RESUMO
BACKGROUND: Previous studies have demonstrated an association between obesity and the decreased male fertility. OBJECTIVE: to observe the mechanisms by which obesity affects semen quality. MATERIALS AND METHODS: A prospective study was performed including 47 male volunteers, of which 27 were obese group (body mass index >30 kg/m2 ) and 20 were eutrophic (body mass index between 18.5 and 25 kg/m2 ) controls. Sperm functional analysis was performed. The remaining seminal plasma was pooled-four pools per group- and submitted to proteomic analysis by liquid chromatography coupled to tandem mass spectrometry. Groups were compared by an unpaired Student's t-test. Differentially expressed proteins were submitted to functional enrichment analysis using the online platform PantherDB. RESULTS: Obese men presented decreased non-progressive motility, morphology, acrosome integrity, mitochondrial activity, and increased sperm DNA fragmentation. In proteomics analysis, 69 proteins were differentially expressed between the two groups. Among them, one protein was absent, 19 were down-regulated, 49 were up-regulated, and one was exclusive in the study group. The main functions enriched were as follows: negative regulation of the intrinsic pathway of apoptosis, activation of immune and inflammatory, antioxidant activity, among others. CONCLUSION: molecular pathways suggest there is a causative link, and that the effector mechanisms alter sperm metabolic status and defective testicular selection 5 mechanisms.
Assuntos
Obesidade/metabolismo , Sêmen/metabolismo , Espermatozoides/fisiologia , Adulto , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Proteoma , Análise do Sêmen , Adulto JovemRESUMO
Varicocelectomy is associated to improved semen quality and sperm functional quality, but individual response is highly variable. Thus, a prospective study was performed including 25 men who collected a semen sample before and 12â¯months after subinguinal microsurgical varicocelectomy. Semen analysis, sperm functional analysis, and seminal plasma proteomic analysis was performed before and 12â¯months after varicocelectomy, and according to improvement or not of semen quality (positive and negative outcome). Varicocelectomy led to an increase in semen volume and sperm count, morphology, and mitochondrial activity. In the pre- vs. post-samples, 698 proteins were quantified - 91 differentially expressed after varicocelectomy. In the positive vs. negative outcome analysis, 647 proteins were identified - 151 differentially expressed in the negative outcome group and 30 differentially expressed in the positive outcome group. Tripeptidyl peptidase-1 offered a predictive value for outcome, with an area under a ROC curve of 84.5%. It seems TPP1 is an outcome predictor for varicocelectomy in adults. More importantly, this study demonstrates that the seminal plasma proteome is different in men with varicocele when compared to post-treatment samples from the same individuals. Understanding and monitoring the molecular mechanisms of semen may further establish therapeutic options for these men. SIGNIFICANCE: Although several large-scale studies have demonstrated varicocele is unequivocally associated to male infertility, these same studies have also demonstrated that varicocele is not a determinant of male infertility. We have yet to answer the question of why don't all men with varicocele present with infertility. Varicocele treatment improves semen quality, but its results are variable, and one cannot know who will and who will not benefit from surgical treatment. Results from this study strongly advance a concept that our previous studies have shown: that men with varicocele present an inflammatory semen profile. We have further demonstrated that men operated for varicocele present a decrease in this inflammatory profile, and that when they do not, semen quality remains unaltered. Trypeptidil peptidase-1, a seminal protein, was 3-fold higher in men with a positive outcome after the procedure, when compared to men with a negative outcome. Therefore, inflammation seems to be a central point to varicocele-derived male infertility.
Assuntos
Infertilidade Masculina/metabolismo , Proteoma/metabolismo , Proteômica , Sêmen/metabolismo , Proteínas de Plasma Seminal/metabolismo , Varicocele/metabolismo , Adulto , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Tripeptidil-Peptidase 1 , Varicocele/patologiaRESUMO
The goal of this study was to determine seminal plasma biomarkers of testicular function in adolescents with varicocoele and to verify enriched gene ontology terms associated to these differential proteomes. An observational study was carried out in an academic research environment. A total of 77 adolescent patients were recruited from a local public school, of which 23 were without varicocoele and with normal semen analysis (control group), 37 were with varicocoele and normal semen (VNS) parameters, and 17 were with varicocoele and altered semen (VAS) parameters. Two semen collections were provided with a 1-week interval, after 2-5 days of ejaculatory abstinence. Seminal plasma proteins were identified and quantified utilizing a label-free shotgun proteomics approach, generating (i) proteins differentially expressed in each group (control, VNS, and VAS) and putative biomarkers using multivariate statistics followed by discriminant analysis. Confirmatory analysis was performed for two proteins by western blotting. Enriched biological processes and molecular functions were determined using gene ontology analysis. In total, 541 proteins were identified and quantified: 108 exclusive or overexpressed in controls, 26 in the VNS group, and 13 in the VAS group. The suggested biomarkers are Cab45/SDF4 (Q9BRK5), protein lefty-1 (O75610), DNase I (P24855), PAP2-alpha (O14494), IBP-7 (Q16270), HDC (P01860), and CRISP-3 (P54108). Western blotting results showed that Cab45 was significantly underexpressed in both varicocoele groups, and CRISP-3 was significantly overexpressed in seminal plasma of adolescents with VAS. In conclusion, specific biomarkers of spermatogenesis and homeostasis are observed in adolescents without varicocoele, and the presence of a palpable varicocoele progressively shifts these adolescents toward initially an immune response, and finally toward a chronic inflammatory profile. This shift is accompanied by decreased semen quality.
Assuntos
Infertilidade Masculina/metabolismo , Proteômica , Proteínas de Plasma Seminal/metabolismo , Varicocele/metabolismo , Adolescente , Biomarcadores/metabolismo , Humanos , Masculino , Análise do SêmenRESUMO
Urinary tract infection (UTI) is a frequent disease of humans and pets and has extra-intestinal pathogenic Escherichia coli (ExPEC) strains as one of the main etiologic agent. ExPEC are characterized by specific virulence factors and are related to a heterogeneous group of human and animal disorders, besides to be a relevant participant in the dissemination of antimicrobial resistance. The purpose of this study was to characterize E. coli strains isolated from UTI of dogs and cats for serotypes, virulence markers, phylogenetic groups and sensitivity to antimicrobial drugs. E. coli was identified as the etiologic agent of UTI in urine samples of 43 pets (7 cats and 36 dogs). Serogroups O2, O4 and O6 corresponded to more than one third of the isolates, being 62% of the total strains classified as B2, 18% as D, 16% as B1 and 4% as A. The iucD (22%), fyuA (80%), traT (51%) and cvaC (20%) genes were distributed among the four phylogenetic groups, whereas the papC/papEF (47%) and malX (67%) genes were found only in groups B2 and D. There were a high number of resistant strains, with 76% of the strains belonging to groups A, B1 and D characterized as multidrug resistant (MDR), whereas only 21% had this phenotype in the group B2. The ExPEC strains isolated in this study displayed pathotypic and phylogenetic similarities with human isolates and high percentages of drug resistance. The finding of MDR ExPEC strains suggests implications for animal and public health and deserves more investigations.
Assuntos
Doenças do Gato/microbiologia , Doenças do Cão/microbiologia , Infecções por Escherichia coli/veterinária , Escherichia coli , Filogenia , Infecções Urinárias/veterinária , Fatores de Virulência/genética , Animais , Antibacterianos/farmacologia , Brasil , Gatos , Cães , Resistência Microbiana a Medicamentos , Escherichia coli/classificação , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/patogenicidade , Infecções por Escherichia coli/microbiologia , Genótipo , Fenótipo , Sorotipagem , Infecções Urinárias/microbiologiaRESUMO
STUDY QUESTION: Does the seminal plasma proteomic profile and functional enrichment of gene ontology terms change after microsurgical varicocelectomy? Are there any potential targets for diagnosis or therapeutic intervention in varicocele? SUMMARY ANSWER: A shift in state from a responsive-to-stress condition before varicocele correction to a responsive-to-environment condition after varicocelectomy was observed in enriched proteomic pathways. WHAT IS KNOWN ALREADY: Varicocele may lead to many adverse effects, including failure of testicular growth and development, and is associated with decreased semen quality and increased semen oxidative stress. Varicocelectomy is the treatment of choice, and is associated with improved semen quality, but little is known regarding the underlying molecular mechanisms and post-genomic pathways following intervention. STUDY DESIGN, SIZE, DURATION: A prospective study was carried out including 18 adult men with varicocele. These patients provided one semen sample before they were submitted for bilateral varicocele repair through microsurgical varicocelectomy, and one other semen sample 90 days after the surgery. PARTICIPANTS/MATERIALS, SETTING, METHODS: An aliquot of each semen sample was used for unbiased proteomics analysis by a label-free quantitative approach (2D nanoUPLC-ESI-MS(E)). Samples were pooled according to group (normalized to protein content) and run in quadruplicate. These quadruplicate runs provided degrees of freedom in order to compare groups using a non-parametric Mann-Whitney test for quantified proteins. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 316 proteins were quantified or identified, of which 91 were exclusively identified or quantified in one of the groups (53 in the pre- and 38 in the post-varicocelectomy group), and 68 were quantified in both groups and submitted to statistical analysis, of which 5 were overrepresented in the pre-varicocelectomy group (P < 0.05). In enriched functional analysis, binding and response to stimulus functions were enriched in a common cluster (present in both groups), nitric oxide metabolism and tetratricopeptide repeat domain-binding functions were enriched in the pre-varicocelectomy group, and response to reactive oxygen species, gluconeogenesis, nicotinamide adenine dinucleotide-binding and protein stabilization were enriched in the post-varicocelectomy. LIMITATIONS, REASONS FOR CAUTION: Because a shotgun proteomics analysis was chosen in order to generate a list of putative biomarkers, a targeted follow-up study should be performed to confirm these biomarkers. WIDER IMPLICATIONS OF THE FINDINGS: The proteins found in both groups possess functions usually found in human semen. The enriched function analysis demonstrated a shift back to homeostasis after varicocelectomy, suggesting that varicocele correction promotes return of semen to a physiological state. STUDY FUNDING/COMPETING INTEREST(S): The funding for this project was received from the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) as a scholarship for Ms Camargo. There was no conflict of interest.
Assuntos
Alostase , Microcirurgia , Proteínas de Plasma Seminal/metabolismo , Cordão Espermático/cirurgia , Varicocele/metabolismo , Varicocele/cirurgia , Adulto , Biomarcadores/química , Biomarcadores/metabolismo , Cromatografia Líquida de Alta Pressão , Perfilação da Expressão Gênica , Humanos , Masculino , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Mapeamento de Peptídeos , Estudos Prospectivos , Proteômica/métodos , Análise do Sêmen , Proteínas de Plasma Seminal/química , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em Tandem , Adulto JovemRESUMO
Individuals with Down syndrome (DS) carry three copies of the Cystathionine ß-synthase (CßS) gene. The increase in the dosage of this gene results in an altered profile of metabolites involved in the folate pathway, including reduced homocysteine (Hcy), methionine, S-adenosylhomocysteine (SAH) and S-adenosylmethionine (SAM). Furthermore, previous studies in individuals with DS have shown that genetic variants in genes involved in the folate pathway influence the concentrations of this metabolism's products. The purpose of this study is to investigate whether polymorphisms in genes involved in folate metabolism affect the plasma concentrations of Hcy and methylmalonic acid (MMA) along with the concentration of serum folate in individuals with DS. Twelve genetic polymorphisms were investigated in 90 individuals with DS (median age 1.29 years, range 0.07-30.35 years; 49 male and 41 female). Genotyping for the polymorphisms was performed either by polymerase chain reaction (PCR) based techniques or by direct sequencing. Plasma concentrations of Hcy and MMA were measured by liquid chromatography-tandem mass spectrometry as previously described, and serum folate was quantified using a competitive immunoassay. Our results indicate that the MTHFR C677T, MTR A2756G, TC2 C776G and BHMT G742A polymorphisms along with MMA concentration are predictors of Hcy concentration. They also show that age and Hcy concentration are predictors of MMA concentration. These findings could help to understand how genetic variation impacts folate metabolism and what metabolic consequences these variants have in individuals with trisomy 21.
Assuntos
Síndrome de Down/genética , Ácido Fólico/sangue , Polimorfismo Genético , Adolescente , Adulto , Brasil , Criança , Pré-Escolar , Cromossomos Humanos Par 21/genética , Síndrome de Down/sangue , Feminino , Ácido Fólico/metabolismo , Frequência do Gene , Estudos de Associação Genética , Genótipo , Homocisteína/sangue , Humanos , Lactente , Modelos Lineares , Masculino , Ácido Metilmalônico/sangue , Análise de Sequência de DNA , Deficiência de Vitamina B 12/sangue , Deficiência de Vitamina B 12/genética , Adulto JovemRESUMO
INTRODUCTION: Although much is known about the increased levels of the 21-hydroxylase substrates 17-hydroxyprogesterone (17OHP) and 21-deoxycortisol (21DF) - the biochemical markers of all forms of 21-hydroxylase deficiency (21OHD), only limited information is available on the zona fasciculata (ZF) products distal to the enzymatic block: 11-deoxycortisol (S), 11-deoxycorticosterone (DOC), and corticosterone (B). OBJECTIVE: To investigate whether basal and post-ACTH levels of S, DOC, and B and the 21-hydroxylase precursor-to-product ratios determined by tandem mass spectrometry preceded by high-performance liquid chromatography separation (liquid chromatography-tandem mass spectrometry) could disclose distinct profiles in genotypically confirmed classic (no.=14) and non-classic (NC) (no.=18) patients, heterozygote carriers (no.=61) and wildtypes (WT) (no.=27) for 21OHD. RESULTS: Salt wasting (SW) and simple virilizing (SV) had higher basal levels of DOC with no further increase in response to ACTH. Stimulated DOC was similar in 21OHD patients and carriers but was reduced as compared to WT. ACTH-stimulated B increased gradually from SW and SV through WT. The post-ACTH 21DF/B ratio was able to detect 92% of the carriers among WT. All NC patients could be detected by post-ACTH 17OHP/DOC and 21DF/B, with no overlap with 21OHD carriers. CONCLUSION: Although 21-hydroxylase is a key enzymatic step in both 17-hydroxy and 17-deoxy pathways of ZF, the reaction is mostly affected in the latter pathway, leading to a significant impairment of B production, which may further characterize the 21OHD subtypes. Also, the precursor-to-product ratios, particularly 21DF/B, can demonstrate the distinctive outline of 21OHD subtypes, including carriers and normal subjects.
Assuntos
17-alfa-Hidroxiprogesterona/metabolismo , Hiperplasia Suprarrenal Congênita/genética , Hiperplasia Suprarrenal Congênita/metabolismo , Cortodoxona/metabolismo , Heterozigoto , Esteroide 21-Hidroxilase/metabolismo , Zona Fasciculada/metabolismo , Hiperplasia Suprarrenal Congênita/fisiopatologia , Adulto , Portador Sadio , Corticosterona/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Esteroide 21-Hidroxilase/genética , Adulto Jovem , Zona Fasciculada/químicaRESUMO
Myelodysplastic syndromes (MDS) are a group of clonal stem cell diseases characterized by ineffective hematopoiesis, bone marrow hyperproliferation, cytopenias in peripheral blood and risk of transformation into acute leukemia. We decided to investigate the effects of a soy concentrate on MDS patients based on the follow-up results of a 61 year-old Japanese female patient who was diagnosed with MDS and refractory cytopenia with multilineage dysplasia in 2003 (hemoglobin = 11g/dL; white blood cells count = 2,500/uL and platelets = 25,000/uL; marrow with mild dysplasia and normal karyotype; paroxysmal nocturnal hemoglobinuria was excluded). She started using soy as a dietary supplementation in May 2004 and presented a gradual increment in blood counts, achieving normalization approximately eight months afterwards. Among the soy components, the main compounds with anti-carcinogenic activity are the isoflavones (genistein and daidzein). Based on these lines of evidence, we proposed to administer daily a standard soy concentrate to 14 MDS out-patients for a minimum period of three months and maximum of 12 months, in an attempt to evaluate prospectively the possible increase in hemoglobin, neutrophils and platelet counts. A historical control group was used to compare results. The use of a soy concentrate in a standardized manner was associated with an increase in neutrophil and/or platelet counts in some cases, but spontaneous increments were also observed in historical controls. This preliminary study does not allow establishing a relation between soy supplementation and blood cell count increase.
As síndromes mielodisplásicas (SMD) são um grupo das doenças clonais de células-tronco caracterizado por hematopoese ineficaz, hiperproliferação de medula óssea, citopenias no sangue periférico e risco de transformação para leucemia aguda. Decidimos investigar os efeitos de um concentrado de soja em pacientes com SMD com base no fato de termos o seguimento de uma paciente japonesa, de 61 anos de idade, que foi diagnosticada em 2003 com SMD, citopenia refratária com displasia subtipo multilinhagens (hemoglobina = 11 g/dL; contagem de glóbulos brancos = 2.500/uL e plaquetas = 25.000/uL; medula com displasia leve e cariótipo normal; hemoglobinúria paroxística excluída), e que começou a usar a soja como suplemento alimentar em maio de 2004, apresentando gradual aumento da contagem das células sanguíneas, atingindo a normalização cerca de oito meses depois. Entre os componentes da soja, os principais compostos com propriedades anticarcinogênese são as isoflavonas (Ge nisteína e daidzeína). Com base nessas linhas de evidência, foi proposto oferecer diariamente um concentrado de soja padrão, por um período mínimo de três meses e máximo de doze meses, a 14 pacientes ambulatoriais, na tentativa de avaliar, prospectivamente, o possível aumento de hemoglobina, neutrófilos e plaquetas. Um grupo controle histórico foi utilizado para comparar os resultados. O uso de um concentrado de soja de forma padronizada foi associado ao aumento na contagem de neutrófilos e/ou de plaquetas em alguns casos, mas aumentos espontâneos também foram observados em controles históricos. Este estudo preliminar não permite estabelecer relação entre o uso de soja e o aumento na contagem sanguínea.
RESUMO
The occurrence of non-mosaic double trisomy is exceptional in newborns. In this paper, a 48,XXY,+21 child, the parental origin of the extra chromosomes and the evaluation of the maternal folate metabolism are presented. The infant was born to a 13-year-old mother and presented with the typical clinical features of Down syndrome (DS). The origin of the additional chromosomes was maternal and most likely resulted from errors during the first meiotic division. Molecular analysis of 12 genetic polymorphisms involved in the folate metabolism revealed that the mother is heterozygous for the MTHFR C677T and TC2 A67G polymorphisms, and homozygous for the mutant MTRR A66G polymorphism. The maternal homocysteine concentration was 4.7 miromol/L, a value close to the one considered as a risk factor for DS in our previous study. Plasma methylmalonic acid and serum folate concentrations were 0.17 micromol/L and 18.4 ng/mL, respectively. It is possible that the presence of allelic variants for the folate metabolism and Hey concentration might have favored errors in chromosomal disjunction during gametogenesis in this young mother. To our knowledge, this is the first patient with non-mosaic Down-Klinefelter born to a teenage mother, resulting from a rare fertilization event combining an abnormal 25,XX,+21 oocyte and a 23,Y spermatozoon.
Assuntos
Alelos , Aneuploidia , Cromossomos Humanos X/genética , Cromossomos Humanos Y/genética , Síndrome de Down/genética , Ferredoxina-NADP Redutase/genética , Ácido Fólico/sangue , Síndrome de Klinefelter/genética , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Polimorfismo Genético/genética , Gravidez na Adolescência/genética , Aberrações dos Cromossomos Sexuais , Trissomia , Adolescente , Brasil , Análise Mutacional de DNA , Síndrome de Down/diagnóstico , Feminino , Triagem de Portadores Genéticos , Genótipo , Cardiopatias Congênitas/diagnóstico , Cardiopatias Congênitas/genética , Homocisteína/sangue , Homozigoto , Humanos , Lactente , Síndrome de Klinefelter/diagnóstico , Masculino , Meiose , Ácido Metilmalônico/sangue , Não Disjunção Genética/genética , GravidezRESUMO
Central giant cell lesion (CGCL) is a reactive bone lesion that occurs mainly in the mandible, characterized by the multinucleated osteoclast-like giant cells in a background of oval to spindle-shaped mononuclear cells. The etiology is unknown and occurs more commonly in young adults. Cherubism, a rare disease found predominantly in females has histologic characteristics indistinguishable from those of CGCL and is caused by mutations mostly present in exon 9 of the SH3BP2 gene. In this study, we investigated four cases of CGCL and one case of cherubism. DNA was extracted from peripheral blood and tumor tissue and all coding and flanking regions of the SH3BP2 amplified by PCR and directly sequenced to identify underlying mutations. Two novel mutations were found; a heterozygous missense mutation c.1442A>T (Q481L) in exon 11 in one sporadic case of CGCL and a heterozygous germline and tumor tissue missense mutation c.320C>T (T107M) in exon 4 in one patient with cherubism. These findings open a new window to investigate the possible relationship between the pathogenesis of the cherubism and CGCL.