Vascular Endothelial Growth Factor as Molecular Target for Bronchopulmonary Dysplasia Prevention in Very Low Birth Weight Infants.

Bronchopulmonary dysplasia (BPD) still represents an important burden of neonatal care. The definition of the disease is currently undergoing several revisions, and, to date, BPD is actually defined by its treatment rather than diagnostic or clinic criteria. BPD is associated with many prenatal and postnatal risk factors, such as maternal smoking, chorioamnionitis, intrauterine growth restriction (IUGR), patent ductus arteriosus (PDA), parenteral nutrition, sepsis, and mechanical ventilation. Various experimental models have shown how these factors cause distorted alveolar and vascular growth, as well as alterations in the composition and differentiation of the mesenchymal cells of a newborn's lungs, demonstrating a multifactorial pathogenesis of the disease. In addition, inflammation and oxidative stress are the common denominators of the mechanisms that contribute to BPD development. Vascular endothelial growth factor-A (VEGFA) constitutes the most prominent and best studied candidate for vascular development. Animal models have confirmed the important regulatory roles of epithelial-expressed VEGF in lung development and function. This educational review aims to discuss the inflammatory pathways in BPD onset for preterm newborns, focusing on the role of VEGFA and providing a summary of current and emerging evidence.

Hypoxic ischemic brain injury: animal models reveal new mechanisms of melatonin-mediated neuroprotection.

Oxidative stress (OS) and inflammation play a key role in the development of hypoxic-ischemic (H-I) induced brain damage. Following H-I, rapid neuronal death occurs during the acute phase of inflammation, and activation of the oxidant-antioxidant system contributes to the brain damage by activated microglia. So far, in an animal model of perinatal H-I, it was showed that neuroprostanes are present in all brain damaged areas, including the cerebral cortex, hippocampus and striatum. Based on the interplay between inflammation and OS, it was demonstrated in the same model that inflammation reduced brain sirtuin-1 expression and affected the expression of specific miRNAs. Moreover, through proteomic approach, an increased expression of genes and proteins in cerebral cortex synaptosomes has been revealed after induction of neonatal H-I. Administration of melatonin in the experimental treatment of brain damage and neurodegenerative diseases has produced promising therapeutic results. Melatonin protects against OS, contributes to reduce the generation of pro-inflammatory factors and promotes tissue regeneration and repair. Starting from the above cited aspects, this educational review aims to discuss the inflammatory and OS main pathways in H-I brain injury, focusing on the role of melatonin as neuroprotectant and providing current and emerging evidence.

Molecular barcoding of an atypical cyprinid population assessed by cytochrome B gene sequencing.

A fish population of the carp family Cyprinidae with atypical phenotypic characteristics was observed in one of the main catchments of the Pollino National Park, a valuable, protected area in southern Italy. In this area, the Italian roach Rutilus rubilio (Bonaparte, 1837), a native endemic fish of Tyrrhenean regions, has been introduced in sympatric conditions with Squalius squalus (Bonaparte, 1837) and Telestes muticellus (Bonaparte, 1837). A molecular investigation was carried out to assess the genetic identity of the population with a view to conservation. Direct sequencing of a cytochrome b gene fragment was performed based on 30 individuals of cyprinid fish with atypical phenotype, in addition to 30 S. squalus, 10 T. muticellus, and 30 R. rubilio pure individuals collected in different Italian regions, which served as reference samples. Multiple sequence alignments demonstrated that 50% of atypical-cyprinid haplotypes were maternally inherited from either S. squalus or R. rubilio. No contribution by T. muticellus was determined. Our results indicate an intergeneric hybridization event between S. squalus and R. rubilio, as a consequence of trans-introduction activities of alien species.

Molecular Barcoding: A Tool to Guarantee Correct Seafood Labelling and Quality and Preserve the Conservation of Endangered Species.

The recent increase in international fish trade leads to the need for improving the traceability of fishery products. In relation to this, consistent monitoring of the production chain focusing on technological developments, handling, processing and distribution via global networks is necessary. Molecular barcoding has therefore been suggested as the gold standard in seafood species traceability and labelling. This review describes the DNA barcoding methodology for preventing food fraud and adulteration in fish. In particular, attention has been focused on the application of molecular techniques to determine the identity and authenticity of fish products, to discriminate the presence of different species in processed seafood and to characterize raw materials undergoing food industry processes. In this regard, we herein present a large number of studies performed in different countries, showing the most reliable DNA barcodes for species identification based on both mitochondrial (COI, cytb, 16S rDNA and 12S rDNA) and nuclear genes. Results are discussed considering the advantages and disadvantages of the different techniques in relation to different scientific issues. Special regard has been dedicated to a dual approach referring to both the consumer's health and the conservation of threatened species, with a special focus on the feasibility of the different genetic and genomic approaches in relation to both scientific objectives and permissible costs to obtain reliable traceability.

Molecular Polymorphisms of Vascular Endothelial Growth Factor Gene and Bronchopulmonary Dysplasia in Very Low Birth Weight Infants.

Background: Bronchopulmonary dysplasia (BPD) is a chronic lung disease affecting primarily preterm and very low birth weight (VLBW) infants. Despite the advances in perinatal care, BPD remains a major clinical and costly complication in premature infants. The pathogenesis of BPD is complex and multifactorial. Prematurity, mechanical ventilation, oxidative stress, and inflammation are recognized as major interrelated contributing factors. Recently, some candidate genes involved in angiogenesis and alveolarization regulating mechanisms have been associated to BPD risk development. The aim of this study was to evaluate the role of vascular endothelial growth factor (VEGF) polymorphisms on BPD onset in VLBW newborns. Methods: Eighty-two VLBW infants, without major anomalies, were consecutively enrolled: 33 developed BPD (BPD group) and 49 infants without BPD served as controls (control group). In all infants, two polymorphisms, respectively (VEGF receptor) VEGFR1-710 C/T and VEGF +936 C/T, were determined through salivary brush. Genomic DNA was extracted and purified from saliva samples by using the MasterAmp Buccal Swab DNA Extraction Kit (Tebu-bio, Milan, Italy). Results: Significant statistic differences were found between BPD newborns and controls with regard to gestational age, birth weight, mechanical ventilation, duration of oxygen therapy, maternal preeclampsia, and chorioamnionitis. No differences were detected between genotypic and allelic levels regarding VEGFR1 and VEGF molecular polymorphisms. Conclusions: Two single nucleotide polymorphisms within VEGF and VEGFR1 genes are not associated with BPD. Further researches are needed to reveal gene polymorphisms involved in vascular development as contributors to the onset of BPD.

Efficiency of DNA Mini-Barcoding to Assess Mislabeling in Commercial Fish Products in Italy: An Overview of the Last Decade.

The problem of fish traceability in processed products is still an important issue in food safety. Major attention is nowadays dedicated to consumer health and prevention of possible frauds regulated by national and international laws. For this reason, a technical approach is fundamental in revealing mislabeling at different levels. In particular, the use of genetic markers has been standardized and DNA barcoding is considered the gold-standard strategy to examine and prevent species substitution. Considering the richness of available DNA databases, it is nowadays possible to rapidly reach a reliable taxonomy at the species level. Among different approaches, an innovative method based on DNA mini barcoding has recently been proposed at an international level. Starting from this evidence, we herein illustrate an investigation dealing with the evolution of this topic in Italy over the last decade. The molecular analysis of 71 commercial fish samples based on mini-COI sequencing with two different primer sets reached an amplification success rate of 87.3 and 97.2%. The investigation revealed four major frauds (5.8%) and four minor ones (5.8%). Results highlighted a decrease in incorrect labeling in Italy from 32% to 11.6% over the last decade, although a recurrent involvement of "endangered" species sensu IUCN was still observed.

Evidence that polymorphic deletion of the glutathione S-transferase gene, GSTM1, is associated with esophageal atresia.

BACKGROUND: Esophageal atresia (EA) is a life-threatening congenital condition whose etiology and pathogenesis are still poorly understood. An increasing trend of this pathology in some Italian regions suggests a possible interaction between xenobiotics and genes involved in detoxification processes during early embryonic development. For the first time polymorphisms of GSTM1, GSTT1, and GSTP1 genes were analyzed in association with EA. METHODS: The study population consisted of 25 EA children, 50 unrelated healthy children, 20 of the EA children's mothers, and 40 unrelated mothers. GSTM1 and GSTT1 null genotypes were identified by PCR amplification, and GSTP1 polymorphism was detected by RFLP analysis. RESULTS: An association was found between homozygosity for the GSTM1 null genotype and EA in affected children (p = 0.0022) and their mothers (p = 0.022). No association was found between GSTT1 and GSTP1 polymorphisms and EA children or their mothers. CONCLUSIONS: Results suggest that the GSTM1(-/-) null genotype may play an important role in the development of EA during early embryogenesis as a consequence of altered detoxification processes both in children and in the mothers. We hypothesize that GSTM1 allelic loss could be responsible for reduced or null catalytic activity in tissues exposed to amniotic fluid, and inefficient detoxification could be a trigger altering proliferation/apoptotic pattern of gut-trachea separation.

Association of dopamine transporter and monoamine oxidase molecular polymorphisms with sudden infant death syndrome and stillbirth: new insights into the serotonin hypothesis.

Recent findings demonstrated the role of neurotransmitters in the aetiopathogenesis of sudden unexpected deaths in infancy. Although genes involved in serotonin metabolism have been proposed as risk factors for sudden infant death syndrome (SIDS), the contribution of additional neurotransmitters and genes different from the serotonin transporter (SLC6A4, 5-HTT) has not been investigated. Considering the common metabolic pathway and synergism between dopamine and serotonin, the role of dopamine transporter (SLC6A3, DAT) and monoamine oxidase A (MAOA) genes in SIDS and stillbirth (sudden intrauterine unexplained death, SIUD) was investigated. Genotypes and allelic frequencies of DAT and MAOA were determined in 20 SIDS and five stillbirth cases and compared with 150 controls. No association was found between DAT polymorphisms and SIDS either at genotype (P = 0.64) or allelic (P = 0.86) level; however, a highly significant association was found between MAOA genotypes (P = 0.047) and alleles (P = 0.002) regulating different expression patterns (3R/3R vs 3.5R/3.5R + 4R/4R) in SIDS + SIUD and controls. Analysis of combined 5-HTTLPR (serotonin transporter linked polymorphic region)/MAOA genotypes revealed that frequency of L/L-4R/4R genotype combination was eightfold higher in SIDS + SIUD than in controls (P < 0.001). Findings are discussed considering the metabolic association among DAT, 5-HTT and MAOA with special emphasis on the linked action of 5-HTT/MAOA in regulating serotonin metabolism of SIDS and SIUD infants.

Genes regulating the serotonin metabolic pathway in the brain stem and their role in the etiopathogenesis of the sudden infant death syndrome.

Genotypes and allelic frequencies of TPH2, 5-HTTLPR, the 5-HTT (SLC6A4) intron 2 variable-number tandem repeat (VNTR) region, and the MAOA VNTR region were determined in brain-stem samples of 20 "genuine" SIDS cases and compared with results obtained from 150 healthy controls. The SNP G1463A responsible for 80% functionality loss of TPH2 (tryptophan hydroxylase 2) was not detected, neither in SIDS infants nor in the controls. In contrast, a strict relation was found between the 5-HTTLPR genotype and its allelic frequencies with SIDS cases. The L/L genotype and the long allele (L) of the promoter region of the serotonin transporter were significantly associated (likelihood ratio (LR) test, p<0.001) with the syndrome (L/L, 60% SIDS vs 14% controls; L, 80% SIDS vs 42.6% controls). Polymorphisms of the intron 2 VNTR of the same gene showed a trend for significant differences between genotypes 10/10 and 12/12 (LR test, p=0.068), with the L-12 haplotype being almost twofold in SIDS (44.5%) with respect to controls (23.4%). Differences were even higher considering the genotype combination L/L-12/12 (20% SIDS vs 2.6%), and variations among categories were statistically highly significant (p<0.001). Although additional differences were observed in the frequency of the MAOA (monoamine oxidase A) VNTR genotype 3R/3R between SIDS and controls (respectively 15% vs 26%), the results were not supported by statistical significance. Molecular polymorphisms are discussed considering their functional role in regulating serotonin synthesis (TPH2), neuronal reuptake (5-HTTLPR and 5-HTT intron 2), and catabolism (MAOA) in the nervous system of Italian SIDS infants. Comparisons are made with previous data obtained in different ethnic groups.

Detoxification genes polymorphisms in SIDS exposed to tobacco smoke.

The best hypothesis to explain Sudden Infant Death Syndrome (SIDS) pathogenesis is offered by the "triple risk model", which suggests that an interaction of different variables related to exogenous stressors and infant vulnerability may lead to the syndrome. Environmental factors are triggers that act during a particular sensible period, modulated by intrinsic genetic characteristics. Although literature data show that one of the major SIDS risk factors is smoking exposure, a specific involvement of molecular components has never been highlighted. Starting from these observations and considering the role of GSTT1 and GSTM1 genes functional polymorphisms in the detoxification process, we analyzed GSTM1 and GSTT1 null genotype frequencies in 47 SIDS exposed to tobacco smoke and 75 healthy individuals. A significant association (p < .0001) between the GSTM1 null genotype and SIDS exposed to smoke was found. On the contrary, no association between GSTT1 polymorphism and SIDS was determined. Results indicated the contribution of the GSTM1 -/- genotype resulting in null detoxification activity in SIDS cases, and led to a better comprehension of the triple risk model, highlighting smoking exposure as a real SIDS risk factor on a biochemical basis.

Serotonin transporter role in identifying similarities between SIDS and idiopathic ALTE.

OBJECTIVE: Considering previous genetic studies on sudden infant death syndrome (SIDS) and the role of L/L serotonin transporter (5HTT) genotype and correlated genes monoamine oxidase A (MAOA) and dopamine transporter (DAT) in unexpected death, an investigation was carried out verifying their involvement in apparent life-threatening events (ALTE and idiopathic form [IALTE]), also assessing common molecular basis with SIDS. METHODS: Differential diagnoses in 76 ALTE infants, distinguishing ALTE from IALTE was elaborated by using clinical-diagnostic data. Genotypes/allelic frequencies of DAT, MAOA, and 5HTT were determined in ALTE and IALTE infants and compared with data obtained from 20 SIDS and 150 controls. RESULTS: No association was found between DAT polymorphisms and ALTE/IALTE groups either at the genotype or allelic level (P range .11-.94). MAOA genotypes and allele data comparison between ALTE and controls was not significant; IALTE data showed a tendency for genotypes (P = .09) and were statistically significant for alleles (P = .036); however, MAOA significance disappeared once the Bonferroni correction was applied. 5HTT polymorphisms in IALTE remarked the role of L/L genotype (P < .00001) and L (P < .00001), as previously demonstrated in SIDS. CONCLUSIONS: Considering correspondence between 5HTT and MAOA in IALTE and SIDS, we hypothesize that the 2 syndromes are different expressions of a common ethiopathogenesis. In particular, genetic data suggest SIDS events could derive from IALTE episodes occurred during sleep, and therefore out of parental control. Despite its functional role, results highlight the usefulness of 5HTT as a valuable tracer of SIDS risk in IALTE infants. Owing to the small sample size, the results are to be considered preliminary and should be reevaluated in an independent sample.