Suppression of Mamu-AG by RNA Interference

Problem  The role of placental major histocompatibility complex (MHC) class I molecules in pregnancy is not well understood. Mamu-AG, the rhesus monkey homology of human leukocyte antigen (HLA)-G expressed in the human placenta, was targeted for degradation by RNA interference (RNAi), a powerful tool to aid in determining gene function, to determine the effect that this knockdown has on NK cell function.
Method of study  A series of potential target short hairpin RNA (shRNA) sequences to suppress Mamu-AG expression was screened, which identified an optimal sequence to use in transfection experiments. Knockdown in two different Mamu-AG-expressing cell lines was measured by flow cytometry. Cytotoxicity assays were performed to correlate Mamu-AG expression with NK cell cytotoxicity.
Results  Decreased expression of Mamu-AG by short interfering RNA (siRNA) (70–80%) in cell types tested was associated with increased lysis of Mamu-AG target cells.
Conclusion  Target sequences have been identified that knocked down Mamu-AG expression by RNAi and increased lysis by NK cells. This supports the concept that NK cell receptors recognize this placental non-classical MHC class I molecule.     

The striopallidal pathway is involved in antiparkinsonian-like effects of the blockade of group I metabotropic glutamate receptors in rats

The aim of the study was to examine the influence of the blockade of group I metabotropic glutamate receptors (mGluRs) on the haloperidol-induced catalepsy and proenkephalin mRNA expression in the rat striatum. Bilateral, intrastriatal injection of AIDA ((RS)-1-aminoindan-1,5-dicarboxylic acid, 3-15 microg/0.5 microl), a selective antagonist of group I mGluRs, inhibited catalepsy induced by haloperidol (0.5 mg/kg i.p.). Repeated intrastriatal AIDA administrations (3 x 15 microg/0.5 microl, 3 h apart) counteracted the haloperidol-induced (3 x 1.5 mg/kg s.c., 3 h apart) increase in the proenkephalin mRNA expression in that structure. The present study indicates that the blockade of the striatal group I mGluRs may inhibit parkinsonian akinesia by normalizing the function of the striopallidal pathway.     

Craniofacial skeletal defects of adult zebrafish Glypican 4 (knypek) mutants

The heparan sulfate proteoglycan Glypican 4 (Gpc4) is part of the Wnt/planar cell polarity pathway, which is required for convergence and extension during zebrafish gastrulation. To observe Glypican 4‐deficient phenotypes at later stages, we rescued gpc4^?/? (knypek) homozygotes and raised them for more than one year. Adult mutants showed diverse cranial malformations of both dermal and endochondral bones, ranging from shortening of the rostral‐most skull to loss of the symplectic. Additionally, the adult palatoquadrate cartilage was disorganized, with abnormal chondrocyte orientation. To understand how the palatoquadrate cartilage normally develops, we examined a juvenile series of wild type and mutant specimens. This identified two novel domains of elongated chondrocytes in the larval palatoquadrate, which normally form prior to endochondral ossification. In contrast, gpc4^?/? larvae never form these domains, suggesting a failure of chondrocyte orientation, though not differentiation. Our findings implicate Gpc4 in the regulation of zebrafish cartilage and bone morphogenesis. Developmental Dynamics 238:2550–2563, 2009. © 2009 Wiley‐Liss, Inc.     

Patient safety culture in obstetrics and gynecology and neonatology units: the nurses’ and the midwives’ opinion

Background: Patients treated in health care facilities that provide services in the fields of obstetrics, gynecology, and neonatology are especially vulnerable. Large multidisciplinary teams of physicians, multiple invasive and noninvasive diagnostic and therapeutic procedures, and the use of advanced technologies increase the probability of adverse events. The evaluation of knowledge about patient safety culture among nurses and midwives working in such units and the identification of critical areas at a health care institution would reduce the number of adverse events and improve patient safety. The aim of the study was to evaluate the opinion of nurses and midwives working in clinical departments that provide services in the fields of obstetrics, gynecology, and neonatology about patient safety culture and to explore potential predictors for the overall perception of safety.

Methods: We used the Hospital Survey on Patient Safety Culture (HSOPSC) to evaluate nurses’ and midwives’ opinion about patient safety issues. The overall response rate in the survey was 100% (n?=?233).

Results: The analysis of the dimensions of safety on the unit level showed that the respondents’ most positive evaluations were in the Organizational Learning – Continuous Improvement (73.2%) and Feedback and Communication about Error (66.8%) dimensions, and the most negative evaluations in the Non-punitive Response to Error (33.5%) and Staffing (44.6%) dimensions. On the hospital level, the evaluation of the safety dimensions ranged between 41.4 and 56.8%. The percentage of positive responses in the outcome dimensions Frequency of Events Reported was 82.4%. We found a significant association between the outcome dimension Frequency of Events Reported and the Hospital Management Support for Patient Safety and Feedback and Communication about Error Dimensions.

Conclusions: On the hospital level, the critical domains in health care facilities that provide services in the fields of obstetrics, gynecology, and neonatology were Teamwork Across Hospital Units, and on the unit level – Communication Openness, Teamwork Within Units, Non-punitive Response to Error, and Staffing. The remaining domains were seen as having a potential for improvement.     

Meta-analysis of putative human bornavirus sequences fails to provide evidence implicating Borna disease virus in mental illness

All Borna disease virus (BDV) sequences derived from human specimens published till date were thoroughly analysed and compared to sequences of BDV laboratory strains and to BDV sequences from animals which succumbed to classical Borna disease (BD). Despite high sequence conservation of the BDV genome, animal-derived BDV sequences clustered according to their geographic origin. However, in marked contrast, human-derived BDV sequences did not cluster according to their geographic origin but showed high sequence identities to BDV laboratory strains and animal-derived BDVs handled in the laboratories reporting the human strains. Japanese, US, Australian and French human-derived BDV sequences proved to be identical or very similar to animal-derived BDV sequences from Germany, although the human specimens were collected hundreds to thousands of miles away from the central European BD endemic regions. These findings suggest that previous studies linking BDV to human neuropsychiatric disease may have been compromised by inadvertent sample contamination.     

Acute appendicitis in children under 3 years of age. Diagnostic and therapeutic problems

Aim: Analysis of diagnostic and therapeutic problems in acute appendicitis in children below 3 years of age. Material and methods: The analysis was based on medical data of 53 children under 3 years of age, treated in our department for acute appendicitis in the years 1988-2008. Among 53 children, 29 (53.7%) were admitted directly to the surgical department and 24 (45.3%) were transferred from the regional pediatric department. In the period of 1 month before admission to the surgical department 13 patients (24.5%) were treated as outpatients due to acute respiratory or alimentary tract infection. On the basis of the data from the case histories, the most frequent symptoms and their duration were evaluated, as well as the clinical signs, intraoperative diagnosis and the postoperative course. Results: The most frequent symptoms and clinical signs in this group of children were: abdominal pain, vomiting and fever, present in 83.0%, 75.5%, and 67.0% patients respectively. The mean time of the symptoms' duration was 3.6 days. The most frequently found physical signs on admission to the surgical ward were: abdominal pain on palpation, increased tonus of abdominal muscles and abdominal distension. On laparotomy gangrenous appendicitis was found in 49% of the children operated. In 24.5% of patients perforation of the appendix was confirmed. Further complications occurred in 9 children (16.9%). The average stay in hospital after the operation lasted 7.9 days. Conclusions: 1. Acute appendicitis in small children is a diagnostic problem not only for primary health care doctors but also for experienced pediatricians and pediatric surgeons. 2. Early surgical consultation should be a standard procedure in small children with acute symptoms of various locations when there is accompaning abdominal pain, not reacting to conservative treatment. Surgical consultation is also indicated in children under 3 years of age with relapses of abdominal pain. 3. Clinical signs of appendicitis in children aged less than 3 years, may differ from those in older children due to changes in their immunological reactivity. This problem should be included in under-graduate and post-graduate medical studies.     

Near-ultraviolet light perceived by the retina generates the signal suppressing melatonin synthesis in the chick pineal gland-an involvement of NMDA glutamate receptors

Exposure of dark-adapted chicks to near ultraviolet (UV-A) light significantly decreased melatonin (MEL) content and the activity of serotonin N-acetyltransferase (AA-NAT; the penultimate and key regulatory enzyme in MEL production) in the pineal glands. Significant reduction in MEL level and AA-NAT activity was also found in pineals of animals whose heads were covered with black opaque tape, an observation suggesting that in the chicken UV-A light perceived by the eyes alone is capable of affecting MEL synthesis in the pineal gland. Covering the chick's eyes, in addition to the head, totally blocked the studied UV-A action. Although SCH 23390 (a selective D1-dopamine receptor antagonist), injected directly into both eyes at a dose of 10 nmol/eye, prevented the decline in pineal AA-NAT activity produced by retinal illumination with white light, the drug did not modify the UV-A light-evoked decrease in the enzyme activity. MK-801 (a selective antagonist of NMDA glutamate receptors; 1 nmol/eye) abolished the suppressive action of UV-A light on pineal AA-NAT activity, but it was inactive in the case of white light. Intraocularly injected sulpiride and CNQX (selective antagonists of D2-dopamine and AMPA/kainite glutamate receptors, respectively) had no effect on the actions of both UV-A and white light (acting on the eyes only) on pineal AA-NAT activity. It is concluded that in the chick retinally perceived UV-A light generates a signal which suppresses MEL production in the pineal gland. At the level of the retina, such signal does not involve dopamine, but is dependent on the stimulation of NMDA glutamate receptors.     

Epitope of the vaccine-type Bordetella pertussis strain 186 lipooligosaccharide and antiendotoxin activity of antibodies directed against the terminal pentasaccharide-tetanus toxoid conjugate

Lipooligosaccharides (LOS) isolated from Bordetella pertussis strains 186 and 606 were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and high-resolution magic angle spinning nuclear magnetic resonance (NMR). These analyses distinguished between the LOS of strains 186 and 606, suggesting that the structure of LOS in B. pertussis is heterogeneous. The pentasaccharide was selectively cleaved from LOS of B. pertussis strain 186, purified, and covalently linked to a monomer fraction of tetanus toxoid. Injection of rabbits with the neoglycoconjugate emulsified in complete Freund's adjuvant yielded immunoglobulin G antibodies that were reactive with the LOS. These antibodies reacted strongly with B. pertussis LOS possessing the complete dodecasaccharide, as determined by an enzyme-linked immunosorbent assay, immunoblotting, and flow cytometry with intact, live bacterial cells. The binding epitope within the pentasaccharide was investigated by saturation transfer difference (STD) NMR spectroscopy. Protons H-1 and H-4 of the terminal alpha-D-GlcpNAc and proton H-6 and protons of an N-methyl group at H-4 of 3-substituted beta-L-FucpNAc4NMe exhibited the largest saturation transfers. STD NMR experiments confirmed that the immunodominant epitope recognized by the antineoglycoconjugate antibodies is located predominantly in the distal trisaccharide of B. pertussis 186 LOS. The antipentasaccharide antibodies induced by the conjugate inhibited the secretion of tumor necrosis factor alpha, interleukin-6, and NO by LOS-stimulated J774A.1 cells.     

A genome-wide association study identifies a novel susceptibility locus for renal cell carcinoma on 12p11.23

Renal cell carcinoma (RCC) is the most lethal urologic cancer. Only two common susceptibility loci for RCC have been confirmed to date. To identify additional RCC common susceptibility loci, we conducted an independent genome-wide association study (GWAS). We analyzed 533 191 single nucleotide polymorphisms (SNPs) for association with RCC in 894 cases and 1516 controls of European descent recruited from MD Anderson Cancer Center in the primary scan, and validated the top 500 SNPs in silico in 3772 cases and 8505 controls of European descent involved in the only published GWAS of RCC. We identified two common variants in linkage disequilibrium, rs718314 and rs1049380 (r(2) = 0.64, D?' = 0.84), in the inositol 1,4,5-triphosphate receptor, type 2 (ITPR2) gene on 12p11.23 as novel susceptibility loci for RCC (P = 8.89 × 10(-10) and P = 6.07 × 10(-9), respectively, in meta-analysis) with an allelic odds ratio of 1.19 [95% confidence interval (CI): 1.13-1.26] for rs718314 and 1.18 (95% CI: 1.12-1.25) for rs1049380. It has been recently identified that rs718314 in ITPR2 is associated with waist-hip ratio (WHR) phenotype. To our knowledge, this is the first genetic locus associated with both cancer risk and WHR.     

The role of genetic breast cancer susceptibility variants as prognostic factors

Recent genome-wide association studies identified 11 single nucleotide polymorphisms (SNPs) associated with breast cancer (BC) risk. We investigated these and 62 other SNPs for their prognostic relevance. Confirmed BC risk SNPs rs17468277 (CASP8), rs1982073 (TGFB1), rs2981582 (FGFR2), rs13281615 (8q24), rs3817198 (LSP1), rs889312 (MAP3K1), rs3803662 (TOX3), rs13387042 (2q35), rs4973768 (SLC4A7), rs6504950 (COX11) and rs10941679 (5p12) were genotyped for 25 853 BC patients with the available follow-up; 62 other SNPs, which have been suggested as BC risk SNPs by a GWAS or as candidate SNPs from individual studies, were genotyped for replication purposes in subsets of these patients. Cox proportional hazard models were used to test the association of these SNPs with overall survival (OS) and BC-specific survival (BCS). For the confirmed loci, we performed an accessory analysis of publicly available gene expression data and the prognosis in a different patient group. One of the 11 SNPs, rs3803662 (TOX3) and none of the 62 candidate/GWAS SNPs were associated with OS and/or BCS at P<0.01. The genotypic-specific survival for rs3803662 suggested a recessive mode of action [hazard ratio (HR) of rare homozygous carriers=1.21; 95% CI: 1.09-1.35, P=0.0002 and HR=1.29; 95% CI: 1.12-1.47, P=0.0003 for OS and BCS, respectively]. This association was seen similarly in all analyzed tumor subgroups defined by nodal status, tumor size, grade and estrogen receptor. Breast tumor expression of these genes was not associated with prognosis. With the exception of rs3803662 (TOX3), there was no evidence that any of the SNPs associated with BC susceptibility were associated with the BC survival. Survival may be influenced by a distinct set of germline variants from those influencing susceptibility.