytodiagnosis, serologic tests revealed a high titer of IgG and the presence of IgM-specific antibodies to Toxoplasma gondii, indicating active/recent disease. CONCLUSION: FNA cytology is a valuable tool for the diagnosis of toxoplasmic lymphadenitis. Papanicolaou stain is appropriate for demonstration of the parasite. Serology is an excellent adjunct in clinching the diagnosis.     

New criterial for cytologic diagnosis of adenoid cystic carcinoma

OBJECTIVE: To formulate new criteria for adenoid cystic carcinoma (ADCC). STUDY DESIGN: The usefulness of 17 items for a cytologically definitive diagnosis of ADCC was examined. The frequency (- - +++) of the 17 items in 18 cases of ADCC and 10 non-ADCC cases (pleomorphic adenoma, basal cell adenoma, myoepithelioma and epithelial-myoepithelial carcinoma) that displayed mimicking cytology was examined cytologically. The total score for cases of ADCC was compared with that for non-ADCC cases. RESULTS: The 17 items included broad intercellular spaces with adhesion; green, granular/cobwebby cytoplasm with translucent intercellular matrix with Papanicolaou staining; coarse hyperchromatin with little nuclear clearing; indistinct and partially distinct nucleoli; small nuclei; broad, smooth margin (SM) space; and translucent M/HG/SM. All cases with a total score of > 21 were ADCC (93.8%). All cases with a total score of < 12 were non-ADCC (87.5%). CONCLUSION: The 17 items appear to be useful as new criteria for ADCC.     

Tetroxanes as New Agents against Leishmania amazonensis

Leishmaniasis is a neglected disease, caused by a parasite of Leishmania genus and widespread in the tropical and subtropical areas of the world. Currents drugs are limited due to their toxicity and parasite resistance. Therefore, the discovery of new treatment, more effective and less toxic, is urgent. In this study, we report the synthesis of six gem‐dihydroperoxides ( 2a – 2f ), with yields ranging from 10 % to 90 %, utilizing a new methodology. The dihydroperoxides were converted into ten tetroxanes ( 3a – 3j ), among which six ( 3b , 3c , 3d , 3g , 3h and 3j ) showed activity against intracellular amastigotes of Leishmania amazonensis. The cytotoxicity of all compounds was also evaluated against canine macrophages (DH82), human hepatoma (HepG2) and monkey renal cells (BGM). Most compounds were more active and less toxic than potassium antimonyl tartrate trihydrate, used as positive control. Amongst all tetroxanes, 3b (IC₅₀=0.64 μm ) was the most active, being more selective than positive control in relation to DH82, HepG2 and BGM cells. In summary, the results revealed a hit compound for the development of new drugs to treat leishmaniasis.     

New structural clues to substrate specificity in the "ubiquitin system"

A 2.5 A crystal structure of a complex between the SUMO-conjugating enzyme Ubc9 and a protein substrate has yielded fresh insight into the specificity of protein modification by SUMO and other ubiquitin-like proteins.     

Sensitivity of lymph node aspiration in localized cutaneous leishmaniasis due to Leishmania (Viannia) braziliensis.

Twenty nine patients with localized cutaneous leishmaniasis had lymph node and skin ulcer aspirations for culture of Leishmania with the modified Marzochís vacuum aspiratory technique. Sensitivity of lymph node aspiration was 58.6% and 34.5% for skin ulcer aspiration (P=0.06). Combined sensitivity of the two methods was 79.3%. There was no agreement between methods (Kappa Index = -0.084; CI95% -0,45; 0,28) showing the potential complementary roles in diagnostic approach.     

New pyrazolopyrimidine derivatives as Leishmania amazonensis arginase inhibitors

Arginase performs the first enzymatic step in polyamine biosynthesis in Leishmania and represents a promising target for drug development. Polyamines in Leishmania are involved in trypanothione synthesis, which neutralize the oxidative burst of reactive oxygen species (ROS) and nitric oxide (NO) that are produced by host macrophages to kill the parasite. In an attempt to synthesize arginase inhibitors, six 1-phenyl-1H-pyrazolo[3,4-d]pyrimidine derivatives with different substituents at the 4-position of the phenyl group were synthesized. All compounds were initially tested at 100 µM concentration against Leishmania amazonensis ARG (LaARG), showing inhibitory activity ranging from 36 to 74%. Two compounds, 1 (R=H) and 6 (R=CF₃), showed arginase inhibition >70% and IC₅₀ values of 12 µM and 47 µM, respectively. Thus, the kinetics of LaARG inhibition were analyzed for compounds 1 and 6 and revealed that these compounds inhibit the enzyme by an uncompetitive mechanism, showing K_is values, and dissociation constants for ternary complex enzyme-substrate-inhibitor, of 8.5 ± 0.9 µM and 29 ± 5 µM, respectively. Additionally, the molecular docking studies proposed that these two uncompetitive inhibitors interact with different LaARG binding sites, where compound 1 forms more H-bond interactions with the enzyme than compound 6. These compounds showed low activity against L. amazonensis free amastigotes obtained from mice lesions when assayed with as much as 30 µM. The maximum growth inhibition reached was between 20 and 30% after 48 h of incubation. These results suggest that this system can be promising for the design of potential antileishmanial compounds.     

Temperature-induced expression of proteins in Leishmania mexicana amazonensis. A 22-kDa protein is possibly localized in the mitochondrion

Temperature increase is an integral part of Leishmania life cycle, and plays a major role in stage transformation. Analysis of the temperature-dependent pattern of protein synthesis on two-dimensional gel electrophoresis shows that, in addition to the conserved heat-shock type of response in which expression of the major 70-kDa and 83-kDa heat-shock proteins is observed, a group of low-molecular-mass (17-40 kDa) proteins is induced in promastigotes of Leishmania mexicana amazonensis at elevated temperatures. Immuno-gold labelling with antibodies raised against the heat-induced 22-kDa proteins was localized mainly in the mitochondrion of Leishmania parasites, though labelling was observed also in the nucleus. The correlation of this finding with various reports on induction of mitochondrial enzymes in response to temperature stress in other organisms is discussed.     

Glycosylphosphatidylinositol biosynthetic enzymes are localized to a stable tubular subcompartment of the endoplasmic reticulum in Leishmania mexicana.

Glycosylphosphatidylinositols (GPI) are essential components in the plasma membrane of the protozoan parasite Leishmania mexicana, both as membrane anchors for the major surface macromolecules and as the sole class of free glycolipids. We provide evidence that L.mexicana dolichol-phosphate-mannose synthase (DPMS), a key enzyme in GPI biosynthesis, is localized to a distinct tubular subdomain of the endoplasmic reticulum (ER), based on the localization of a green fluorescent protein (GFP)-DPMS chimera and subcellular fractionation experiments. This tubular membrane (termed the DPMS tubule) is also enriched in other enzymes involved in GPI biosynthesis, can be specifically stained with the fluorescent lipid, BODIPY-C5-ceramide, and appears to be connected to specific subpellicular microtubules that underlie the plasma membrane. Perturbation of microtubules and DPMS tubule structure in vivo results in the selective accumulation of GPI anchor precursors, but not free GPIs. The DPMS tubule is closely associated morphologically with the single Golgi apparatus in non-dividing and dividing cells, appears to exclude luminal ER resident proteins and is labeled, together with the Golgi apparatus, with another GFP chimera containing the heterologous human Golgi marker beta1,2-N-acetylglucosaminyltransferase-I. The possibility that the DPMS-tubule is a stable transitional ER is discussed.     

Effect of HIV protease inhibitors on New World Leishmania

The incidence of HIV/Leishmania co-infection decreases after antiretroviral drug therapy; therefore, the in vitro and in vivo activity of three antiretroviral drugs against Leishmania (Viannia) braziliensis and L. (L.) amazonensis was evaluated. Different concentrations of indinavir (IDV), atazanavir (ATV), and ritonavir (RTV) were added to promastigote cultures, and the 50% inhibitory concentration (IC50) was determined. IDV and RTV were also evaluated against intracellular amastigotes, and the Infection Index determined. BALB/c mice, infected with L. (L.) amazonensis in the left footpad, were treated orally with IDV and RTV for 30days, and monitored by measuring the footpad thickness and parasite load of regional lymph nodes and spleen. For promastigotes, IDV exhibited an IC50 value of 100μM against L.(L.) amazonensis. The RTV IC50 for L. (L.) amazonensis and L. (V.) braziliensis were 40 and 2.3μM, respectively, and the ATV IC50 for L. (V.) braziliensis was 266μM. For intracellular amastigotes, IDV (25, 50, and 100μM) significantly decreased the Infection Index of L. (L.) amazonensis (56.8%, 47.9%, and 65.0%) and L. (V.) braziliensis (37.8%, 48.7%, and 43.2%). RTV (12.5, 25, and 50μM) decreased the infection index of L. (L.) amazonensis by 26.3%, 42.4%, and 44.0%, and that of L. (V.) braziliensis by 27.6%, 37.3%, and 39.2%. Antiretroviral-treated mice had a significant reduction in footpad thickness after the third week of IDV and after the fifth week of RTV treatment. However, there was no reduction in parasite load. These results suggest that IDV and RTV have anti-Leishmania activity, but only in higher concentrations.