Genetic and cytological analyses reveal the recombination landscape of a partially differentiated plant sex chromosome in kiwifruit.

BACKGROUND: Angiosperm sex chromosomes, where present, are generally recently evolved. The key step in initiating the development of sex chromosomes from autosomes is the establishment of a sex-determining locus within a region of non-recombination. To better understand early sex chromosome evolution, it is important to determine the process by which recombination is suppressed around the sex determining genes. We have used the dioecious angiosperm kiwifruit Actinidia chinensis var. chinensis, which has an active-Y sex chromosome system, to study recombination rates around the sex locus, to better understand key events in the development of sex chromosomes. RESULTS: We have confirmed the sex-determining region (SDR) in A. chinensis var. chinensis, using a combination of high density genetic mapping and fluorescent in situ hybridisation (FISH) of Bacterial Artificial Chromosomes (BACs) linked to the sex markers onto pachytene chromosomes. The SDR is a subtelomeric non-recombining region adjacent to the nucleolar organiser region (NOR). A region of restricted recombination of around 6 Mbp in size in both male and female maps spans the SDR and covers around a third of chromosome 25. CONCLUSIONS: As recombination is suppressed over a similar region between X chromosomes and between and X and Y chromosomes, we propose that recombination is suppressed in this region because of the proximity of the NOR and the centromere, with both the NOR and centromere suppressing recombination, and this predates suppressed recombination due to differences between X and Y chromosomes. Such regions of suppressed recombination in the genome provide an opportunity for the evolution of sex chromosomes, if a sex-determining locus develops there or translocates into this region.

Measuring affiliation in group therapy for substance use disorders in the Women's Recovery Group study: Does it matter whether the group is all-women or mixed-gender?

BACKGROUND AND OBJECTIVES: A Stage II, two-site randomized clinical trial compared the manualized, single-gender Women's Recovery Group (WRG) to mixed-gender group therapy (Group Drug Counseling; GDC) and demonstrated efficacy. Enhanced affiliation and support in the WRG is a hypothesized mechanism of efficacy. This study sought to extend results of the previous small Stage I trial that showed the rate of supportive affiliative statements occurred more frequently in WRG than GDC. METHODS: Participants (N = 158; 100 women, 58 men) were 18 years or older, substance dependent, and had used substances within the past 60 days. Women were randomized to WRG (n = 52) or GDC (n = 48). Group therapy videos were coded by two independent raters; Rater 1 coded 20% of videos (n = 74); Rater 2 coded 25% of videos coded by Rater 1 (n = 19). RESULTS: The number of affiliative statements made in WRG was 66% higher than in GDC. Three of eight affiliative statement categories occurred more frequently in WRG than GDC: supportive, shared experience, and strategy statements. DISCUSSION AND CONCLUSIONS: This larger Stage II trial provided a greater number of group therapy tapes available for analysis. Results extended our previous findings, demonstrating both greater frequency of all affiliative statements, as well as specific categories of statements, made in single-gender WRG than mixed-gender GDC. SCIENTIFIC SIGNIFICANCE: Greater frequency of affiliative statements among group members may be one mechanism of enhanced support and efficacy in women-only WRG compared with standard mixed-gender group therapy for substance use disorders. (Am J Addict 2016;25:573-580).

Changes in sleep disruption in the treatment of co-occurring posttraumatic stress disorder and substance use disorders.

Sleep disruption appears not only to reflect a symptom of posttraumatic stress disorder (PTSD), but also a unique vulnerability for its development and maintenance. Studies examining the impact of psychosocial treatments for PTSD on sleep symptoms are few and no studies to date of which we are aware have examined this question in samples with co-occurring substance use disorders. The current study is a secondary analysis of a large clinical trial comparing 2 psychological treatments for co-occurring PTSD and substance use disorders. Women (N = 353) completed measures of PTSD at baseline, end of treatment, and 3-, 6-, and 12-month follow-ups. Results indicated that the prevalence of insomnia, but not nightmares, decreased during treatment, and that 63.8% of participants reported at least 1 clinical-level sleep symptom at the end of treatment. Improvement in sleep symptoms during treatment was associated with better overall PTSD outcomes over time, χ(2) (1) = 33.81, p < .001. These results extend the existing literature to suggest that residual sleep disruption following PTSD treatment is common in women with co-occurring PTSD and substance use disorders. Research on the benefits of adding sleep-specific intervention for those with residual sleep disruption in this population may be a promising future direction.

The cannabinoid WIN 55212-2 mitigates apoptosis and mitochondrial dysfunction after hypoxia ischemia.

Perinatal hypoxia-ischemia has significant mortality and morbidity due to there is still no specific treatment as a consequence of the complexities of hypoxic-ischemic pathophysiology. The aim of this work was to evaluate the effects of the cannabinoid agonist WIN 55212-2 on apoptotic cell death and mitochondrial dysfunction after perinatal asphyxia in fetal lambs. Animals were assigned to: one SHAM group and two hypoxic-ischemic groups that received a dose of 0.01 µg/kg WIN 55,212-2 (HI + WIN) or not (HI + VEH) after 60 min of partial occlusion of the umbilical cord, and sacrificed 3 h later. Different brain regions were separated for morphological studies, and the same territories were dissociated and analyzed by flow cytometry to quantify apoptosis, to determine mitochondrial integrity and transmembrane potential and to analyze intracellular calcium levels. Our results showed that WIN 55,212-2 reduced apoptotic cell death in all regions studied through the maintenance of mitochondrial integrity and functionality.

Treatment of esophageal perforation: A review of our experience at a tertiary referral hospital spanning the past 19 years.

INTRODUCTION AND AIMS: Esophageal perforation is an uncommon event that is a medical/surgical emergency, with a 15-30% mortality rate. The aim of the present study was to communicate our experience in the management of esophageal perforation, evaluating the different strategies utilized, in an effort to establish measures to guide decision-making in selecting treatment. MATERIALS AND METHODS: A retrospective descriptive study was conducted on patients diagnosed with esophageal perforation at our hospital center, within the time frame of 2000 and 2019. RESULTS: Over the past 19 years, 15 patients were diagnosed with esophageal perforation. Surgical treatment was carried out in 80% of the cases. Primary closure, reinforced with plasty, was performed in 67% of the patients, of whom 62.5% had early diagnosis and a 100% survival rate. Diagnosis was late in 37.5% of the cases, with a 33.3% survival rate. Esophagectomy and gastric pull-up were performed on 25% of the patients, 66.6% of whom had early diagnosis and a 100% survival rate. In the 33.3% that had late diagnosis, the mortality rate was 100%. Esophagectomy, with cervical esophagostomy and feeding jejunostomy, was performed on one of the patients (8.3%) that had early diagnosis and a 100% survival rate. CONCLUSIONS: The main survival predictor in esophageal perforation is the interval of time between the injury and its diagnosis, and in turn, the resulting treatment. Each patient with esophageal perforation should have individualized treatment to adequately manage the condition.

Characterization of Gene Expression in the Rat Brainstem After Neonatal Hypoxic-Ischemic Injury and Antioxidant Treatment.

The perinatal brainstem is known to be very vulnerable to hypoxic-ischemic events which can lead to deafness, swallowing dysfunction, and defective respiratory control. The aim of the present work was to evaluate the potential neuroprotective effects of nicotine, melatonin, resveratrol, and docosahexaenoic acid on the expression of a panel of genes in the brainstem following hypoxic-ischemic damage. Quantitative PCR was used to examine gene expression 3 and 12 h after the damage, and immunohistochemistry was employed to evaluate neurons, astrocytes, and synaptic vesicles 24 h post insult. We found that the expression of some immediate-early genes, as well as that of inflammatory genes TNF-α, COX2, and caspase 3, was upregulated in response to the insult. Twenty-four hours after the damage, the percentage of NeuN and synaptophysin immunolabeled cells was found to be reduced while GFAP expression was upregulated. No differences were observed in ROS gene expression following treatments.

Docosahexaenoic Acid Reduces Cerebral Damage and Ameliorates Long-Term Cognitive Impairments Caused by Neonatal Hypoxia-Ischemia in Rats.

As the interest in the neuroprotective possibilities of docosahexaenoic acid (DHA) for brain injury has grown in the recent years, we aimed to investigate the long-term effects of this fatty acid in an experimental model of perinatal hypoxia-ischemia in rats. To this end, motor activity, aspects of learning, and memory function and anxiety, as well as corticofugal connections visualized by using tracer injections, were evaluated at adulthood. We found that in the hours immediately following the insult, DHA maintained mitochondrial inner membrane integrity and transmembrane potential, as well as the integrity of synaptic processes. Seven days later, morphological damage at the level of the middle hippocampus was reduced, since neurons and myelin were preserved and the astroglial reactive response and microglial activation were seen to be diminished. At adulthood, the behavioral tests revealed that treated animals presented better long-term working memory and less anxiety than non-treated hypoxic-ischemic animals, while no difference was found in the spontaneous locomotor activity. Interestingly, hypoxic-ischemic injury caused alterations in the anterograde corticofugal neuronal connections which were not so evident in rats treated with DHA. Thus, our results indicate that DHA treatment can lead to long-lasting neuroprotective effects in this experimental model of neonatal hypoxia-ischemic brain injury, not only by mitigating axonal changes but also by enhancing cognitive performance at adulthood.

The V-ATPase A subunit gene (vma-1) from Giardia lamblia.

The sequence of the gene encoding the A subunit of the vacuolar type ATPase from Giardia lamblia is reported. Comparison of the encoded protein with the homologous subunits of eukaryotic and archaebacterial ATPases reveals high levels of similarity throughout the sequence (e.g., overall 49.1 and 44.6% identity to the homologous subunit from carrot and Halobacterium, respectively). An exception are three regions which are unique to the Giardia subunit. The largest of these regions contains motifs characteristic for eukaryotic spliceosomal introns; however, comparison to the cDNA shows that this region is also present in the mRNA.

Co-expression of laminin and a 67 kDa laminin-binding protein in teratocarcinoma embryoid bodies.

The synthesis of laminin chains is usually correlated to specific functions of laminin during embryo development. In this study we show that the CE44 teratocarcinoma embryoid bodies synthesize B1 and B2 chains of laminin as well as a 67 kDa laminin-binding protein while simultaneously differentiating into parietal endoderm. The intracystic presence of laminin and the 67 kDa cell surface laminin-receptor in teratocarcinoma differentiated cells suggest that the B chains of laminin play an important role in induction and/or mediation of cell differentiation and confirm the importance of laminin A chain in cell polarization and the supramolecular rearrangement of definitive basement membrane.

Denial of urinalysis-confirmed opioid use in prescription opioid dependence.

Although research has generally supported the validity of substance use self-reports, some patients deny urine-verified substance use. We examined the prevalence and patterns of denying urinalysis-confirmed opioid use in a sample of prescription opioid dependent patients. We also identified characteristics associated with denial in this population of increasing public health concern. Opioid use self-reports were compared with weekly urinalysis results in a 12-week multi-site treatment study for prescription opioid dependence. Among those who used opioids during the trial (n=246/360), 44.3% (n=109) denied urinalysis-confirmed opioid use, although usually only once (78%). Overall, 22.9% of opioid-positive urine tests (149/650) were denied on self-report. Multivariable analysis found that initially using opioids to relieve pain was associated with denying opioid use. These findings support the use of both self-reports and urine testing in treating prescription opioid dependence.

Initial response as a predictor of 12-week buprenorphine-naloxone treatment response in a prescription opioid-dependent population.

OBJECTIVE: Initial medication response has been shown to predict treatment outcome across a variety of substance use disorders, but no studies have examined the predictive power of initial response to buprenorphine-naloxone in the treatment of prescription opioid dependence. We therefore conducted a secondary analysis of data from the Prescription Opioid Addiction Treatment Study to determine whether initial response to buprenorphine-naloxone predicted 12-week treatment outcome in a prescription opioid-dependent population. METHOD: Using data from a multisite, randomized controlled trial of buprenorphine-naloxone plus counseling for DSM-IV prescription opioid dependence (June 2006-July 2009), we conducted a secondary analysis to investigate the relationship between initial medication response and 12-week treatment outcome to establish how soon the efficacy of buprenorphine-naloxone could be predicted (N = 360). Outcomes were determined from the Substance Use Report, a self-report measure of substance use, and confirmatory urinalysis. Predictive values were calculated to determine the importance of abstinence versus use at various time points within the first month of treatment (week 1, weeks 1-2, 1-3, or 1-4) in predicting successful versus unsuccessful treatment outcome (based on abstinence or near-abstinence from opioids) in the last 4 weeks of buprenorphine-naloxone treatment (weeks 9-12). RESULTS: Outcome was best predicted by medication response after 2 weeks of treatment. Two weeks of initial abstinence was moderately predictive of treatment success (positive predictive value = 71%), while opioid use in both of the first 2 weeks was strongly predictive of unsuccessful treatment outcome (negative predictive value [NPV] = 84%), especially when successful outcome was defined as total abstinence from opioids in weeks 9-12 (NPV = 94%). CONCLUSIONS: Evaluating prescription opioid-dependent patients after 2 weeks of buprenorphine-naloxone treatment may help determine the likelihood of successful outcome at completion of the current treatment regimen. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT00316277.

Cells from the inner mass of blastocyst as a source of neural derivates for differentiation studies.

Our results show that cells derived from the inner cell mass (ICM) show a clear tendency to differentiate into the neural lineage, showing both cells and structures in different degrees of differentiation. Among the experimental paradigms used to learn about neural differentiation, there have been several lines of investigation on stem cells, including embryonic stem (ES) cells isolated from the inner cell mass of embryo and also stem cells derived from embryonic carcinoma (EC). In this work, we have used a cellular line obtained from the inner cell mass of a blastocyst. The cells were cultured and after inoculated subcutaneously in syngenic mice. The neural differentiation was predominant, and could be observed both by morphological and immunohistochemical methods. It was represented by neural-tubes, neurons and glial cells, as expressed by the presence of Microtubule-associated protein-2 (MAP-2) and glial fibrilary acidic protein. Moreover, tyrosine hydroxilase positive labelling was found in neuron-like cells, which suggest the chatecolaminergic differentiation. These results show that isolation of cells from the inner mass of blastocyst represents an easy, reproducible and cheap source of neural derivates suitable for both in vivo and in vitro differentiation studies.

Presence of laminin and 67KDa laminin-receptor on endothelial surface of lung capillaries. An immunocytochemical study.

The existence of cell surface-associated molecules has been claimed to play a major role in cellular recognition and interaction. In this respect, different tumor cell lines express laminin and its receptor, and this expression has been correlated with metastatic potential. In the present work, we have studied, by electron microscopic immunolabeling methods, the presence of laminin and 67KDa laminin-receptor on the surface of endothelial cells of lung blood capillaries. To label these molecules, we have developed an easy method in which the labeling is carried out "in situ", in previously excised lungs. The presence of both molecules was observed on the luminal surface of endothelial capillaries and, in many cases, gold particles were associated to small open vesicles of the endothelial cells. The results suggest that these molecules, traditionally associated to extracellular matrix, are also expressed in cellular surface of the lung vascular bed.

Epigenetic modulation of differentiation in CE44 teratocarcinoma.

Teratocarcinoma is a mixed germ cell tumor histologically composed of embryonal carcinoma cells and embryonic and extraembryonic tissues. In the present work we have used the CE44 teratocarcinoma, which is a tumor cell line derived from the OTT6050 experimental tumor, to appreciate the influence the microenvironment has on the modulation of tumoral differentiation. For this, we have studied the development of CE44 teratocarcinoma in primary tumors (subcutaneous and intrasplenic) and in experimental metastases (hepatic and pulmonary). CE44 teratocarcinoma shows variations in its capacity for differentiation in so far as development is concerned and, in hepatic metastases, we noticed a reparative process of the intratumoral necrotic areas which in the same cases were substituted by loose connective tissue. Our results clearly suggest that the microenvironment is decisive in the biological behaviour of the teratocarcinoma cells and that epigenetic factors influence the capacity for differentiation of the undifferentiated tumoral cells.

The intein of the Thermoplasma A-ATPase A subunit: structure, evolution and expression in E. coli.

BACKGROUND: Inteins are selfish genetic elements that excise themselves from the host protein during post translational processing, and religate the host protein with a peptide bond. In addition to this splicing activity, most reported inteins also contain an endonuclease domain that is important in intein propagation. RESULTS: The gene encoding the Thermoplasma acidophilum A-ATPase catalytic subunit A is the only one in the entire T. acidophilum genome that has been identified to contain an intein. This intein is inserted in the same position as the inteins found in the ATPase A-subunits encoding gene in Pyrococcus abyssi, P. furiosus and P. horikoshii and is found 20 amino acids upstream of the intein in the homologous vma-1 gene in Saccharomyces cerevisiae. In contrast to the other inteins in catalytic ATPase subunits, the T. acidophilum intein does not contain an endonuclease domain.T. acidophilum has different codon usage frequencies as compared to Escherichia coli. Initially, the low abundance of rare tRNAs prevented expression of the T. acidophilum A-ATPase A subunit in E. coli. Using a strain of E. coli that expresses additional tRNAs for rare codons, the T. acidophilum A-ATPase A subunit was successfully expressed in E. coli. CONCLUSIONS: Despite differences in pH and temperature between the E. coli and the T. acidophilum cytoplasms, the T. acidophilum intein retains efficient self-splicing activity when expressed in E. coli. The small intein in the Thermoplasma A-ATPase is closely related to the endonuclease containing intein in the Pyrococcus A-ATPase. Phylogenetic analyses suggest that this intein was horizontally transferred between Pyrococcus and Thermoplasma, and that the small intein has persisted in Thermoplasma apparently without homing.

Determination of the physicochemical constants and spectrophotometric characteristics of the metallochromic Zincon and its potential use in biological systems.

In this work we report a detailed characterization of the metallochromic Zincon. Zincon forms complexes with Zn2+ and Cu2+, producing change in colour; the complexes with Fe2+, Mn2+ and Ca2+ cause the bleaching of the Zincon solutions. Mg2+ does not interact with Zincon nor does it change its spectral characteristics. The presence of Ca2+ and Mg2+ does not interfere with the spectral characteristics of the Zn-Zincon complex. The Kd, Ks and delta epsilon values for the complexes were determined. The delta epsilon values were very high, making this spectrophotometric method very sensitive. The complex Zn-Zincon is fully reversible; however, the complex Cu-Zincon is only partially reversible. The free Zincon, and the complexes Zn-Zincon and Cu-Zincon, does not partition into organic solvents, does not permeate liposome membrane, and neither does it interact with biological membranes. All these characteristics make the metallochromic Zincon useful in biological systems.

Presence of four stem cell populations in monolayer cultures derived from teratocarcinoma embryoid bodies.

In the present work, flow cytometry techniques together with morphologic studies were used to perform multiparametric analyses in cell cultures derived from CE44 teratocarcinoma embryoid bodies. The intrinsic cell parameters studied by flow cytometry were size (FALS), cytoplasmic complexity (ISS) and autofluorescence, expressed as LIGFL/FALS (green fluorescence intensity on a logarithmic scale/FALS). Our results showed that CE44 teratocarcinoma yields monolayers whose cells show a marked morphological heterogeneity and can be grouped according to flow cytometric criteria into four populations that remain stable throughout the entire time of culture. Moreover, these populations showed a different immunolabelling with the differentiation markers SSEA-1, TROMA-1 and anti-vimentin.

Horizontal transfer of ATPase genes--the tree of life becomes a net of life.

An ancient gene duplication gave rise to the catalytic and non-catalytic subunits of each of the three types of proton pumping ATPases: vacuolar, archaebacterial and eubacterial. Previously, this gene duplication has been used to root the universal tree of life. However, recent findings of archaebacterial type ATPases in eubacteria and of eubacterial type in an archaebacterium suggested that both types of ATPases may have been already present in the last common ancestor. Here we show that a phylogenetic analysis of these ATPase subunits indicates that this conclusion is premature. We suggest that horizontal gene transfer can explain the data. In addition, we show that the analysis of glutamate dehydrogenases data neither affirm nor contradict any particular placement of the last common ancestor in the universal tree of life. The prevalence and the mode of horizontal gene transfer is discussed.