RESUMO
AIMS: The aim of this work was to refine the taxonomy and the functional characterization of publicly available Lactiplantibacillus plantarum complete genomes through a pan-genome analysis. Particular attention was paid in depicting the probiotic potential of each strain. METHODS AND RESULTS: Complete genome sequence of 127 L. plantarum strains, without detected anomalies, was downloaded from NCBI. Roary analysis of L. plantarum pan-genome identified 1436 core, 414 soft core, 1858 shell and 13,203 cloud genes, highlighting the 'open' nature of L. plantarum pan-genome. Identification and characterization of plasmid content, mobile genetic elements, adaptative immune system and probiotic marker genes (PMGs) revealed unique features across all the L. plantarum strains included in the present study. Considering our updated list of PMGs, we determined that approximatively 70% of the PMGs belongs to the core/soft-core genome. CONCLUSIONS: The comparative genomic analysis conducted in this study provide new insights into the genomic content and variability of L. plantarum. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides a comprehensive pan-genome analysis of L. plantarum, including the largest number (N = 127) of complete L. plantarum genomes retrieved from publicly available repositories. Our effort aimed to determine a solid reference panel for the future characterization of newly sequenced L. plantarum strains useful as probiotic supplements.
Assuntos
Lactobacillus plantarum , Probióticos , Genoma Bacteriano , Genômica , Lactobacillaceae , Lactobacillus plantarum/genética , Análise de Sequência de DNARESUMO
BACKGROUND: Acute or chronic irreversible respiratory failure may occur in patients undergoing pneumonectomy. Aim of this study was to determine transcriptome expression changes after experimental pneumonectomy in swine model. Experimental left pneumonectomy was performed in five pigs under general anaesthesia. Both the resected and the remaining lung, after 60 post-operative completely uneventful days, underwent genome-wide bulk RNA-Sequencing (RNA-Seq). RESULTS: Histological analysis showed dilation of air spaces and rupture of interalveolar septa. In addition, mild inflammation, no fibrosis, radial stretch of the bronchus, strong enlargement of airspaces and thinning of the blood supply were observed. Bioinformatic analyses of bulk RNA-Seq data identified 553 Differentially Expressed Genes (DEGs) at adjusted P-value below 0.001, between pre- and post-pneumonectomy. The top 10 up-regulated DEGs were Edn1, Areg, Havcr2, Gadd45g, Depp1, Cldn4, Atf3, Myc, Gadd45b, Socs3; the top 10 down-regulated DEGs were Obscn, Cdkn2b, ENSSSCG00000015738, Prrt2, Amer1, Flrt3, Efnb2, Tox3, Znf793, Znf365. Leveraging digital cytometry tools, no difference in cellular abundance was found between the two experimental groups, while the analysis of cell type-specific gene expression patterns highlighted a striking predominance of macrophage-specific genes among the DEGs. DAVID-based gene ontology analysis showed a significant enrichment of "Extrinsic apoptotic signaling pathway" (FDR q = 7.60 × 10- 3) and "Response to insulin" (FDR q = 7.60 × 10- 3) genes, along with an enrichment of genes involved as "Negative regulators of DDX58/IFIH1 signaling" (FDR q = 7.50 × 10- 4) found by querying the REACTOME pathway database. Gene network analyses indicated a general dysregulation of gene inter-connections. CONCLUSION: This translational genomics study highlighted the existence both of individual genes, mostly dysregulated in certain cellular populations (e.g., macrophages), and gene-networks involved in pulmonary reaction after left pneumonectomy. Their involvement in lung homeostasis is largely supported by previous studies, carried out both in humans and in other animal models (under homeostatic or disease-related conditions), that adopted candidate-gene approaches. Overall, the present findings represent a preliminary assessment for future, more focused, studies on compensatory lung adaptation, pulmonary regeneration and functional reload.
Assuntos
Perfilação da Expressão Gênica , Pneumonectomia , Animais , Biologia Computacional , Redes Reguladoras de Genes , Humanos , Pulmão , SuínosRESUMO
Gene drives have enormous potential for the control of insect populations of medical and agricultural relevance. By preferentially biasing their own inheritance, gene drives can rapidly introduce genetic traits even if these confer a negative fitness effect on the population. We have recently developed gene drives based on CRISPR nuclease constructs that are designed to disrupt key genes essential for female fertility in the malaria mosquito. The construct copies itself and the associated genetic disruption from one homologous chromosome to another during gamete formation, a process called homing that ensures the majority of offspring inherit the drive. Such drives have the potential to cause long-lasting, sustainable population suppression, though they are also expected to impose a large selection pressure for resistance in the mosquito. One of these population suppression gene drives showed rapid invasion of a caged population over 4 generations, establishing proof of principle for this technology. In order to assess the potential for the emergence of resistance to the gene drive in this population we allowed it to run for 25 generations and monitored the frequency of the gene drive over time. Following the initial increase of the gene drive we observed a gradual decrease in its frequency that was accompanied by the spread of small, nuclease-induced mutations at the target gene that are resistant to further cleavage and restore its functionality. Such mutations showed rates of increase consistent with positive selection in the face of the gene drive. Our findings represent the first documented example of selection for resistance to a synthetic gene drive and lead to important design recommendations and considerations in order to mitigate for resistance in future gene drive applications.
Assuntos
Anopheles/genética , Genes Essenciais , Genética Populacional , Seleção Genética , Alelos , Sequência de Aminoácidos , Animais , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Feminino , Fertilidade/genética , Frequência do Gene , Biblioteca Gênica , Engenharia Genética , Haplótipos , Sequenciamento de Nucleotídeos em Larga Escala , Insetos Vetores/genética , Malária/prevenção & controle , Masculino , Modelos Genéticos , Controle de Mosquitos/métodos , Mutação , Análise de Sequência de RNARESUMO
Several recent lines of evidence are proving an important role for dopamine in the aging process and in the determination of life span. Components of the dopaminergic system may represent good candidates for longevity studies. Herein, we tested the possible association of the functional SLC6A3/DAT1 40-bp VNTR with life-expectancy in a healthy population of Central Italy (N = 993) by applying a genetic-demographic approach that takes into account the demographic information and different survival rates between sexes for modeling the survival of specific allele carriers in the population. Male carriers of S*/S* genotype showed a lower survival chance across most of the lifespan respect to the survival of DAT1*L-carriers (P = 0.021). The same analyses gave non-significant results in females. Several studies already reported significant sex differences in dopamine metabolism and its related biological pathways. Thus, we can hypothesize that the SLC6A3/DAT1 40 bp-VNTR may affect life expectancy in a sex-specific way. Moreover, it is conceivable that DAT1 S*/S* carriers, who are prone to assume "risk" type behaviors, may be dropped out of the "healthy" population by a sort of "demographic selection".
Assuntos
Pareamento de Bases/genética , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Longevidade/genética , Repetições Minissatélites/genética , Grupos Populacionais/genética , Fatores Sexuais , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Pareamento de Bases/fisiologia , Demografia , Dopamina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Dopamina/fisiologia , Feminino , Genótipo , Humanos , Itália , Longevidade/fisiologia , Masculino , Pessoa de Meia-Idade , Repetições Minissatélites/fisiologia , Modelos Genéticos , Adulto JovemRESUMO
CD38 (EC 3.2.2.6, NAD(+)-glycohydrolase) is a multifunctional enzyme catalyzing the synthesis and hydrolysis of cyclic ADP-ribose from NAD(+) to ADP-ribose. The loss of CD38 function is associated with impaired immune responses, metabolic disturbances, and behavioral modifications. Notably, it has been linked to HIV infection, leukemias, myelomas, solid tumors, Type II Diabetes mellitus, bone metabolism, as well as Autism Spectrum Disorder. Taking into account the crucial role played by CD38 in many diseases and in clinical practice, here we assessed the distribution of CD38 NADase activity in a healthy population (104 sex-matched unrelated individuals, 12-98 years) and determined its main predictors among genetic and physiological factors (age and sex). The mean value of CD38 NADase activity was 0.051±0.023 mU/mg (0.010-0.099 mU/mg), following a normal distribution in the study population (Kolmogorov-Smirnov test P=0.200). The TNF-α -308G>A (rs1800629) resulted the main predictor (ß=0.364, P=0.00008), followed by Age (ß=0.280, P=0.002) and the CD38 184C>G (rs6449182) (ß=0.193, P=0.033). Our study contributes to understanding CD38 enzyme physiological functions, by reporting, for the first time, its activity distribution in healthy individuals and demonstrating a significant positive correlation with age. Moreover, the possible use of TNF-α -308G>A (rs1800629) and the CD38 184C>G (rs6449182) SNPs as predictive genetic markers of CD38 activity, clearly point toward possible pharmacogenomic applications and to a more refined use of CD38 in clinical settings.
Assuntos
ADP-Ribosil Ciclase 1/genética , ADP-Ribosil Ciclase 1/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , ADP-Ribosil Ciclase 1/biossíntese , Adenosina Difosfato Ribose/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Feminino , Marcadores Genéticos , Variação Genética , Genótipo , Humanos , Itália , Masculino , Pessoa de Meia-Idade , NAD+ Nucleosidase/genética , NAD+ Nucleosidase/metabolismo , Polimorfismo de Nucleotídeo Único , Valores de Referência , Fator de Necrose Tumoral alfa/genética , Adulto JovemRESUMO
Aging is a complex phenotype with multiple determinants but a strong genetic component significantly impacts on survival to extreme ages. The dysregulation of immune responses occurring with increasing age is believed to contribute to human morbidity and mortality. Conversely, some genetic determinants of successful aging might reside in those polymorphisms for the immune system genes regulating immune responses. Here we examined the main effects of single loci and multi-locus interactions to test the hypothesis that the adenosine deaminase (ADA) and tumor necrosis factor alpha (TNF-α) genes may influence human life-expectancy. ADA (22G>A, rs73598374) and TNF-α (-308G>A, rs1800629; -238G>A, rs361525) functional SNPs have been determined for 1071 unrelated healthy individuals from Central Italy (18-106 years old) divided into three gender-specific age classes defined according to demographic information and accounting for the different survivals between sexes: for men (women), the first class consists of individuals<66 years old (<73 years old), the second class of individuals 66-88 years old (73-91 years old), and the third class of individuals>88 years old (>91 years old). Single-locus analysis showed that only ADA 22G>A is significantly associated with human life-expectancy in males (comparison 1 (age class 2 vs. age class 1), O.R. 1.943, P=0.036; comparison 2 (age class 3 vs. age class 2), O.R. 0.320, P=0.0056). Age- and gender-specific patterns of epistasis between ADA and TNF-α were found using Generalized Multifactor Dimensionality Reduction (GMDR). In comparison 1, a significant two-loci interaction occurs in females between ADA 22G>A and TNF-α -238G>A (Sign Test P=0.011). In comparison 2, both two-loci and three-loci interaction are significant associated with increased life-expectancy over 88 years in males. In conclusion, we report that a combination of functional SNPs within ADA and TNF-α genes can influence life-expectancy in a gender-specific manner and that males and females follow different pathways to attain longevity.
Assuntos
Adenosina Desaminase/genética , Epistasia Genética , Expectativa de Vida , Fator de Necrose Tumoral alfa/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Bases , Primers do DNA , Feminino , Humanos , Masculino , Reação em Cadeia da Polimerase , Polimorfismo de Nucleotídeo Único , Adulto JovemRESUMO
Cytidine deaminase (CDA) is a pyrimidine salvage pathway enzyme that catalyzes the hydrolytic deamination of cytidine and deoxycytidine to their corresponding uracil nucleosides. CDA also catalyzes the inactivation of some chemotherapeutic nucleoside analogues such as cytosine arabinoside and gemcitabine. CDA 79A > C (K27Q, rs2072671) and 208G > A (A70T, rs60369023) were found to be associated either with clinical outcomes as well as with pharmacokinetics and toxicity of drugs administered to different subsets of patients. In this paper we reported two PCR-based methods for CDA 79A > C (K27Q) and 208G > A (A70T) genotyping and tested their feasibility using DNA extracted from whole blood as well as from buccal swabs. The aim of this study was also to assess the distribution of genotypic variants in a central Italy population. The allele frequencies were 56.3% (K*) and 43.7% (Q*) for K27Q and 100% (A*) and 0% (T*) for A70T. The genotype frequencies were 32.8% (K*/K*), 46.9% (K*/Q*) and 20.3% (Q*/Q*) for K27Q. The genotype frequencies did not deviate from Hardy-Weinberg equilibrium. The results were compared with those of other reported populations. They showed marked ethnic group differences.
Assuntos
Alelos , Substituição de Aminoácidos/genética , Citidina Desaminase/genética , Reação em Cadeia da Polimerase/métodos , Polimorfismo de Nucleotídeo Único/genética , Estudos de Casos e Controles , Feminino , Frequência do Gene/genética , Genótipo , Humanos , Itália , Masculino , Pessoa de Meia-IdadeRESUMO
Detection of BRAFV600E within cell free tumor DNA (ctDNA) is emerging as a promising means to improve patients' stratification or enable BRAF inhibitor (BRAFi) therapeutic monitoring in a minimally invasive manner. Here, we investigated whether extracellular vesicle-(EV)-associated-DNA (EV-DNA) has value as an alternative source of circulating BRAFV600E. To do so, we identified a clinical practice-compatible protocol for the isolation of EV-DNA and assessed BRAF gene status on plasma samples from metastatic melanoma patients at the beginning and during BRAFi therapy. This protocol uses a peptide with high affinity for EVs and it has been found to recover more mutant DNA from plasma than standard ultracentrifugation. Molecular analyses revealed that mutant DNA is largely unprotected from nuclease digestion, interacting with the outer side of the EV membrane or directly with the peptide. When used on clinical samples, we found that the protocol improves the detection of BRAFV600E gene copies in comparison to the reference protocol for ctDNA isolation. Taken together, these findings indicate that EVs are a promising source of mutant DNA and should be considered for the development of next-generation liquid biopsy approaches.
Assuntos
Exossomos/genética , Melanoma/tratamento farmacológico , Nivolumabe/uso terapêutico , Proteínas Proto-Oncogênicas B-raf/genética , Idoso , Linhagem Celular Tumoral , DNA Tumoral Circulante , Feminino , Humanos , Masculino , Melanoma/sangue , Melanoma/genética , Mutação , Metástase NeoplásicaRESUMO
Recent evidence demonstrated a relevant role of adenosine deaminase (ADA) in replicative senescence of T cells through its capacity to modulate telomerase activity (TA). Herein, we tested the impact of the functional polymorphism ADA rs73598374:G>A (c.22G>A, p.Asp8Asn) on telomere biology, by measuring TA and leukocyte telomere length (LTL) in healthy subjects selected according to rs73598374 genotype. rs73598374-A carriers showed lower TA (P=0.019) and shorter LTL (P=0.003), respectively, compared to G/G carriers. rs73598374-A carriers showed a stronger cross-sectional age reduction of LTL (r=-0.314, P=0.005) compared to G/G carriers (r=-0.243, P=0.022). The reduced ADA activity associated to rs73598374-A variant predisposes those carriers to display higher levels of adenosine compared to G/G carriers. Consequently, it may lead to an accelerated process of replicative senescence, causing a stronger reduction of TA and in turn shorter LTL. In conclusion, the crucial role played by replicative senescence of the immune system in several human diseases and in the aging process underscores the relevance of the present findings and also spurs interest into the possible involvement of rs73598374 in shaping the susceptibility to several age-related diseases.
Assuntos
Adenosina Desaminase/genética , Polimorfismo de Nucleotídeo Único , Telomerase/metabolismo , Homeostase do Telômero/genética , Adenosina Desaminase/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Leucócitos/metabolismo , MasculinoRESUMO
The dysregulation of both immune and inflammatory responses occurring with aging is believed to substantially contribute to morbidity and mortality in humans. We have already reported the association of the functional Variable Number of Tandem Repeat (VNTR) at the Immunoglobulin heavy chain (IGH) enhancer HS1.2 with Immunoglobulin levels and with several autoimmune diseases. Herein we tested the association of the VNTR at the HS1.2 enhancer with human longevity, also evaluating the possible modulatory effect of TNFA promoter diplotype (rs361525/rs1800629). HS1.2 enhancer genotypes have been determined for 193 unrelated healthy individuals from Central Italy divided into two groups: Group 1 (18-84 yrs, mean age 56.8 ± 19.4) and Group 2 (85-100 yrs, mean age 93.0 ± 3.5). Homozygous subjects for 2 allele were significantly disadvantaged in reaching higher life-expectancy (OR=0.457, p=0.021). A significant interaction between TNFA promoter diplotype status, HS1.2 2/2 genotype and the two Groups was found (p=0.014). Of note, TNFA -308A allele seems to exert a protective effect in HS1.2 2/2 carriers. These results support the hypothesis of an important role of HS1.2 VNTR in the puzzle of the immune-system regulation, evidenced also by the potential interaction with TNFA. Moreover, the previous results showing the association of HS1.2 2 allele with inflammatory phenomena are consistent with the hypothesis that this allele is a detrimental factor in reaching advanced age.
Assuntos
Cadeias Pesadas de Imunoglobulinas/genética , Repetições Minissatélites/genética , Regiões Promotoras Genéticas/genética , Fator de Necrose Tumoral alfa/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Bases , Elementos Facilitadores Genéticos/genética , Epistasia Genética , Feminino , Frequência do Gene , Genética Populacional , Genótipo , Humanos , Itália , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Polimorfismo de Nucleotídeo Único , Homologia de Sequência do Ácido Nucleico , Adulto JovemRESUMO
Cytidine deaminase (CDA), is one of the enzymes involved in the pyrimidine salvage pathways, which catalyzes the formation of uridine and deoxyuridine by the hydrolytic deamination of cytidine and deoxycytidine, respectively. Human CDA is a tetrameric enzyme of identical 15 kDa subunits, each containing an essential zinc atom in the active site. The substrate binds to each active site independently and the cooperativity between subunits has not been reported. CDA is able to recognize as substrates some antitumor and antiviral cytidine analogs rendering them pharmacologically inactive. In light of the role played by this enzyme, a deep knowledge of CDA active site and mechanism of catalysis is required. Site-directed mutagenesis, associated with molecular modeling studies, may be an important tool to discover the active site structure of an enzyme and consequently its mechanism of action. In this review are summarized the site-directed mutagenesis experiments performed on human CDA: through these studies it was possible to understand the role exerted by specific amino acid residues in CDA active site and in the contacts between subunits. The obtained results may open a way for designing new cytidine based drugs or more potent CDA inhibitors.
Assuntos
Citidina Desaminase/química , Citidina Desaminase/metabolismo , Sequência de Aminoácidos , Domínio Catalítico , Citidina Desaminase/genética , Humanos , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Subunidades Proteicas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alinhamento de SequênciaRESUMO
AIM: To assess the distribution of CDA activity from whole blood of 142 healthy subjects, determining its main predictors among genetic (six CDA SNPs) and physiological factors (age and gender). Moreover, we performed a kinetic study of the two CDA protein variants (Q27 and K27) determined by the rs2072671 SNP. MATERIALS & METHODS: CDA activity was assessed by HPLC. Selected CDA SNPs were genotyped by PCR-based methods. Recombinant CDA protein variants (Q27 and K27) were expressed in an Escherichia coli strain SØ5201 and kinetic assays were performed. RESULTS: The mean value of CDA activity was 0.051 ± 0.024 mU/mg and followed a normal distribution in the study population. Carriers of the CDA*2B (-451T/-92G/-31Del/79C/435C) haplotype displayed higher CDA activity compared with the others. CDA -451G>A, -92A>G and 79A>C (K27Q) SNPs displayed significant associations with CDA activity. The best predictive model of CDA activity included the variables gender and CDA 79A>C (K27Q). Cytidine is the preferential substrate for the variant Q27. CONCLUSION: We suggest the analysis of both CDA activity and CDA 79A>C (K27Q) SNP in future prospective trials with cytidine analogs, alone or in combination, in order to identify the best marker to secure the administration of these anticancer therapies.
Assuntos
Citidina Desaminase/genética , Citidina Desaminase/metabolismo , Estudos de Associação Genética , Haplótipos/genética , População Branca/genética , Feminino , Humanos , Itália , Masculino , Pessoa de Meia-Idade , Fenótipo , Polimorfismo de Nucleotídeo ÚnicoRESUMO
The association of TP53 P72R (rs1042522) with longevity remains uncertain and varies with ethnicity. Here, we tested its association with longevity in a cross-sectional population of Central Italy (18-106 years, N = 1,072), by integrating demographic information and frequency data to account for the different survival rates between sexes through the application of a genetic-demographic approach. rs1042522 affects females longevity, showing significant associations in Comparison 2 (Age Class 3 [>91 years] vs Age Class 2 [73-91 years]) under both additive (odds ratio [OR] 0.574; p = .006) and dominant (OR 0.513; p = .006) models. The TP53*P72 allele is significantly underrepresented in Age Class 3 only in women (OR 0.575; p = .008). The genetic-demographic approach demonstrated that the frequency of female TP53*P72 carriers underwent a significant reduction after 82 years (OR 0.586; p = .002). The same analyses gave nonsignificant results in men. The discrepancies among the results obtained on rs1042522 for longevity could result from the pleiotropic effects of p53 and the potential ethnic variation of its functional variants.
Assuntos
Genes p53/genética , Expectativa de Vida , Longevidade/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Estudos Transversais , Demografia , Feminino , Humanos , Itália , Pessoa de Meia-Idade , Fatores Sexuais , Adulto JovemRESUMO
BACKGROUND: Telomerase, encoded by TERT, is the ribonucleoprotein polymerase that maintains telomere ends and it plays a crucial role in cellular senescence. TERT single nucleotide polymorphisms (SNPs) have been associated both with various malignancies and telomere length (TL). The association of TERT SNPs with longevity remains uncertain and varies with ethnicity. The aim of this study was to investigate whether the functional variable number of tandem repeat (VNTR) MNS16A of TERT is associated with longevity. METHODS: MNS16A genotypes have been determined for 1072 unrelated healthy individuals from Central Italy (18-106 years old) divided into three gender-specific age classes defined according to demographic information and accounting for the different survivals between sexes: for men (women), the first class consists of individuals <66 years old (<73 years old), the second class of individuals 66-88 years old (73-91 years old), and the third class of individuals >88 years old (>91 years old). TL was assessed using genomic DNA from whole blood of 72 selected individuals by a multiplex real-time PCR assay. RESULTS: MNS16A appears associated to longevity, showing significant associations in Comparison 2 (Age Class 3 vs. Age Class 2) under both additive (odds ratio [O.R.] 0.749; p=0.019) and dominant (O.R. 0.579; p=0.011) models. The MNS16A*L allele is significantly underrepresented in Age Class 3 (O.R. 0.759; p=0.020) compared to Age Class 2. A significant telomere attrition is reported along the three age classes (p=0.0001), that remains significant only in L*/L* genotype carriers (p=0.002) when the analysis was conducted according to MNS16A genotype. CONCLUSIONS: The TERT MNS16A*L allele appears negatively associated with longevity. The concomitant significant telomere cross sectional attrition rate observed for L*/L* genotype suggests that this polymorphism could influence human longevity by affecting TL.
Assuntos
Predisposição Genética para Doença , Longevidade/genética , Repetições Minissatélites/genética , Polimorfismo de Nucleotídeo Único , Telomerase/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Itália , Masculino , Pessoa de Meia-Idade , Fatores de Risco , População Branca/genética , Adulto JovemRESUMO
Dopamine is a neurotransmitter whose functions are mediated by five receptors expressed in several organs and tissues. Dopaminergic system dysfunctions are involved in the etiology or treatment of several pathological conditions, including drug addiction. Alcohol dependence (AD) is a widespread psychiatric disorder, affecting 5.4% of the general population lifetime. Family and twins studies support the role of a genetic component in AD. Since dopamine neurotransmission has been shown to be involved in drug reward, related genes are plausible candidates for susceptibility to AD. Here, we evaluated both the DRD2/ANKK1 TaqIA (rs1800497) and SLC6A3 40 bp-VNTR SNP and gene-gene interaction analysis in AD patients from a population of Central Italy. The study design was a case-control. In total, 280 alcoholic subjects (213 men and 67 woman) and 280 age- and sex-matched control subjects were recruited for this study. Case subjects met the DSM-IV criteria for AD and they are free from any psychiatric co-morbidities. Controls were subjects who had non-alcohol problem either never drank; those who have smoked at least one pack of cigarettes per day for at least 1 year were excluded. Genotyping was performed by allele-specific PCR and RFLP-PCR. SLC6A3 40 bp 3'UTR-VNTR displays no association with AD. DRD2/ANKK1 TaqIA genotype distribution is significantly associated to AD (O.R.=1.551, p=0.023), with A1* allele displaying an O.R.=1.403 (p=0.029). Gene-gene interaction analysis using three-way contingency table analysis by a log-linear model yielded no significant result. Our study in a population of Central Italy extends and confirms previous results and, for the first time, tested the gene-gene interaction between SLC6A3 and DRD2 in AD.
Assuntos
Transtornos Relacionados ao Uso de Álcool/genética , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Proteínas Serina-Treonina Quinases/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Alcoolismo/genética , Estudos de Casos e Controles , Epistasia Genética , Feminino , Estudos de Associação Genética , Humanos , Itália , Masculino , Pessoa de Meia-Idade , Repetições Minissatélites , Polimorfismo de Nucleotídeo Único , Adulto JovemRESUMO
Polyamines (putrescine, spermidine, and spermine) are a family of molecules that derive from ornithine through a decarboxylation process. They are essential for cell growth and proliferation, stabilization of negative charges of DNA, RNA transcription, translation, and apoptosis. Recently, it has been demonstrated that exogenously administered spermidine promotes longevity in yeasts, flies, worms, and human cultured immune cells. Here, using a cross-sectional observational study, we determined whole-blood polyamines levels from 78 sex-matched unrelated individuals divided into three age groups: Group 1 (31-56 years, n=26, mean age 44.6±6.07), group 2 (60-80 years, n=26, mean age 68.7±6.07), and group 3 (90-106 years, n=26, mean age 96.5±4.59). The total content of polyamines is significantly lower in groups 2 and 3 compared to group 1 (p=3.6×10(-12)). Interestingly, this reduction is mainly attributable to the lower putrescine content. Group 2 displays the lowest levels of spermidine and spermine. On the other hand, nona/centenarians (group 3) display a significantly higher median relative percentage content of spermine with respect to total polyamines, compared to the other groups (13.2% vs. 14.1% vs. 30.6%, p=6.0×10(-4)). For the first time, we report profiles of polyamines from the whole blood of healthy nona/centenarians, and our results confirm and extend previous findings on the role of polyamines in determining human longevity. However, although we found an important correlation between polyamines levels and age groups, further studies are warranted to fully understand the role of polyamines in determining life span. Also, longitudinal and nutritional studies might suggest potential therapeutic approaches to sustain healthy aging and to increase human life span.
Assuntos
Espermidina/sangue , Espermina/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Redes e Vias Metabólicas , Metionina/metabolismo , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Haptoglobin (HP), which scavenges free, cell-toxic hemoglobin and has anti-inflammatory and immune-modulatory function in extravascular tissues, may represent an excellent candidate gene to investigate the life-span expectancy. METHODS: HP 1/2 polymorphism has been determined for 1072 (569 females, 503 males) unrelated healthy individuals from Central Italy, 18-106 years old, divided into three gender-specific age classes defined according to demographic information and accounting for the different survivals between sexes. HP*1F/S subtyping was also performed to check the possible existence for a preferential advantage of HP*1F or HP*1S allele. RESULTS: HP*1/*1 genotype results associated to increased probability of young subjects of attaining longevity (Comparison 1: O.R. 1.709, p=0.0114) with a concomitant advantage of HP*1 allele (Comparison 1: O.R. 1.273, p=0.0194). On the other side, carriers of HP*2 allele displayed an overall significant disadvantage in reaching Age Class 2 (O.R. 0.585, p=0.0092). No significant differences were noticed between age groups either considering total HP*1F and HP*1S allele frequencies or according to HP 1/2 genotypes. CONCLUSION: The crucial role played by HP in aging process is warranted by its many established functions and its related phenotypes so that it may be considered an important gene involved in the determination of human survival.
Assuntos
Haptoglobinas/genética , Longevidade/genética , Polimorfismo Genético , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Sequência de Bases , Estudos Transversais , Primers do DNA , Feminino , Humanos , Itália , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
BACKGROUND: Apolipoprotein E (APOE) functional haplotypes determined by rs429358 and rs7412 SNPs have been extensively studied and found to be one of the most consistent association in human longevity studies. However, the search for longevity-determining genes in human has largely neglected the operation of genetic interactions. METHODS: APOE haplotypes have been determined for 1072 unrelated healthy individuals from Central Italy, 18-106 years old, divided into three gender-specific age classes defined according to demographic information and accounting for the different survival between sexes. The epistasis between APOE haplotypes and Haptoglobin (HP) 1/2 polymorphism was tested according to three-way contingency table analysis by a log-linear model. RESULTS: APOE genotype and haplotype distributions differ significantly along the age classes (Genotype: p=0.014; Haplotype: p=0.005) with APOE*ε4 genotype status and haplotype displaying negative association (Genotype: O.R.=0.377, p=0.002, Haplotype: O.R.=0.447, p=0.005). A significant interaction between APOE*ε4 genotype status, HP 1/2 genotype and age classes is reported (p=0.006). CONCLUSION: APOE haplotypes are significantly associated with longevity in our population. Of note, HP*1/*1 genotype seems to protects APOE*ε4 carriers from age-related negative selection. Collectively, these results also suggest and claim for further investigations on APOE/HP interaction in other age-related diseases such as Alzheimer's disease, atherosclerosis and Parkinson's disease.
Assuntos
Apolipoproteínas E/genética , Epistasia Genética , Haplótipos , Longevidade/genética , Polimorfismo Genético , Sequência de Bases , Primers do DNA , Humanos , ItáliaRESUMO
Since the pioneer experiments conducted by Friedrich Miescher in 1861, extraordinary advances have been achieved in the field of DNA handling. Today nucleic acids can be extracted from any type of biological material such as tissues, cells and viruses. Moreover, increasing knowledge of human genome is paving the way to an effective employment of pharmacogenomics and genetic-based predictive tests in medicine. In this context, the recovery of DNA from different sources of biological samples (e.g. archived formalin-fixed autopsy tissues, dried blood spots, frozen serum or plasma, long-term stored whole blood) is also an emerging field in genetic epidemiology studies. Thus, given the crucial role played by DNA in bio-medical research and in its related applications, here we review the main relevant issued patents and recently published advances in the field of DNA extraction and purification from human specimens.
Assuntos
DNA/isolamento & purificação , Patentes como Assunto , Fixadores/química , Formaldeído/química , Humanos , Manejo de Espécimes/métodosRESUMO
Cytidine deaminase (ECâ 3.5.4.5, CDA), an enzyme of the pyrimidine salvage pathways, is responsible for the degradation and inactivation of several cytidine-based antitumor drugs such as cytarabine, gemcitabine, decitabine, and azacytidine. Thus, CDA inhibitors are highly sought after as compounds to be co-administered with said drugs to improve their effectiveness. Alternatively, the design of antitumor drugs not susceptible to the action of CDA is also regarded as an attractive solution. Herein we describe a virtual screen for CDA ligands based on chemical similarity and molecular docking. The campaign led to the identification of three novel inhibitors and one novel substrate, with a 19 % hit rate, and allowed a significant extension of the structure-activity relationships, also in light of the compounds that resulted inactive. The most active compound identified through the screen is the inhibitor pseudoisocytidine, which has the potential to serve as a lead for highly stable compounds. The study also delineated the detrimental effect of 5-aza and 6-aza substitutions, the incompatibility of the presence of an amino group at the 3'-position, as well as the presence of very strict steric requirements around the 2'-arabino position and, even more, the N4-position. Importantly, these features can be exploited for the design of novel anti-neoplastic agents resistant to the action of CDA.