Mapping the neuroanatomical abnormalities in a phenotype of male compulsive rats.

Compulsivity is considered a transdiagnostic dimension in obsessive-compulsive and related disorders, characterized by heterogeneous cognitive and behavioral phenotypes associated with abnormalities in cortico-striatal-thalamic-cortical circuitry. The present study investigated the structural morphology of white and gray matter in rats selected for low- (LD) and high- (HD) compulsive drinking behavior on a schedule-induced polydipsia (SIP) task. Regional brain morphology was assessed using ex-vivo high-resolution magnetic resonance imaging (MRI). Voxel-based morphometry of segmented MRI images revealed larger white matter volumes in anterior commissure and corpus callosum of HD rats compared with LD rats. HD rats also showed significantly larger regional volumes of dorsolateral orbitofrontal cortex, striatum, amygdala, hippocampus, midbrain, sub-thalamic nucleus, and cerebellum. By contrast, the medial prefrontal cortex was significantly smaller in HD rats compared with LD rats with no significant group differences in whole brain, ventricular, or cerebrospinal fluid volumes. These findings show that limbic cortico-basal ganglia structures implicated in impulse control disorders are distinct in rats that are vulnerable to develop compulsive behavior. Such abnormalities may be relevant to the etiology of compulsive disorders in humans.

Reduced Expression of the Htr2a, Grin1, and Bdnf Genes and Cognitive Inflexibility in a Model of High Compulsive Rats.

Compulsivity is a core symptom in different psychopathological disorders, characterized by excessive behaviors and behavioral inflexibility. The selection of high drinker (HD) versus low drinker (LD) rats by schedule-induced polydipsia (SIP) is a valid model for studying the compulsive phenotype. The compulsive HD rats showed cognitive inflexibility and reduced serotonin 2A (5-HT2A) receptor binding levels in the frontal cortex (FC). According to that, we hypothesize that compulsive HD rats might have an alteration in the cognitive control domain regarding inflexibility, assessed by spatial memory on the Morris Water Maze (MWM), working and reference memory by the Radial Arm Maze, and behavioral deficits in stimulus processing by the Novel Object Recognition test. The possible underlying mechanisms might be linked to the brain gene expression of 5HT2A, 5HT2C, glutamate NMDA receptors, and brain-derived neurotrophic factor (BDNF) in FC, hippocampus, and amygdala. HD rats confirmed a cognitive inflexibility profile on the reversal condition in the MWM compared to LD rats, while no differences were observed on stimulus processing, spatial, and working memory. Moreover, HD rats showed a reduced expression of the Htr2a, Grin1, and Bdnf genes in FC. Furthermore, there was a negative correlation between the relative expression of the Htr2a, Grin1, and Bdnf genes in FC and the level of compulsive water intake in HD rats on SIP. These data reveal that cognitive inflexibility may not be associated with a memory or stimulus processing deficit in compulsive individuals but may result by a region-specific alteration of the Htr2a, Grin1, and Bdnf gene expression in FC.

Behavioral domains in compulsive rats: implications for understanding compulsive spectrum disorders.

Introduction: Compulsive behavior has been proposed as a transdiagnostic trait observed in different neuropsychiatric disorders, such as obsessive-compulsive disorder, autism, and schizophrenia. Research Domain Criteria (RDoC) strategy could help to disentangle the neuropsychological basis of compulsivity for developing new therapeutic and preventive approaches. In preclinical research, the selection of high-drinker (HD) vs. low-drinker (LD) animals by schedule-induced polydipsia (SIP) is considered a putative model of compulsivity, which includes a well-differentiated behavioral pattern. Methods: The purpose of this research was to assess the cognitive control and the negative valence system domains in a phenotype of compulsive HD rats. After the selection of animals as HD or LD, we assessed behavioral inflexibility by probabilistic spatial reversal learning (PSRL), motor and cognitive impulsivity by variable delay-to-signal (VDS), and risky decision-making by rodent gambling task (rGT). Results: HD rats performed fewer reversals and showed less probability of pressing the same lever that was previously reinforced on PSRL, more premature responses after the exposure to longer delays on VDS, and more disadvantageous risky choices on rGT. Moreover, HD animals performed more perseverative responses under the punishment period on rGT. Discussion: These results highlight that HD compulsive phenotype exhibits behavioral inflexibility, insensitivity to positive feedback, waiting impulsivity, risky decision-making, and frustrative non-reward responsiveness. Moreover, these findings demonstrate the importance of mapping different behavioral domains to prevent, treat, and diagnose compulsive spectrum disorders correctly.

Avoidance and inhibitory control are possible transdiagnostic traits? A systematic review in animal models.

In clinical research, aberrant avoidance behavior and inhibitory control deficit have a high comorbidity in different psychopathological disorders. Therefore, avoidance and impulsive and/or compulsive behaviors might be classified as transdiagnostic traits, where the assessment through animal models could address evidence of their contribution as neurobehavioral mechanisms in psychopathology. The objective of the present review has been to assess the avoidance trait and the implication of inhibitory control behaviors, through studies using passive and active avoidance tests in rodents, and a preclinical model using selective breeding of high- or low-avoidance Roman rats (RHA, RLA). A systematic search strategy was carried out in the PubMed and Web of Science databases, where a total of 40 studies were accepted in the qualitative synthesis. The results of the different studies reviewed pointed to a relation between a reduced avoidance profile in passive avoidance (PA) with impulsive decision making and novelty-seeking behaviors; an increased avoidance profile in PA with compulsive drinking; a high active avoidance profile, including RHA rats, with different types of impulsivity and novelty- seeking behaviors; and regarding compulsivity depending on its measure, a low active avoidance profile, including RLA rats, has been associated with increased anxiety in the EPM and increased grooming, while a high active avoidance profile, including RHA rats, has been associated with increased rearing, compulsive drinking including alcohol, and cognitive inflexibility. The results have been discussed in terms of environmental factors and the underlying mechanisms between these possible transdiagnostic traits in psychopathology.

Differential Neurobiological Markers in Phenotype-stratified Rats Modeling High or Low Vulnerability to Compulsive Behavior: A Narrative Review.

Compulsivity is a key manifestation of inhibitory control deficit and a cardinal symptom in different neuropsychopathological disorders such as obsessive-compulsive disorder, schizophrenia, addiction, and attention-deficit hyperactivity disorder. Schedule-induced polydipsia (SIP), is an animal model to study compulsivity. In this procedure, rodents develop excessive and persistent drinking behavior under different food-reinforcement schedules, that are not related to homeostatic or regulatory requirements. However, there are important individual differences that support the role of high-drinker HD rats as a compulsive phenotype, characterized in different paradigms by inhibitory response deficit, cognitive inflexibility, and resistant to extinction behavior; with significant differences in response to pharmacological challenges, and relevant neurobiological alterations in comparison with the control group, the non-compulsive low drinker LD group on SIP. The purpose of this review is to collate and update the main findings on the neurobiological bases of compulsivity using the SIP model. Specifically, we reviewed preclinical studies on SIP, that have assessed the effects of serotonergic, dopaminergic, and glutamatergic drugs; leading to the description of the neurobiological markers, such as the key role of the serotonin 5-HT2A receptor and glutamatergic signaling in a phenotype vulnerable to compulsivity as high drinker HD rats selected by SIP. The review of the main findings of HD rats on SIP helps in the characterization of the preclinical compulsive phenotype, disentangles the underlying neurobiological, and points toward genetic hallmarks concerning the vulnerability to compulsivity.

Negative valence system as a relevant domain in compulsivity: review in a preclinical model of compulsivity.

Compulsive behavior is observed in different neuropsychiatric disorders such as Obsessive-Compulsive Disorder (OCD), anxiety, phobia, schizophrenia and addiction. Compulsivity has been proposed as a transdiagnostic symptom, where the Research Domain Criteria (RDoC) strategy could help to understand its neuropsychological basis for a better understanding, and development of therapeutic and preventive strategies. However, research on compulsivity has been focused on the cognitive control domain, and the contribution of an altered negative valence system has been less considered. In this review, we collate the main findings in an animal model of compulsivity, the high drinker (HD) rats selected by Schedule-Induced Polydipsia (SIP) regarding these two research domains. This preclinical model of compulsivity has shown a phenotype characterized by a lack of behavioral inhibition, impulsive decision-making and cognitive inflexibility. Moreover, the results in compulsive HD rats, suggests that there is also a relevant alteration in the emotional dimension, linked to the negative valence system domain, as for example by: the increased perseverative responses in a withdrawal condition, associated with the behavioral construct of frustrative non-reward; and an inhibition or extinction deficit in memory retrieval associated with an alteration in the behavioral response to sustained threat. However, the precise nature of the link between these shared altered domains, cognitive control and negative valence system, remains unknown. These results point towards relevant behavioral aspects of the compulsive phenotype that should be taken into account when studying the vulnerability to compulsivity that could help in the development of a better transdiagnostic assessment, preventive and therapeutic strategies.

Infectious Salmon Anemia Virus Infectivity Is Determined by Multiple Segments with an Important Contribution from Segment 5.

Infectious salmon anemia virus (ISAV) is the etiological agent of infectious salmon anemia. It belongs to the genus isavirus, one of the genera of the Orthomyxoviridae family, as does Influenzavirus A. The ISAV genome comprises eight negative-sense single-stranded RNA segments that code for at least 10 proteins. Although some ISAV strains can reach 100% mortality rates, the factors that determine isavirus infectivity remain unknown. However, some studies suggest that segments 5 and 6 are responsible for the different degrees of virulence and infectivity among ISAV subtypes, unlike the influenza A virus, where most segments are involved in the virus infectivity. In this work, synthetic reassortant viruses for the eight segments of ISAV were generated by reverse genetics, combining a highly virulent virus, ISAV 752_09 (HPR7b), and an avirulent strain, SK779/06 (HPR0). We characterized the rescued viruses and their capacity to replicate and infect different cell lines, produce plaques in ASK cells, and their ability to induce and modulate the cellular immune response in vitro. Our results show that the majority of ISAV segments are involved in at least one of the analyzed characteristics, segment 5 being one of the most important, allowing HPR0 viruses, among other things, to produce plaques and replicate in CHSE-214 cells. We determined that segments 5 and 6 participate in different stages of the viral cycle, and their compatibility is critical for viral infection. Additionally, we demonstrated that segment 2 can modulate the cellular immune response. Our results indicate a high degree of genetic compatibility between the genomic segments of HPR7b and HPR0, representing a latent risk of reassortant that would give rise to a new virus with an unknown phenotype.

Prenylated Flavonoids with Potential Antimicrobial Activity: Synthesis, Biological Activity, and In Silico Study.

Prenylated flavonoids are an important class of naturally occurring flavonoids with important biological activity, but their low abundance in nature limits their application in medicines. Here, we showed the hemisynthesis and the determination of various biological activities of seven prenylated flavonoids, named 7-13, with an emphasis on antimicrobial ones. Compounds 9, 11, and 12 showed inhibitory activity against human pathogenic fungi. Compounds 11, 12 (flavanones) and 13 (isoflavone) were the most active against clinical isolated Staphylococcus aureus MRSA, showing that structural requirements as prenylation at position C-6 or C-8 and OH at positions C-5, 7, and 4' are key to the antibacterial activity. The combination of 11 or 12 with commercial antibiotics synergistically enhanced the antibacterial activity of vancomycin, ciprofloxacin, and methicillin in a factor of 10 to 100 times against drug-resistant bacteria. Compound 11 combined with ciprofloxacin was able to decrease the levels of ROS generated by ciprofloxacin. According to docking results of S enantiomer of 11 with ATP-binding cassette transporter showed the most favorable binding energy; however, more studies are needed to support this result.

Rapid sequence modification in the highly polymorphic region (HPR) of the hemagglutinin gene of the infectious salmon anaemia virus (ISAV) suggests intra-segmental template switching recombination.

The ISAV has a genome composed of eight segments of (-)ssRNA, segment 6 codes for the hemagglutinin-esterase protein, and has the most variable region of the genome, the highly polymorphic region (HPR), which is unique among orthomyxoviruses. The HPR has been associated with virulence, infectivity and pathogenicity. The full length of the HPR is called HPR0 and the strain with this HPR is avirulent, in contrast to strains with deleted HPR that are virulent to varying degrees. The molecular mechanism that gives rise to the different HPRs remains unclear. Here, we studied in vitro the evolution of reassortant recombinant ISAV (rISAV) in Atlantic salmon head kidney (ASK) cells. To this end, we rescued and cultivated a set of rISAV with different segment 6-HPR genotypes using a reverse genetics system and then sequencing HPR regions of the viruses. Our results show rapid multiple recombination events in ISAV, with sequence insertions and deletions in the HPR, indicating a dynamic process. Inserted sequences can be found in four segments of the ISAV genome (segments 1, 5, 6, and 8). The results suggest intra-segmental heterologous recombination, probably by class I and class II template switching, similar to the proposed segment 5 recombination mechanism.

Biological Properties and Absolute Configuration of Flavanones From Calceolaria thyrsiflora Graham.

Flavanones (-)-(2S)-5,4'-dihydroxy-7-methoxyflavanone (1) and (-)-(2S)-5,3',4'-trihydroxy-7-methoxyflavanone (2) were isolated from the extracts of Calceolaria thyrsiflora Graham, an endemic perennial small shrub growing in the central zone of Chile. The absolute configuration of these compounds was resolved by optical rotation experiments and in silico calculations. Three analogs (3, 4, and 5) were synthesized to do structure-activity relationships with the biological assays studied. Biological tests revealed that only flavanone 2 exhibited a moderate inhibitory activity against the methicillin-resistant strain S. aureus MRSA 97-77 (MIC value of 50 µg/ml). In addition, flavanone 2 showed a potent, selective, and competitive inhibition of 5-hLOX, which supports the traditional use of this plant as an anti-inflammatory in diseases of the respiratory tract. Also, 2 exhibited cytotoxic and selective effects against B16-F10 (8.07 ± 1.61 µM) but 4.6- and 17-fold lesser activity than etoposide and taxol.

Development of an Isavirus minigenome system to study the function of the pocket RNA-binding domain of the viral nucleoprotein (NP) in salmon cells.

The Isavirus is an orthomyxovirus with a genome composed of eight segments of negative single-strand RNA (-ssRNA). It has been proposed that the eight genomic segments of the Isavirus are organized as a ribonucleoprotein (RNP) complex called a minigenome, which contains all the viral RNA segments, a viral heterotrimeric polymerase and multiple copies of the viral nucleoprotein (NP). Here, we develop an Isavirus minigenome system and show the importance of the formation of active RNPs and the role of viral NP R189, R194, R302 and K325 residues in the NP RNA-binding domain in the context of RNPs. The results indicate it is possible to generate a minigenome in salmon cells, a composite ISAV RNPs with EGFP-based chimeric vRNA with heterotrimeric polymerase (PB1, PB2, PA) and NP protein using CMV-based auxiliary plasmids. It was also shown that NP R189, R194, R302 and K325 residues are important to generate viral mRNA from the constituted RNPs and a detectable reporter protein. This work is the first salmon cell-based minigenome assay for the Isavirus, which was evaluated by a bioinformatic and functional study of the NP protein in viral RNPs, which showed that correct NP-vRNA interaction is key to the functioning of RNPs.

Deficient mitochondrial biogenesis in IL-2 activated NK cells correlates with impaired PGC1-α upregulation in elderly humans.

Immunosenescence has been described as age-associated changes in the immune function which are thought to be responsible for the increased morbidity with age. Human Natural Killer (NK) cells are a specialized heterogeneous subpopulation of lymphocytes involved in immune defense against tumor and microbial diseases. Interestingly, aging-related NK cell dysfunction is associated with features of aging such as tumor incidence, reduced vaccination efficacy, and short survival due to infection. It is known that NK cell effector functions are critically dependent on cytokines and metabolic activity. Our aim was to determine whether there is a difference in purified human NK cell function in response to high concentration of IL-2 between young and elder donors. Here, we report that the stimulation of human NK cells with IL-2 (2000 U/mL) enhance NK cell cytotoxic activity from both young and elderly donors. However, while NK cells from young people responded to IL-2 signaling by increasing mitochondrial mass and mitochondrial membrane potential, no increase in these mitochondrial functional parameters was seen in purified NK cells from elderly subjects. Moreover, as purified NK cells from the young exhibited an almost three-fold increase in PGC-1α expression after IL-2 (2000 U/mL) stimulation, PGC-1α expression was inhibited in purified NK cells from elders. Furthermore, this response upon PGC-1α expression after IL-2 stimulation promoted an increase in ROS production in NK cells from elderly humans, while no increase in ROS production was observed in NK cells of young donors. Our data show that IL-2 stimulates NK cell effector function through a signaling pathway which involves a PGC-1α-dependent mitochondrial function in young NK cells, however it seems that NK cells from older donors exhibit an altered IL-2 signaling which affects mitochondrial function associated with an increased production of ROS which could represent a feature of NK cell senescence.

Preparation of Novel Homodimers Derived from Cytotoxic Isoquinolinequinones. A Twin Drug Approach.

The synthesis of five novel homodimers is reported based on the anilinoisoquinolinequinone scaffold. In these twin-drug derivatives, two units of the anilinoquinone pharmacophores are linked through a methylene spacer. The formation of dimers was achieved by reaction of isoquinolinequinones with 4, 4'-diaminodiphenylmethane via a sequence of two oxidative amination reactions. A preliminary in vitro screening of the homodimers reveals moderate to high cytotoxic activities against MDA-MB-21 breast adenocarcinoma and B16-F10 murine metastatic melanoma cell lines. The asymmetrical homodimer 15 stands out due to its cytotoxic potencies at submicromolar concentrations and high selectivity index (mean IC50 = 0.37 µM; SI = 6.97) compared to those of etoposide (mean IC50 = 3.67; SI = 0.32) and taxol (mean IC50 = 0.35; SI = 0.91) employed as reference anticancer drugs.

Dead Tumor Cells Expressing Infectious Salmon Anemia Virus Fusogenic Protein Favor Antigen Cross-Priming In Vitro.

Antigen cross-presentation is a crucial step in the assembly of an antitumor immune response leading to activation of naïve CD8 T cells. This process has been extensively used in clinical trials, in which dendritic cells generated in vitro are loaded with tumor antigens and then autotransplanted to the patients. Recently, the use of autologous transplant of dendritic cells fused with dying tumor cells has demonstrated good results in clinical studies. In this work, we generated a similar process in vivo by treating mice with dead tumor cells [cell bodies (CBs)] expressing the fusogenic protein of the infectious salmon anemia virus (ISAV). ISAV fusion protein retains its fusogenic capability when is expressed on mammalian cells in vitro and the CBs expressing it facilitates DCs maturation, antigen transfer by antigen-presenting cells, and increase cross-presentation by DCs in vitro. Additionally, we observed in the melanoma model that CBs with or without ISAV fusion protein reduce tumor growth in prophylactic treatment; however, only ISAV expressing CBs showed an increase CD4 and CD8 cells in spleen. Overall, our results suggest that CBs could be used as a complement with other type of strategies to amplify antitumor immune response.

PGC-1α-Dependent Mitochondrial Adaptation Is Necessary to Sustain IL-2-Induced Activities in Human NK Cells.

Human Natural Killer (NK) cells are a specialized heterogeneous subpopulation of lymphocytes involved in antitumor defense reactions. NK cell effector functions are critically dependent on cytokines and metabolic activity. Among various cytokines modulating NK cell function, interleukin-2 (IL-2) can induce a more potent cytotoxic activity defined as lymphokine activated killer activity (LAK). Our aim was to determine if IL-2 induces changes at the mitochondrial level in NK cells to support the bioenergetic demand for performing this enhanced cytotoxic activity more efficiently. Purified human NK cells were cultured with high IL-2 concentrations to develop LAK activity, which was assessed by the ability of NK cells to lyse NK-resistant Daudi cells. Here we show that, after 72 h of culture of purified human NK cells with enough IL-2 to induce LAK activity, both the mitochondrial mass and the mitochondrial membrane potential increased in a PGC-1α-dependent manner. In addition, oligomycin, an inhibitor of ATP synthase, inhibited IL-2-induced LAK activity at 48 and 72 h of culture. Moreover, the secretion of IFN-γ from NK cells with LAK activity was also partially dependent on PGC-1α expression. These results indicate that PGC-1α plays a crucial role in regulating mitochondrial function involved in the maintenance of LAK activity in human NK cells stimulated with IL-2.

Neuroendocrine mechanisms for immune system regulation during stress in fish.

In the last years, the aquaculture crops have experienced an explosive and intensive growth, because of the high demand for protein. This growth has increased fish susceptibility to diseases and subsequent death. The constant biotic and abiotic changes experienced by fish species in culture are challenges that induce physiological, endocrine and immunological responses. These changes mitigate stress effects at the cellular level to maintain homeostasis. The effects of stress on the immune system have been studied for many years. While acute stress can have beneficial effects, chronic stress inhibits the immune response in mammals and teleost fish. In response to stress, a signaling cascade is triggered by the activation of neural circuits in the central nervous system because the hypothalamus is the central modulator of stress. This leads to the production of catecholamines, corticosteroid-releasing hormone, adrenocorticotropic hormone and glucocorticoids, which are the essential neuroendocrine mediators for this activation. Because stress situations are energetically demanding, the neuroendocrine signals are involved in metabolic support and will suppress the "less important" immune function. Understanding the cellular mechanisms of the neuroendocrine regulation of immunity in fish will allow the development of new pharmaceutical strategies and therapeutics for the prevention and treatment of diseases triggered by stress at all stages of fish cultures for commercial production.

Relationship between drug use and psychopathological variables of risk in university students / Relaciones entre el consumo de drogas y variables de riesgo psicopatológicas en jóvenes universitarios

Background: Research has been studying the relationships between drug use and the risk of suffering psychopathological disorders. This study analyzed the relationships existing between this use and certain psychotic disorder risk variables: hallucination, schizotypy and cognitive fusion. Method: Several screening questionnaires on drug use (CAGE), a questionnaire on «cognitive fusion» (TAFS), another on hallucination proneness (LSHS-R) and another on schizotypy (O-LIFE-R) were given to a sample of 308 students at the University of Almería with a mean age of 19.51 years (SD= 2.11). Results: The results found show how cognitive fusion is positively related to use of cannabis and cocaine, the scores on the schizotypy scale correlated positively with use of alcohol and cannabis, and the scores on a hallucination proneness correlated positively to use of cannabis. Regression equations were found that predicted the use of these substances from the variables of vulnerability to suffering from schizophrenia spectrum disorders. Conclusions: The results show an association between drug use and the risk variables studied (AU)

Antecedentes: la investigación ha venido estudiando las relaciones entre el consumo de sustancias y el riesgo de padecimiento de trastornos psicopatológicos. En este estudio se han analizado las relaciones existentes entre dicho consumo y determinadas variables de riesgo para los trastornos psicóticos: alucinaciones, esquizotipia y fusión cognitiva. Método: se administraron diversos cuestionarios de screening relativos al consumo de drogas (CAGE), un cuestionario de «fusión cognitiva» (TAFS), otro de predisposición a las alucinaciones (LSHS-R) y otro de esquizotipia (O-LIFE-R), a una muestra de 308 estudiantes universitarios de la Universidad de Almería, con una media de edad de 19,51 años (DT= 2,11). Resultados: los resultados hallados muestran cómo la fusión cognitiva se relaciona positivamente con el consumo de cannabis y cocaína; cómo las puntuaciones en la escala de esquizotipia correlacionan positivamente con el consumo de alcohol y cannabis y cómo las puntuaciones en una escala de predisposición a las alucinaciones correlaciona positivamente con el consumo de cannabis. Se establecieron ecuaciones de regresión que predecían el consumo de dichas sustancias a partir de las variables de vulnerabilidad al padecimiento de trastornos del espectro esquizofrénico. Conclusiones: los resultados muestran una asociación entre el consumo de sustancias y las variables de riesgo estudiadas (AU)

Relationship between drug use and psychopathological variables of risk in university students.

BACKGROUND: Research has been studying the relationships between drug use and the risk of suffering psychopathological disorders. This study analyzed the relationships existing between this use and certain psychotic disorder risk variables: hallucination, schizotypy and cognitive fusion. METHOD: Several screening questionnaires on drug use (CAGE), a questionnaire on "cognitive fusion" (TAFS), another on hallucination proneness (LSHS-R) and another on schizotypy (O-LIFE-R) were given to a sample of 308 students at the University of Almeria with a mean age of 19.51 years (SD= 2.11). RESULTS: The results found show how cognitive fusion is positively related to use of cannabis and cocaine, the scores on the schizotypy scale correlated positively with use of alcohol and cannabis, and the scores on a hallucination proneness correlated positively to use of cannabis. Regression equations were found that predicted the use of these substances from the variables of vulnerability to suffering from schizophrenia spectrum disorders. CONCLUSIONS: The results show an association between drug use and the risk variables studied.

Natural killer cell cytotoxicity is not regulated by folic acid in vitro.

OBJECTIVES: Folate supplementation may be associated with an increased risk of developing several types of cancer and a derangement of immune function. Among the latter, Natural killer (NK) cells are involved in non-MHC-restricted natural immunity against malignant target cells. Abnormalities in NK cell number or function have been associated with a higher cancer risk. The aim of this study was to study in vitro the possible effect of different concentrations of 5-methyltetrahydrofolic acid (5-MTHF) or folic acid on NK cell cytotoxic function, and expression of the stimulatory and inhibitory receptors KIRDL4, KIRDL3, and NKG2D. METHODS: Volunteer-derived peripheral mononuclear cells (PBMC) and highly enriched NK cells (95% CD56+ CD16+) were grown in folic acid free-RPMI 1640, supplemented either with folic acid or 5-MTHF (15-100 nM) during 72 h to 96 h. RESULTS: No differences in the cytolytic activity of PBMC and enriched NK cells were observed. After 96 h of in vitro culture without folate or supplemented with FA or 5-MTHF (30 or 100 nM), there were no changes in the percentage of HPNK receptor-positive cells. CONCLUSIONS: Our data indicate that a high dose of 5-MTHF or folic acid does not influence NK cell function in vitro.

Mechanisms and clinical correlates of sperm DNA damage.

Among the different DNA anomalies that can be present in the male gamete, DNA fragmentation is the most frequent, particularly in infertile subjects. There is now consistent evidence that a sperm containing fragmented DNA can be alive, motile, morphologically normal and able to fertilize an oocyte. There is also evidence that the oocyte is able to repair DNA damage; however, the extent of this repair depends on the type of DNA damage present in the sperm, as well as on the quality of the oocyte. Thus, it is important to understand the possible consequences of sperm DNA fragmentation (SDF) for embryo development, implantation, pregnancy outcome and the health of progeny conceived, both naturally and by assisted reproductive technology (ART). At present, data on the consequences of SDF for reproduction are scarce and, in many ways, inconsistent. The differences in study conclusions might result from the different methods used to detect SDF, the study design and the inclusion criteria. Consequently, it is difficult to decide whether SDF testing should be carried out in fertility assessment and ART. It is clear that there is an urgent need for the standardisation of the methods and for additional clinical studies on the impact of SDF on ART outcomes.