Levels of seminal tRNA-derived fragments from normozoospermic men correlate with the success rate of ART.

Decreased fertility is becoming an important social and medical problem and the male factor is involved in at least half of infertility cases. Since conventional semen analysis provides limited prediction of male fertility; in this work, we evaluated the potential use of seminal small RNAs (sRNA) as markers of semen quality in ART. Our bioinformatic analyses of available sRNA-seq databases showed that the most abundant sRNA species in seminal plasma of normozoospermic men are tRNA-derived fragments (tRFs), a novel class of regulatory sRNAs. These molecules not only exert their function within cells but also are released into the extracellular environment where they could carry out signaling functions. To evaluate whether the assessment of seminal tRFs in normozoospermic men has a predictive value for the clinical outcome in ART, we performed a prospective study with couples who underwent ICSI cycles with donated oocytes. The results obtained demonstrated that levels of 5'tRF-Glu-CTC, 5'tRF-Lys-CTT, and 5'tRF-Gly-GCC are significantly elevated in seminal samples from cases with repeated failed ICSI cycles, suggesting a potential association between increased seminal tRFs and unexplained male infertility. Interestingly, these tRFs showed a negative association with seminal testosterone, highlighting their involvement in male endocrinology. Our findings also suggest that tRFs could play a role in modulating male reproductive function in response to physiological stress since they showed significant associations with the levels of sperm DNA fragmentation in couples that achieved pregnancy but not in cases with failed ICSI cycles where seminal cortisol levels correlate with sperm quality.

CALR, JAK2 and MPL mutation status in Argentinean patients with BCR-ABL1- negative myeloproliferative neoplasms.

OBJECTIVES: To establish the frequency of JAK2, MPL and CALR mutations in Argentinean patients with BCR-ABL1-negative  myeloproliferative neoplasms (MPN) and to compare their clinical and haematological features. METHODS: Mutations of JAK2V617F, JAK2 exon 12, MPL W515L/K and CALR were analysed in 439 Argentinean patients with BCR-ABL1-negative MPN, including 176 polycythemia vera (PV), 214 essential thrombocythemia (ET) and 49 primary myelofibrosis (PMF). RESULTS: In 94.9% of PV, 85.5% ET and 85.2% PMF, we found mutations in JAK2, MPL or CALR. 74.9% carried JAK2V617F, 12.3% CALR mutations, 2.1% MPL mutations and 10.7% were triple negative. In ET, nine types of CALR mutations were identified, four of which were novel. PMF patients were limited to types 1 and 2, type 2 being more frequent. DISCUSSION: In ET, patients with CALR mutation were younger and had higher platelet counts than those with JAK2V617F and triple negative. In addition, JAK2V617F patients had high leucocyte and haemoglobin values compared with CALR-mutated and triple-negative patients. In PMF, patients with mutant CALR were associated with higher platelet counts. CONCLUSION: Our study underscores the importance of JAK2, MPL and CALR genotyping for accurate diagnosis of patients with BCR-ABL1-negative MPN.

Benznidazole modulates cell proliferation in acute leukemia cells.

CONTEXT: We have previously reported that benznidazole (BZL), known for its trypanocidal action, has anti-proliferative activity against different cell lines like HeLa and Raw 264.7 among others. At the moment, it has not been reported if the anti-proliferative effect of BZL is similar for non-adherent hematopoietic cells like was reported for adherent cancer cell lines. OBJECTIVE: We aimed to investigate the efficacy of BZL on the growth of the leukemic cell lines THP-1 and OCI/AML3. MATERIALS AND METHODS: We evaluated cell proliferation by [³H]-thymidine incorporation and MTT reduction as well as cell death by lactate dehydrogenase (LDH) activity. We assessed apoptosis by flow cytometry for detection of annexin V-positive and propidium iodide-negative cells, along with nuclear morphology by diamidino-2-phenolindole (DAPI) staining. Western blot studies were performed to evaluate changes in cell cycle proteins in BZL-treated cells. RESULTS: BZL significantly reduced proliferation of both cell lines without inducing cell death. Likewise it produced no significant differences in apoptosis between treated cells and controls. In addition, flow cytometry analysis indicated that BZL caused a larger number of THP-1 cells in G0/G1 phase and a smaller number of cells in S phase than controls. This was accompanied with an increase in the expression of the CDK inhibitor p27 and of cyclin D1, with no significant differences in the protein levels of CDK1, CDK2, CDK4, cyclins E, A and B as compared to controls. CONCLUSION: BZL inhibits the proliferation of leukemic non-adherent cells by controlling cell cycle at G0/G1 cell phase through up-regulation of p27.

[Hemolytic anemia due to hemoglobin Evans in an Argentinean family]. / Anemia hemolítica causada por hemoglobina Evans en una familia argentina.

Unstable hemoglobins are structural variants of the hemoglobin molecule, mostly originated by single amino-acid replacement in some globin chains. These changes affect molecule stability, leading to loss of solubility, precipitation, and cellular lysis. Patients carrying these unstable hemoglobins may present mild to severe chronic hemolytic anemia. Hemoglobin Evans is an unstable variant originated by replacement of valine with methionine at position 62 of the α-globin chain. We have identified this variant in a girl with an acute hemolytic crisis associated to pharyngitis, as well as in two of her family members. This is the third case of hemolytic anemia due to hemoglobin Evans reported in the literature.

Compound heterozygosity of Hb Q India (α64 (E13) ASP→HIS) and -α3,7 thalassemia. First report from Argentina / Heterocigosis de la Hb Q India (α64 (E13) ASP →HIS) y -α3,7 thalassemia. Primer informe desde Argentina

Hemoglobine (Hb) Q-India is an innocuous αglobin variant: α64 Asp → His. DNA sequencing studies have shown that the Hb Q India mutation is GAC → CAC in codon 64 of the α1 gene. Hb Q-India is a well-known hemoglobin variant in South-East Asia but only isolated case reports exist in literature to describe this rare entity in the rest of de world. The variant has been found with various forms of αand ß thalassemia. This hemoglobin has the same electrophoretic mobility as Hb S. We report, for the first time, the identification of Hb Q-India in an Argentinian woman (her parents came from Gibraltar), referred to our laboratory bearing a mild microcytic hypocromic anemia; a co-inherited α+ thalassemia (-α3.7 th) was also found.

La hemoglobina (Hb) Q India es una hemoglobina anormal e inocua que afecta la cadena α de esta. Los análisis de secuencia han demostrado que la mutación se encuentra en el codon 64 GAC → CAC del gen α1. Si bien es una variante muy conocida en el sudeste asiático, solo se han reportado pocos casos en el resto del mundo. Esta hemoglobina anormal se ha encontrado asociada con diversas formas de α y ß talasemia y su posición electroforética es idéntica a la de la Hb S. Reportamos, por primera vez, la identificación de la Hb Q India en una mujer Argentina (cuyos padres procedían del Peñón de Gibraltar), enviada a nuestro laboratorio por padecer de anemia microcítica hipocrómica, en la que se encontró también la coexistencia de α+ talasemia (-α3,7 th).

Anemia hemolítica causada por hemoglobina evans en una familia argentina / Hemolytic anemia due to hemoglobin evans in an Argentinean family

Las hemoglobinas inestables son variantes estructurales de lamolécula de hemoglobina originadas, en su mayoría, por sustitucionespuntuales de aminoácidos en alguna de las cadenas de globina. Estos cambios afectan la estabilidad de la molécula,causan pérdida de la solubilidad y precipitación dentro del eritrocito, lo cual provoca su destrucción acelerada. Desde el punto de vista clínico, las hemoglobinas inestables puedenpresentar anemia hemolítica crónica de gravedad variable.La hemoglobina Evans es una hemoglobina inestable causadapor la sustitución de valina por metionina en la posición 62 de la cadena de alfa globina. Hemos identificado esta variante en una niña con crisis hemolítica aguda asociada a faringitis y en dos miembros de su grupo familiar. Éste es el tercer caso de anemia hemolítica congénita causada por hemoglobina Evans comunicado en la bibliografía mundial.

Unstable hemoglobins are structural variants of the hemoglobin molecule, mostly originated by single amino-acid replacement in some globin chains. These changes affect molecule stability, leading to loss of solubility, precipitation, and cellular lysis. Patients carrying these unstable hemoglobins may present mild to severe chronic hemolytic anemia. Hemoglobin Evans is an unstable variant originated by replacement of valine with methionine at position 62 of the αa-globin chain. We have identified this variant in a girl with an acute hemolytic crisis associated to pharyngitis, as well as in two of her family members. This is the third case of hemolytic anemia due to hemoglobin Evans reported in the literature.

Hb S-San Martin: a new sickling hemoglobin with two amino acid substitutions [ß6(A3)Glu→Val;ß105(G7)Leu→Pro] in an Argentinean family.

A new sickling hemoglobin (Hb) detected in an Argentinean family from San Martín, Buenos Aires, Argentina, is hereby described. Two mutations were identified on the same ß-globin gene resulting in a new variant named Hb San Martin. One mutation was found on exon 1, corresponding to Hb S [ß6Glu→Val, GAG>GTG] and the second one on exon 3 at ß105(G7)Leu→Pro, CTC>CCC. The replacement of leucine by proline will likely impair the structure breaking helix G and causing instability of the molecule and the clinical manifestations typical of unstable Hbs. The mutation at ß105 seemed to be a de novo one in our patients, arising on a previously mutated gene, due to the fact that Hb S is the most frequent structural variant.

[Severe hemolytic anemia due to hemoglobin Hammersmith]. / Anemia hemolítica grave causada por hemoglobina Hammersmith.

Most of the hemoglobin variants are the result of single amino acid replacement in one of the globin chains. In many cases, these hemoglobinopathies are harmless, while in others they determine alterations in the physical and chemical properties, raising clinical manifestations of variable severity. In the unstable hemoglobinopathies, the changes reduce solubility, inducing the formation of precipitates of denaturated hemoglobin (Heinz bodies), which damage the membrane and finally destroy the red blood cells prematurely. Up to now, more than 150 different unstable hemoglobins have been described; most of them cause chronic hemolysis, increased by infections or drugs. We report the clinical presentation of an unstable hemoglobin (hemoglobin Hammersmith) in a girl with severe hemolytic anemia, splenomegaly and red blood cell requirement.

Anemia hemolítica grave causada por hemoglobina Hammersmith / Severe hemolytic anemia due to hemoglobin Hammersmith

Las variantes estructurales de la hemoglobina resultan, en su mayoría, de sustituciones concretas de aminoácidos en una de las cadenas de globina. En muchos casos, estas hemoglobinopatías son inocuas, pero en otros determinan alteraciones de las propieddes físicas y químicas, cuyas manifestaciones clínicas son de gravedad y variable. En el caso de las hemoglobinas inestables, las alteraciones disminuyen la solubilidad y facilitan la formación de complejos de hemoglobina precipitada y desnaturalizada (cuerpos de Heinz), que ocasionan el daño de la membrana, y finalmente, la destrucción prematura de los eritrocitos. Hasta la actualidad se han descrito más de 150 hemogloginas inestables diferentes, la mayoría ocasionan hemólisis crónica, exacerbada por infecciones o la ingesta de medicamentos. Presentamos un caso clínico de hemoglobina inestable (hemoglobina Hammersmith) en una niña con anemia hemolítica grave, esplenomegalia y requerimiento transfusional.

Anemia hemolítica grave causada por hemoglobina Hammersmith / Severe hemolytic anemia due to hemoglobin Hammersmith

Las variantes estructurales de la hemoglobina resultan, en su mayoría, de sustituciones concretas de aminoácidos en una de las cadenas de globina. En muchos casos, estas hemoglobinopatías son inocuas, pero en otros determinan alteraciones de las propieddes físicas y químicas, cuyas manifestaciones clínicas son de gravedad y variable. En el caso de las hemoglobinas inestables, las alteraciones disminuyen la solubilidad y facilitan la formación de complejos de hemoglobina precipitada y desnaturalizada (cuerpos de Heinz), que ocasionan el daño de la membrana, y finalmente, la destrucción prematura de los eritrocitos. Hasta la actualidad se han descrito más de 150 hemogloginas inestables diferentes, la mayoría ocasionan hemólisis crónica, exacerbada por infecciones o la ingesta de medicamentos. Presentamos un caso clínico de hemoglobina inestable (hemoglobina Hammersmith) en una niña con anemia hemolítica grave, esplenomegalia y requerimiento transfusional.(AU)

Detection of the nucleophosmin gene mutations in acute myelogenous leukemia through RT-PCR and polyacrylamide gel electrophoresis.

OBJECTIVES: Mutations in the C-terminal region of the nucleophosmin (NPM1) gene occur in approximately 60% of acute myeloid leukemia (AML) cases with normal karyotype and represent the most common genetic lesion presently known in this disease. Because of their frequency and favorable impact on prognostic outcome, screening for this aberration is currently recommended in routine diagnostic characterization of AML. Several techniques enabling to detect NPM1 mutation have been reported, but all require sophisticated equipment, which represent an obstacle particularly in countries with limited resources. METHODS: We designed an RT-PCR strategy to amplify NPM1 exon 12 followed by electrophoresis and fragment visualization on polyacrylamide gels to discriminate a 4-5 bp size difference resulting from mutations in this gene. A hemi-nested method was designed to increase sensitivity for the study of minimal residual disease (MRD). RESULTS: The assay enabled specific detection of NPM1 mutations in 12/36 patients. A 10(-2) sensitivity level was obtained using one amplification round, while the hemi-nested PCR approach yielded a 10(-5) sensitivity level, therefore proving useful to assess MRD in patients carrying the mutation. The results were independently validated in 24 AML cases by sequencing analysis. CONCLUSIONS: This simple and low-cost assay may integrate diagnostic work-up of AML and could be used for assessment of response to therapy in patients with NPM1 mutations.