The Presence of Mycotoxins in Human Amniotic Fluid

Mycotoxin exposure assessments through biomonitoring studies, based on the analysis of amniotic fluid, provides useful information about potential exposure of mothers and fetuses to ubiquitous toxic metabolites that are routinely found in food and the environment. In this study, amniotic fluid samples (n = 86) were collected via abdominal amniocentesis at 15–22 weeks of gestation from pregnant women with a high risk of chromosomal anomalies or genetic fetal defects detected during 1st trimester prenatal screening. These samples were analyzed for the presence of the most typical Aspergillus, Penicillium and Fusarium mycotoxins, with a focus on aflatoxins, ochratoxins and trichothecenes, using the LC-FLD/DAD method. The results showed that the toxin was present in over 75% of all the tested samples and in 73% of amniotic fluid samples from fetuses with genetic defects. The most frequently identified toxins were nivalenol (33.7%) ranging from <LOQ to 4037.6 ng/mL, and aflatoxins (31.4%), including aflatoxin G1, ranging from <LOQ to 0.4 ng/mL. Ochratoxin A and deoxynivalenol were identified in 26.7% and 27.9% of samples, respectively. Bearing in mind the above, the detection of mycotoxin levels in amniotic fluid is useful for the estimation of overall risk characterization with an attempt to link the occurrence of fetal abnormalities with exposure to mycotoxins in utero.     

Vitamin D supplementation increases adipokine concentrations in overweight or obese adults

European Journal of Nutrition - Vitamin D regulates adipokine production in vitro; however, clinical trials have been inconclusive. We conducted secondary analyses of a randomized controlled trial...     

Spanish BH4-responsive phenylalanine hydroxylase-deficient patients: evolution of seven patients on long-term treatment with tetrahydrobiopterin

A novel subtype of patients with mutations in the phenylalanine hydroxylase (PAH) gene that show a positive response during a tetrahydrobiopterin (BH4) loading test has recently been recognized. These studies suggest that a number of phenylketonuric (PKU) patients may benefit from BH4 substitution, eliminating the need of life-long dietary restrictions. In our unit, we performed BH4 overload tests in 50 PAH-deficient patients. Overall, 38% of the patients had a positive response, mostly MHP and mild PKU patients, all with at least one missense mutation with presumed residual activity. Seven of the patients that required dietary restrictions have received treatment with BH4 from 5 to 18 months. All the patients included in the long-term treatment protocol had a mild PKU phenotype. BH4 therapy began at 10 mg/kg/day and changes were made over time depending on Phe levels. All patients at least doubled their protein ingestion and some could follow a completely free diet. Patients with a smaller decrease in Phe levels during the BH4 overload required higher BH4 doses and/or dietary restrictions to maintain adequate Phe levels over time. The genotype and the potential mechanisms underlying BH4 responsiveness and interindividual differences in pharmacokinetics of the administered cofactor are probably the basis for the differences in prolonged treatment.     

Effect of exogenous opioid peptides on TNF-alpha-induced human neutrophil apoptosis in vitro

Polymorphonuclear leukocyte (neutrophil) apoptosis is an important mechanism regulating the life span and some functions of neutrophils at inflamed sites. Opioid peptides are present in the peripheral circulation and their concentrations rapidly increase as a result of stress and inflammation. The effect of opioid peptides such as met-enkephalin (M-ENK) and beta-endorphin (beta-END) on tumor necrosis factor (TNF)-alpha-induced apoptosis in human neutrophils in vitro was investigated. Neutrophils isolated from peripheral blood were cultured in the absence or presence of 10(-6)-10(-10) M of opioid peptides for 8, 12 and 18 h. Features of apoptotic neutrophils were measured by a flow cytometric method based on analysis of the apoptotic nuclei (DNA content). We found that M-ENK and beta-END enhanced both uninduced and TNF-alpha-induced neutrophil apoptosis in vitro in a dose-dependent manner. The effect of opioid peptides on the modulation of neutrophil apoptosis was not reversed by the opioid-receptor antagonist naloxone. The results suggest that M-ENK and beta-END can regulate neutrophil life span via apoptosis and in this way may participate in the resolution of inflammation.     

Kinetic and stability analysis of PKU mutations identified in BH4-responsive patients

From all the different molecular mechanisms put forward to explain the basis of BH4 responsiveness in PKU patients, a clear picture is now emerging based on the results from expression studies performed with a number of missense mutations identified in patients with a positive response in BH4 loading tests. Two of the proposed mechanisms, namely decreased binding affinity of the mutant proteins for the natural cofactor and stabilization effect of BH4, have been confirmed for several PKU mutations and the results are reviewed here. The actual view supports a multifactorial basis of the response, highlighting the necessity of detailed in vitro characterization of each mutant PAH protein. Several of the confirmed molecular mechanisms may be operating simultaneously, as exemplified in the data presented, and this may result in different degrees of BH4 responsiveness.     

Induction of peptide-specific primary cytotoxic T lymphocyte responses from human peripheral blood

Various protocols were developed and compared for eliciting specific cytotoxic T lymphocyte (CTL) cell lines from the unselected human peripheral blood mononuclear cells of naive donors. Interleukin-7 and CD4⁺ T cells primed in vitro by keyhole limpet hemocyanin were shown to act together in the generation of these responses. Primary responses were consistently induced with a variety of different HLA class I-binding malarial peptides. Primary CTL responses could be induced from unselected CD8⁺ and from CD45RA⁺CD8⁺ T cells. The CTL lines derived from these naive donors were CD8⁺ and demonstrated a high level of HLA class I-restricted killing for > 3 months after priming in vitro. They were also able to recognize and kill targets infected with a recombinant vaccinia virus containing the full-length antigen. In addition, this same protocol enhanced up to fourfold the levels of secondary CTL responses induced. The optimal method presented for naive cytotoxic T cell stimulation is simple, rapid and generally applicable and should provide a useful tool for both basic research and human therapy.