Dysregulation of methylation and expression of imprinted genes in oocytes and reproductive tissues in mice of advanced maternal age.

PURPOSE: To evaluate reproductive outcomes in aged compared to young female mice, and determine associated methylation and expression of imprinted genes in reproductive tissues. METHODS: Fetal, placental, and ovarian tissue were collected on d16.5 of pregnancy from young (4-5 weeks) and aged (15 months) mice. Uterine tissue and in vivo matured oocytes were collected from non-pregnant females. Methylation of imprinted genes was determined by restriction enzyme based assays, and transcript abundance of imprinted and nutrient supply genes were analyzed by quantitative PCR (qPCR). RESULTS: Maternal age was associated with fetal growth restriction and placental overgrowth. In maternally aged mice, methylation was minimally dysregulated in fetal tissue, while placental tissue showed aberrant methylation and transcript abundance of imprinted genes. Ovarian methylation and gene expression was severely dysregulated, although oocyte gene expression was only minimally altered. Abundance of Kcnq1 transcripts was significantly (P < 0.05) increased in oocytes obtained from aged females compared to young females. Gene expression was also severely dysregulated in the uterus, including nutrient transport genes. CONCLUSION: Fetal and placental growth abnormalities correspond to aberrant methylation and gene expression in reproductive tissues from maternally aged mice. Significant alterations in gene expression and methylation in the aged ovary suggests that the follicular environment may be compromised. Aberrant methylation and expression of imprinted genes in the aged uterus may contribute to reduced implantation. Maternal age negatively affects imprinted gene methylation and expression in both germ cells and somatic cells of the reproductive tract, contributing to the reduced fertility observed with advanced maternal age.

Fatty acid metabolism during maturation affects glucose uptake and is essential to oocyte competence.

Fatty acid ß-oxidation (FAO) is essential for oocyte maturation in mice. The objective of this study was to determine the effect of etomoxir (a FAO inhibitor; 100 µM), carnitine (1 mM), and palmitic acid (1 or 100 µM) during maturation on metabolism and gene expression of the oocyte and cumulus cells, and subsequent embryo development in the mouse. Carnitine significantly increased embryo development, while there was a decrease in development following maturation with 100 µM palmitic acid or etomoxir (P<0.05) treatment. Glucose consumption per cumulus-oocyte complex (COC) was decreased after treatment with carnitine and increased following etomoxir treatment (P<0.05). Intracellular oocyte lipid content was decreased after carnitine or etomoxir exposure (P<0.05). Abundance of Slc2a1 (Glut1) was increased after etomoxir treatment in the oocyte and cumulus cells (P<0.05), suggesting stimulation of glucose transport and potentially the glycolytic pathway for energy production when FAO is inhibited. Abundance of carnitine palmitoyltransferase 2 (Cpt2) tended to increase in oocytes (P=0.1) after treatment with 100 µM palmitic acid and in cumulus cells after exposure to 1 µM palmitic acid (P=0.07). Combined with carnitine, 1 µM palmitic acid increased the abundance of Acsl3 (P<0.05) and Cpt2 tended to increase (P=0.07) in cumulus cells, suggesting FAO was increased during maturation in response to stimulators and fatty acids. In conclusion, fatty acid and glucose metabolism are related to the mouse COC, as inhibition of FAO increases glucose consumption. Stimulation of FAO decreases glucose consumption and lipid stores, positively affecting subsequent embryo development, while an overabundance of fatty acid or reduced FAO negatively affects oocyte quality.

Alterations in the transcriptome of porcine oocytes derived from prepubertal and cyclic females is associated with developmental potential.

The developmental competence of oocytes is progressively attained as females approach puberty. The poor quality of prepubertally derived oocytes suggests that essential processes during cytoplasmic maturation have not been completed. The objective of this experiment was to identify genes in oocytes that are associated with good (cyclic females) and poor (prepubertal females) developmental competence. Development to the blastocyst stage in vitro was significantly decreased in oocytes derived from prepubertal females compared with cyclic females (5.26 and 12.86%, respectively). Approximately 10% of the oocyte transcriptome was differentially expressed between in vitro-matured oocytes derived from cyclic and prepubertal females (P < 0.05); 58% of differentially expressed genes had increased transcript abundance in oocytes derived from cyclic females. Genes involved in the metabolism and regulation of biological processes had increased transcript abundance in oocytes derived from cyclic females, whereas genes involved in translation were increased in prepubertally derived oocytes. Quantitative PCR confirmed differential expression (P < 0.05) for 6 out of 11 selected genes [DPYD (dihydropyrimidine dehydrogenase), RDH11 (retinol dehydrogenase 11), SFRS4 (serine/arginine-rich splicing factor 4), SFRS7 (serine/arginine-rich splicing factor 7), TL4 (transcribed loci 4), and TOP2B (topoisomerase II ß)] that were differentially expressed with greater than a 2-fold change by microarray, although 3 of these genes, DPYD, TL4, and TOP2B, were in opposing directions by the 2 methods. In conclusion, expression of multiple genes involved in metabolism and translation was significantly altered in oocytes from prepubertal females compared with cyclic females, which was associated with reduced in vitro development to the blastocyst stage. These genes may represent important cellular mechanisms that regulate oocyte quality.

Economic downturns and population mental health: research findings, gaps, challenges and priorities.

Prior research suggests that the current global economic crisis may be negatively affecting population mental health. In that context, this paper has several goals: (1) to discuss theoretical and conceptual explanations for how and why economic downturns might negatively affect population mental health; (2) present an overview of the literature on the relationship between economic recessions and population mental health; (3) discuss the limitations of existing empirical work; and (4) highlight opportunities for improvements in both research and practice designed to mitigate any negative impact of economic declines on the mental health of populations. Research has consistently demonstrated that economic crises are negatively associated with population mental health. How economic downturns influence mental health should be considered in policies such as social protection programs that aim to promote recovery.

Women's preferences for obstetric care in rural Ethiopia: a population-based discrete choice experiment in a region with low rates of facility delivery.

BACKGROUND: Delivery attended by skilled professionals is essential to reducing maternal mortality. Although the facility delivery rate in Ethiopia's rural areas is extremely low, little is known about which health system characteristics most influence women's preferences for delivery services. In this study, women's preferences for attributes of health facilities for delivery in rural Ethiopia were investigated. METHODS: A population-based discrete choice experiment (DCE) was fielded in Gilgel Gibe, in southwest Ethiopia, among women with a delivery in the past 5 years. Women were asked to select a hypothetical health facility for future delivery from two facilities on a picture card. A hierarchical Bayesian procedure was used to estimate utilities associated with facility attributes: distance, type of provider, provider attitude, drugs and medical equipment, transport and cost. RESULTS: 1006 women completed 8045 DCE choice tasks. Among them, 93.8% had delivered their last child at home. The attributes with the greatest influence on the overall utility of a health facility for delivery were availability of drugs and equipment (mean ß=3.9, p<0.01), seeing a doctor versus a health extension worker (mean ß=2.1, p<0.01) and a receptive provider attitude (mean ß=1.4, p<0.01). CONCLUSION: Women in rural southwest Ethiopia who have limited personal experience with facility delivery nonetheless value health facility attributes that indicate high technical quality: availability of drugs and equipment and physician providers. Well-designed policy experiments that measure the contribution of quality improvements to facility delivery rates in Ethiopia and other countries with low health service utilisation and high maternal mortality may inform national efforts to reduce maternal mortality.

Primary kidney allograft dysfunction due to myeloma-cast nephropathy: a case report.

Cast nephropathy is a rare event among renal transplants, usually associated with multiple myeloma or light chain nephropathy. Herein we have reported primary graft dysfunction early posttransplantation due to cast nephropathy, associated with thrombotic microangiopathy.

The influence of antisense oligonucleotides against STAT5 on the regulation of normal haematopoiesis in a bone marrow model.

Cytokines and growth factors that take part in the regulation of haematopoietic cell development activate many signalling pathways in target cells. The STAT5 (signal transducers and activators of transcription) proteins are members of a family of signal transducers and activators of transcription that can be activated after cytokine stimulation. Their binding to promoters of different genes influences cell proliferation, differentiation and survival. It is suggested that they play an important role in haematopoiesis, however, the question of the real function of STAT5 proteins requires further examination. The aim of our study was to investigate the role of STAT5 in the proliferation and apoptosis of normal haematopoietic bone marrow cells derived from heparinized cadaveric organ donors (HCOD). We applied antisense oligodeoxynucleotides (ODNs) to block STAT5A and STAT5B at the mRNA level and the reverse transcription polymerase chain reaction method to study STAT5 mRNA expression in the cells after incubation with ODNs. Moreover, we performed Western blot analysis of the STAT5A protein after exposure to antisense STAT5A. We analysed the clonogenicity of the colony-forming unit of granulocytes-macrophages and the burst-forming unit of erythrocytes in methylcellulose cultures according to the type and the dose of ODNs. We also examined apoptosis induced in bone marrow mononuclear and CD34(+) cells by employing annexin V staining and the TUNEL method using flow cytometry (FACScan). We found that the perturbation of STAT5 expression decreased the clonogenicity of bone marrow haematopoietic cells. However, we did not observe any significant increase in the percentage of apoptotic cells after incubation with antisense ODNs. It was concluded that the STAT5 proteins play a significant role in the proliferation of human bone marrow cells harvested from HCOD. These proteins might be critical in the regulation of haematopoiesis, especially under stress conditions.

An optimization of isolation of early hematopoietic cells from heparinized cadaveric organ donors.

Heparinized cadaveric organ donors are a potential source of hematopoietic cells for transplantation purposes. The aim of this study was to optimize the harvest of early hematopoietic cells from cadaver donors. Accordingly, we noticed that bone marrow cells harvested from cadaver donors should be resuspended in RPMI or Iscove's medium supplemented with heparin or ACD. Bags with harvested marrow should contain 20% atmosphere air during short-term storage/transportation. Finally, we noticed that BMMNC survive short-term storage better if the collected marrow is not depleted of erythrocytes.

Factors that impact on immediate graft function in patients after renal transplantation.

Kidney transplantation has become therapy of choice for patients with end-stage renal failure. However, many factors may cause graft rejection or delayed graft function, both of which decrease the prognosis for graft survival. For transplantologists the most important endeavor is to eliminate factors responsible for shortening graft function and to find those predictive of immediate graft function. The aim of the study was to investigate which factors influence early graft function. We retrospectively reviewed 442 renal transplant patients performed between 1990 and 1995 in two Szezecin units. All patients received an identical immunosuppressive drug schedule. Three hundred twelve patients who displayed immediate graft function were included in the study group to analyze donor and recipient age and sex, etiology of ESRD, HLA compatibility AB0 and Rh compatibility cold ischemia time, warm ischemia time, antileukocytes antibodies (PRA), and period of dialysis therapy before transplantation. We observed statistical significance for HLA and AB0 compatibility, younger donor age, and shorter cold ischemia time as the most important factors predictive of early graft function and an improved prognosis for graft survival.

Variations in temperature of the donor kidney during cold ischemia time and subsequent assessment of reperfusion using the application of thermovision camera.

Cold ischemia time (CIT) and graft reperfusion events are important nonimmunological factors that influence kidney graft function. The optimal temperature for the organs during CIT ranges from 4 degrees C to 8 degrees C. However, preservation of the designated temperature is usually not controlled during standard storage procedures. Aspects of initial graft reperfusion are usually assessed indirectly. Better evaluation of the effectiveness of the early blood supply may improve the surgical outcome. The aim of the study was to monitor the temperature during CIT in the kidney and surrounding area and subsequently to assess graft reperfusion using thermography. Temperature values of the area surrounding the kidney were registered during 8 organ procurements. We observed that the area surrounding the kidney displayed the optimal temperature range only during the first 5 minutes of CIT; later the temperature oscillated between 1 degrees C and 3.5 degrees C. The study proved that the temperature of the preservation fluid is frequently below prescribed. Analysis of the thermograms of 40 transplanted kidneys with the use of a thermovision camera ThermaCAM SC500 showed usefulness of this method to assess reperfusion and blood distribution in the transplanted kidney.

Impact of selected factors on early graft function in patients after renal transplantation.

Transplantation is the best treatment for end-stage renal diseases. For transplantologists, it is most important to know the factors that worsen graft survival prognosis. The aim of the study was to investigate factors predictive of graft loss and shortened graft survival. We retrospectively reviewed 442 renal transplant patients between 1990 and 1995 in two Szczecin units, all of whom received a triple-drug immunosuppressive regimen. One hundred thirty patients showed graft disorders such as delayed graft function or primary nonfunction. The occurrence of these disorders was examined as a function of donor and recipient age and sex, cause of ESRD, HLA compatibility, ABO and Rh compatibility, cold ischemia time, warm ischemia time, antileukocyte antibody level (PRA), and period of dialysis therapy before transplantation. The study showed that a high maximal PRA level, incompatibility for ABO group, and a longer warm ischemia time increase the probability of early graft function disorders.

The binding interface database (BID): a compilation of amino acid hot spots in protein interfaces.

SUMMARY: To make information about protein interactive function easily accessible, we are mining the primary scientific literature for detailed data about protein interfaces. The Binding Interface Database (BID) organizes the vast amount of protein interaction information into tables, graphical contact maps and descriptive functional profiles. Currently data on 170 interacting protein pairs are available with over 1300 mutations described. AVAILABILITY: The BID database is freely available at http://tsailab.org/BID/ To have your protein of interest entered, contact Tiffany Fischer (tiffbrink@neo.tamu.edu) or Jerry Tsai at the email below

Factors influencing low incidence of living donors for renal transplantation in Poland.

PURPOSE OF THE STUDY: To answer the question: "Why are only 3.8% of kidney transplantations in Poland from living donors?" MATERIAL AND METHODS: The research was conducted using data from anonymous polls addressed to family members of dialyzed patients (potential donors). RESULTS: Almost everybody (98%) had heard about the possibility of treating renal insufficiency by kidney transplantation, but only 77% knew about the possibility of giving their own kidney to a related person. Merely 36% had been informed about this treatment method by a doctor. The most common fears within respondents were those of: health worsening after donating a kidney--45%, and the operation itself (taking an organ)--39%.

Heparinized cadaveric organ donors (HCOD)--a potential source of hematopoietic cells for transplantation and gene therapy.

BACKGROUND: Hematopoietic stem cells (HSC) from unrelated HLA-matched heparinized cadaveric organ donors (HCOD) are a new potential source of cells for transplantation and gene therapy. In addition, these cells could also be used as adjuvant therapy to increase microchimerism and graft tolerance after transplantations of various solid organs. Our purpose was to develop an efficient method for harvesting hematopoietic cells from HCODs, METHODS: Bone marrow cells were harvested from pelvic bones and/or vertebral bodies from 50 adult HCODs before or up to 3 hr after disconnecting the donor from the respirator. Subsequently, we evaluated the hematological and gasometric parameters of aspirated marrow samples as well as the proliferative potential, viability, and expression of CD34 and AC133 antigens on these cells. RESULTS: We noticed that up to 2-3 hr after disconnecting the donor from the respirator bone marrow cavities do not clot and remain uninfected and that it is possible to aspirate bone marrow mononuclear cells in quantities sufficient to perform allotransplantation. Nevertheless, due to the developing hypoxia and acidosis of the hematopoietic microenvironment the number and proliferative potential of CD34+ and AC133+ cells gradually decreases. Hence, to obtain viable early hematopoietic cells, bone marrow should be aspirated without delay; optimally before HCOD is disconnected from the respirator or at the very latest 2 hr after organ harvest. CONCLUSIONS: Collectively, our results show that early hemopoietic cells may be efficiently harvested from HCOD in large quantities and used for research and/or transplantation purposes. We postulate to create an international network of banks in which hemopoietic stem cells from HCODs could be preserved for therapeutic purposes.

Survival of Rhizobium phaseoli in Coal-Based Legume Inoculants.

The long-term survival of Rhizobium phaseoli strains 127K17, 127K26, and 127K35 in legume inoculants prepared with eight different coals (one strain and one coal per inoculant) was studied. The coals used were Pennsylvania anthracite, bituminous coals from Illinois, Pennsylvania, and Utah, lignite from North Dakota and Texas, and subbituminous coals from New Mexico and Wyoming; they ranged in pH from 4.7 to 7.5 All coals, with the exceptions of Illinois bituminous coal and Texas lignite (pH's of 5.0 and 4.7, respectively), supported the growth and survival of all R. phaseoli strains. All coal-based inoculants in which rhizobial viability was maintained had more than 10 rhizobia per g for at least 7 months, and most contained more than 10 rhizobia per g after 12 months. It appears that most coals, regardless of grade or source, may be acceptable carriers for R. phaseoli inoculants.