Adverse drug events among adult inpatients: a meta-analysis of observational studies.

WHAT IS KNOWN AND OBJECTIVE: Studies in a number of countries have shown that adverse drug events (ADE) occur frequently among hospital inpatients. The objective of this study was to conduct a systematic review of observational studies of the frequency of ADE in adult inpatients and to examine factors associated with observed heterogeneity in the reported results. METHODS: The systematic review included observational studies, which identified and analysed ADE during hospitalization of adult inpatients. The literature search was conducted on MEDLINE, Embase, Lilacs and Google Scholar (January of 2000 to June of 2013). Article selection, quality assessment and information extraction were performed by two of the authors, working independently. Using the random-effects model, the proportion of patients with adverse events was used as an outcome measure. Proportion was estimated for subgroups based on event identification method: stimulated reporting (SR), retrospective monitoring (RM) and prospective monitoring (PM). For the latter group, meta-regression was used to identify sources of heterogeneity in the estimates. RESULTS AND DISCUSSION: Twenty-eight articles from the 7550 identified met our inclusion criteria. The articles were heterogeneous in terms of quality, outcome definition and event identification method and in the corresponding descriptions. Of the 28 articles selected, 25 were included in the corresponding quantitative summary: four used SR, six RM and 15 PM, returning incidences of 2·3% (CI 95%: 1·6-4·5), 8·7% (CI 95%: 4·8-15·3) and 21·3% (CI 95%: 15·7-28·3), respectively, and I(2) greater than 95%. There were other sources of heterogeneity, including the use of combined strategies within each subgroup. In the PM subgroup, using multivariate meta-regression model, no variables were found to associate with proportion. WHAT IS NEW AND CONCLUSION: Event frequency seems to associate with the event identification method. PM returned the highest estimates. This subgroup used a greater diversity of approaches for event identification and more diverse data sources. Improved recording of information on the event identification method, the characteristics of the events and the conduct of the study would enable more reliable and precise estimates of the frequency of ADE among hospital inpatients.

Genetic interactions within TFIIIC, the promoter-binding factor of yeast RNA polymerase III.

TFIIIC is a heteromultimeric protein, made of six distinct subunits in Saccharomyces cerevisiae, that binds to RNA polymerase III promoters and triggers the assembly of the transcription complex. The largest yeast subunit tau138, encoded by TFC3, binds to the B-box promoter element. This binding is defective in the temperature-sensitive mutant tfc3-G349E; the mutation responsible is located in one of two conserved motifs shared with the B-binding component of human TFIIIC. Rare dominant gain-of-function mutations that restore growth at high temperature were obtained following ultraviolet mutagenesis of tfc3-G349E. All of them resulted from single amino acid substitutions that alter the structure of TFIIIC. Three were due to reversion or intragenic suppression (TFC3-K754E and TFC3-L804H) events. Three were identical isolates of TFC6-E330K, a previously described mutation of the tau91 subunit. The remaining suppressors mapped in TFC4, and resulted in amino acid replacements in the second largest subunit of TFIIIC (tau131). With the exception TFC4-E711K, these affect positions that are invariant between the S. cerevisiae and Homo sapiens proteins, and are localised in conserved tetratricopeptide motifs. These findings demonstrate a close functional interaction between the two largest subunits of TFIIIC and underscore the importance of the tetratricopeptide motif of tau131.

A genetic look at the active site of RNA polymerase III.

rpc160-112, a mutant of the RNA polymerase III active site, is corrected in vivo by six second-site mutants obtained by random mutagenesis. These mutants introduce single-site amino acid replacements at the two large subunits of the enzyme. The mutated motifs are conserved in RNA polymerases I and II and, for some of them, in the bacterial enzyme, thus delineating key elements of the active site in eukaryotic RNA polymerases.

Changes in HOXB6 homeodomain protein structure and localization during human epidermal development and differentiation.

HOX homeodomain proteins are master developmental regulators, which are now thought to function as transcription factors by forming cooperative DNA binding complexes with PBX or other protein partners. Although PBX proteins exhibit regulated subcellular localization and function in the nucleus in other tissues, little data exists on HOX and PBX protein localization during skin development. We now show that the HOXB6 protein is expressed in the suprabasal layer of the early developing epidermis and throughout the upper layers of late fetal and adult human skin. HOXB6 signal is cytoplasmic throughout fetal epidermal development, but substantially nuclear in normal adult skin. HOXB6 protein is also partially nuclear in hyperproliferative skin conditions, but appears to be cytoplasmic in basal and squamous cell carcinomas. Although all three PBX genes are expressed in fetal epidermis, none of the three PBX proteins exhibit nuclear co-localization with HOXB6 in either fetal or adult epidermis. RNA and protein data suggest that a truncated HOXB6 protein, lacking the homeodomain, is expressed in undifferentiated keratinocytes and that the full-length protein is induced by differentiation. GFP-fusion proteins were used to demonstrate that the full-length HOXB6 protein is localized to the nucleus while the truncated protein is largely cytoplasmic. Taken together, these data suggest that during epidermal development the truncated HOXB6 isoform may function by a mechanism other than as DNA binding protein, and that most of the nuclear, homeodomain-containing HOXB6 protein does not utilize PBX proteins as DNA binding partners in the skin. Published 2000 Wiley-Liss, Inc.

[Long-term benzodiazepine use in women at a daycare center for older people]. / O uso prolongado de benzodiazepínicos em mulheres de um centro de convivência para idosos.

While experts recommend caution against long-term benzodiazepine use in the elderly, survey data suggests that the use of benzodiazepine increase with age. The patterns of benzodiazepine use and factors associated with long-term use in population at risk were studied with a standarlized questionnaire applied to 634 women over 60, who attended a daycare center for older people in Rio de Janeiro between May, 1992 and December, 1995. Prevalence of benzodiazepine use in the last 15 days was 21.3% (CI 95% 18.1-24.5), and prevalence of daily use for 12 or more months was 7.4% (CI 95% 5.4-9.4). In a multivaried analysis the amount of drugs being consumed displayed an important and progressive association with long-term benzodiazepine use, with OR = 2.77 (CI 95% 1.17-6.57) for those who take from four to six drugs, and OR = 7.62 (CI 95% 3.18-18.26) for those who take more than seven drugs. Insomnia (OR = 8.87 CI 95% 2.53-31.06) and chronic headache (OR = 3.53 CI 95% 1.82-6.89) have also been associated with this pattern of use.

Frequent co-expression of the HOXA9 and MEIS1 homeobox genes in human myeloid leukemias.

There is increasing evidence that HOX homeobox genes play a role in leukemogenesis. Recent studies have demonstrated that enforced co-expression of HOXA9 and MEIS1 in murine marrow leads to rapid development of myeloid leukemia, and that these proteins exhibit cooperative DNA binding. However, it is unclear whether co-activation of HOXA9 and MEIS genes is a common occurrence in human leukemias. We surveyed expression of HOXA9 and MEIS1 in 24 leukemic cell lines and 80 patient samples, using RNase protection analyses and immunohistochemistry. We demonstrate that the expression of HOXA9 and MEIS1 in leukemia cells is uniquely myeloid, and that these genes are commonly co-expressed in myeloid cell lines and in samples of acute myelogenous leukemia (AML) of all subtypes except in promyelocytic leukemia. While HOXA9 is expressed in most cases of chronic myelogenous leukemia, MEIS1 is weakly expressed or not at all. Immunohistochemical staining of selected AML samples showed moderate to high levels of HOXA9 protein, primarily cytoplasmic, in leukemic myeloblasts, with weaker and primarily nuclear staining for MEIS1. These data support the concept that co-activation of HOXA9 and MEIS1 is a common event in AML, and may represent a common pathway of many different oncogenic mutations.

[Quality assessment of drug use in the elderly]. / Avaliação da qualidade do uso de medicamentos em idosos.

INTRODUCTION: The objective is to evaluate the quality of medication utilization through the analysis of the pattern of usage, the degree of compliance to essential drug lists, therapeutic value and by drug interactions found among women over 60 years of age. METHODS: Six hundred thirty-four women enrolled at the Open University of the Third Age were studied. Data was collected through pattern-oriented, tested questionnaires. The variables examined were related to drugs and to drug utilization. The units of analysis used were the drugs and the individual. RESULTS: Of 634 women that participated in the study, 9,1% did not use drugs. The number of medications taken vary from 1 to 17. The average is 4,0 drugs/woman. Among the 2.510 pharmaceutical specialties mentioned by the interviewed, 538 different substances were identified. About 26% of the medications were in agreement with the recommendations of the World Health Organization and 17% with recommendations of the "Relação Nacional de Medicamentos Essenciais". Seventeen percent of the drugs are inappropriate for use in seniors; 14,1% of the women may suffer consequences for taking drugs of the same therapeutic class, and 15, 5% are exposed to interactions. CONCLUSIONS: The data suggest that the pattern of the medication utilization is considerably influenced by the medical prescription and that their quality is harmed by the low selectiveness of the pharmaceutical market

HOXA9 forms triple complexes with PBX2 and MEIS1 in myeloid cells.

Aberrant activation of the HOX, MEIS, and PBX homeodomain protein families is associated with leukemias, and retrovirally driven coexpression of HOXA9 and MEIS1 is sufficient to induce myeloid leukemia in mice. Previous studies have demonstrated that HOX-9 and HOX-10 paralog proteins are unique among HOX homeodomain proteins in their capacity to form in vitro cooperative DNA binding complexes with either the PBX or MEIS protein. Furthermore, PBX and MEIS proteins have been shown to form in vivo heterodimeric DNA binding complexes with each other. We now show that in vitro DNA site selection for MEIS1 in the presence of HOXA9 and PBX yields a consensus PBX-HOXA9 site. MEIS1 enhances in vitro HOXA9-PBX protein complex formation in the absence of DNA and forms a trimeric electrophoretic mobility shift assay (EMSA) complex with these proteins on an oligonucleotide containing a PBX-HOXA9 site. Myeloid cell nuclear extracts produce EMSA complexes which appear to contain HOXA9, PBX2, and MEIS1, while immunoprecipitation of HOXA9 from these extracts results in coprecipitation of PBX2 and MEIS1. In myeloid cells, HOXA9, MEIS1, and PBX2 are all strongly expressed in the nucleus, where a portion of their signals are colocalized within nuclear speckles. However, cotransfection of HOXA9 and PBX2 with or without MEIS1 minimally influences transcription of a reporter gene containing multiple PBX-HOXA9 binding sites. Taken together, these data suggest that in myeloid leukemia cells MEIS1 forms trimeric complexes with PBX and HOXA9, which in turn can bind to consensus PBX-HOXA9 DNA targets.

Cellular proliferation and transformation induced by HOXB4 and HOXB3 proteins involves cooperation with PBX1.

The products of PBX homeobox genes, which were initially discovered in reciprocal translocations occurring in human leukemias, have been shown to cooperate in the in vitro DNA binding with HOX proteins. Despite the growing body of data implicating Hox genes in the development of various cancers, little is known about the role of HOX-PBX interactions in the regulation of proliferation and induction of transformation of mammalian cells. We build on the existing model of Hox-induced transformation of Rat-1 cells to show that both cellular transformation and proliferation induced by Hoxb4 and Hoxb3 are greatly modulated by the levels of available PBX1 present in these cells. Furthermore, we show that the transforming capacity of these two HOX proteins depends on their conserved tetrapeptide and homeodomain regions which mediate binding to PBX and DNA, respectively. Taken together, results of this study demonstrate that cooperation between HOX and PBX proteins modulates cellular proliferation and strongly suggest that cooperative DNA binding by these two groups of proteins represent the basis for Hox-induced cellular transformation.

Loss of function of the homeobox gene Hoxa-9 perturbs early T-cell development and induces apoptosis in primitive thymocytes.

Hox homeobox genes play a crucial role in specifying the embryonic body pattern. However, a role for Hox genes in T-cell development has not been explored. The Hoxa-9 gene is expressed in normal adult and fetal thymuses. Fetal thymuses of mice homozygous for an interruption of the Hoxa-9 gene are one eighth normal size and have a 25-fold decrease in the number of primitive thymocytes expressing the interleukin-2 receptor (IL-2R, CD25). Progression to the double positive (CD4+CD8+) stage is dramatically retarded in fetal thymic organ cultures. This aberrant development is associated with decreased amounts of intracellular CD3 and T-cell receptor beta (TCRbeta) and reduced surface expression of IL-7R and E-cadherin. Mutant thymocytes show a significant increase in apoptotic cell death and premature downregulation of bcl-2 expression. A similar phenotype is seen in primitive thymocytes from adult Hoxa-9-/- mice and from mice transplanted with Hoxa-9-/- marrow. Hoxa-9 appears to play a previously unsuspected role in T-cell ontogeny by modulating cell survival of early thymocytes and by regulating their subsequent differentiation.

Drug surveillance: topics for discussion and prospects

Drug surveillance can be grouped into two areas. The first includes registration and inspection, activities that are more familiar to society at large. The second involves research and monitoring of adverse effects. Brazilian legislation has regulated this subject since 1970. A recent directive also provides for a national pharmaceutical surveillance system. This current article provides an overview of the history, definitions, pharmacological concepts, classification, and diagnosis of the adverse effects of drugs. An analysis is presented of the goals and sources of information for drug monitoring as well as some Brazilian experience in this field.

HOX homeobox genes exhibit spatial and temporal changes in expression during human skin development.

The spatial and temporal deployment of HOX homeobox genes along the spinal axis and in limb buds during fetal development is a key program in embryonic pattern formation. Although we have previously reported that several of the HOX homeobox genes are expressed during murine skin development, there is no information about developmental expression of HOX genes in human skin. We have now used reverse transcriptase polymerase chain reaction, in conjunction with a set of degenerate oligonucleotide primers, to identify a subset of HOX genes that are expressed during human fetal skin development. In situ hybridization analyses demonstrated that there were temporal and spatial shifts in expression of these genes. Strong HOXA4 expression was detected in the basal cell layers of 10 wk fetal epidermis and throughout the epidermis and dermis of 17 wk skin, whereas weak signal was present in the granular layer of newborn and adult skin. The expression patterns of HOXA5 and HOXA7 were similar, but their expression was weaker. In situ hybridization analysis also revealed strong HOXC4 and weaker HOXB7 expression throughout fetal development, whereas HOXB4 was expressed at barely detectable levels. Differential HOX gene expression was also observed in developing hair follicles, and sebaceous and sweat glands. None of the HOX genes examined were detected in the adult dermis.

Tau91, an essential subunit of yeast transcription factor IIIC, cooperates with tau138 in DNA binding.

Transcription factor IIIC (TFIIIC) (or tau) is a large multisubunit and multifunctional factor required for transcription of all class III genes in Saccharomyces cerevisiae. It is responsible for promoter recognition and TFIIIB assembly. We report here the cloning and characterization of TFC6, an essential gene encoding the 91-kDa polypeptide, tau91, present in affinity-purified TFIIIC. Tau91 has a predicted molecular mass of 74 kDa. It harbors a central cluster of His and Cys residues and has basic and acidic amino acid regions, but it shows no specific similarity to known proteins or predicted open reading frames. The TFIIIC subunit status of tau91 was established by the following biochemical and genetic evidence. Antibodies to tau91 bound TFIIIC-DNA complexes in gel shift assays; in vivo, a B block-deficient U6 RNA gene (SNR6) harboring GAL4 binding sites was reactivated by fusing the GAL4 DNA binding domain to tau91; and a point mutation in TFC6 (tau91-E330K) was found to suppress the thermosensitive phenotype of a tfc3-G349E mutant affected in the B block binding subunit (tau138). The suppressor mutation alleviated the DNA binding and transcription defects of mutant TFIIIC in vitro. These results indicated that tau91 cooperates with tau138 for DNA binding. Recombinant tau91 by itself did not interact with a tRNA gene, although it showed a strong affinity for single-stranded DNA.

AbdB-like Hox proteins stabilize DNA binding by the Meis1 homeodomain proteins.

Recent studies show that Hox homeodomain proteins from paralog groups 1 to 10 gain DNA binding specificity and affinity through cooperative binding with the divergent homeodomain protein Pbx1. However, the AbdB-like Hox proteins from paralogs 11, 12, and 13 do not interact with Pbx1a, raising the possibility of different protein partners. The Meis1 homeobox gene has 44% identity to Pbx within the homeodomain and was identified as a common site of viral integration in myeloid leukemias arising in BXH-2 mice. These integrations result in constitutive activation of Meis1. Furthermore, the Hoxa-9 gene is frequently activated by viral integration in the same BXH-2 leukemias, suggesting a biological synergy between these two distinct classes of homeodomain proteins in causing malignant transformation. We now show that the Hoxa-9 protein physically interacts with Meis1 proteins by forming heterodimeric binding complexes on a DNA target containing a Meis1 site (TGACAG) and an AbdB-like Hox site (TTTTACGAC). Hox proteins from the other AbdB-like paralogs, Hoxa-10, Hoxa-11, Hoxd-12, and Hoxb-13, also form DNA binding complexes with Meis1b, while Hox proteins from other paralogs do not appear to interact with Meis1 proteins. DNA binding complexes formed by Meis1 with Hox proteins dissociate much more slowly than DNA complexes with Meis1 alone, suggesting that Hox proteins stabilize the interactions of Meis1 proteins with their DNA targets.

The Abd-B-like Hox homeodomain proteins can be subdivided by the ability to form complexes with Pbx1a on a novel DNA target.

Previous studies showed that the Hox homeodomain proteins from paralog groups 1-8 display cooperative DNA binding with the non-Hox homeodomain protein Pbx, mediated by a canonical YPWM. Although the Abd-B-like Hox proteins in paralogs 9-13 lack this sequence, Hoxb-9 and Hoxa-10 were reported to bind with Pbx1a to DNA. We show that these interactions require a tryptophan 6 amino acids N-terminal to the homeodomain. Binding site selection for Hoxb-9 with Pbx1a yielded ATGATTTACGAC, containing a novel TTAC Hox-binding site adjacent to a Pbx site. In the presence of Pbx1a, Hoxb-9 and Hoxa-10 bound to targets containing either TTAC or TTAT. These data extend previous findings that interactions with Pbx define a Hox protein binding code for different DNA sequences across paralog groups 1 through 10. Members of the 11, 12, and 13 paralogs do not cooperatively bind DNA with Pbx1a, despite the presence of tryptophan residues N-terminal to the homeodomain in Hoxd-12 and Hoxd-13. Hoxa-11, Hoxd-12, or Hoxd-13, in the presence of Pbx1a, selected a TTAC Hox site but lacking a Pbx1a site. These data suggest that Abd-B-like Hox proteins bind to a novel TTAC site and can be divided by their cooperative binding to DNA with Pbx1a.

The nuclear Kluyveromyces lactis MRF1 gene encodes a mitochondrial class I peptide chain release factor that is important for cell viability.

We report the isolation and characterization of the Kluyveromyces lactis MRF1 gene encoding mitochondrial peptide chain release factor mRF-1. Over-expression of the KlMRF1 gene has a strong antisuppressive effect in a Saccharomyces cerevisiae mitochondrial nonsense suppressor strain. Inactivation of KlMRF1 results in a dual phenotype: most cells die after about 10-13 generations, while a small number of cells exceed this limit. We propose that the lethality is related to a loss of mitochondrial genome integrity. Surviving Klmrf1 cells are able to grow slowly on the non-fermentable substrate glycerol, indicating the existence of a second mitochondrial release factor activity. Our previous comparative analysis of class I release factors is refined by the incorporation of KlmRF-1 and ten recently identified prokaryotic release factor sequences.Keywords Kluyveromyces lactis middle dot Mitochondrial release factor middle dot MRF1 middle dot Peptide chain termination

Hox homeodomain proteins exhibit selective complex stabilities with Pbx and DNA.

Eight of the nine homeobox genes of the Hoxb locus encode proteins which contain a conserved hexapeptide motif upstream from the homeodomain. All eight proteins (Hoxb-1-Hoxb-8) bind to a target oligonucleotide in the presence of Pbx1a under conditions where minimal or no binding is detected for the Hox or Pbx1a proteins alone. The stabilities of the Hox-Pbx1a-DNA complexes vary >100-fold, with the proteins from the middle of the locus (Hoxb-5 and Hoxb-6) forming very stable complexes, while Hoxb-4, Hoxb-7 and Hoxb-8 form complexes of intermediate stability and proteins at the 3'-side of the locus (Hoxb-1-Hoxb-3) form complexes which are very unstable. Although Hox-b proteins containing longer linker sequences between the hexapeptide and homeodomains formed unstable complexes, shortening the linker did not confer complex stability. Homeodomain swapping experiments revealed that this motif does not independently determine complex stability. Naturally occurring variations within the hexapeptides of specific Hox proteins also do not explain complex stability differences. However, two core amino acids (tryptophan and methionine) which are absolutely conserved within the hexapeptide domains appear to be required for complex formation. Removal of N- and C-terminal flanking regions did not influence complex stability and the members of paralog group 4 (Hoxa-4, b-4, c-4 and d-4), which share highly conserved hexapeptides, linkers and homeodomains but different flanking regions, form complexes of similar stability. These data suggest that the structural features of Hox proteins which determine Hox-Pbx1a-DNA complex stability reside within the precise structural relationships between the homeodomain, hexapeptide and linker regions.

Pbx proteins display hexapeptide-dependent cooperative DNA binding with a subset of Hox proteins.

The human proto-oncogene PBX1 codes for a homolog of Drosophila extradenticle, a divergent homeo domain protein that modulates the developmental and DNA-binding specificity of select HOM proteins. We demonstrate that wild-type Pbx proteins and chimeric E2a-Pbx1 oncoproteins cooperatively bind a consensus DNA probe with HoxB4, B6, and B7 of the Antennapedia class of Hox/HOM proteins. Specificity of Hox-Pbx interactions was suggested by the inability of Pbx proteins to cooperatively bind the synthetic DNA target with HoxA10 or Drosophila even-skipped. Site-directed mutagenesis showed that the hexapeptide motif (IYPWMK) upstream of the Hox homeo domain was essential for HoxB6 and B7 to cooperatively bind DNA with Pbx proteins. Engraftment of the HoxB7 hexapeptide onto HoxA10 endowed it with robust cooperative properties, demonstrating a functional role for the highly conserved hexapeptide element as one of the molecular determinants delimiting Hox-Pbx cooperativity. The Pbx homeo domain was necessary but not sufficient for cooperativity, which required conserved amino acids carboxy-terminal of the homeo domain. These findings demonstrate that interactions between Hox and Pbx proteins modulate their DNA-binding properties, suggesting that Pbx and Hox proteins act in parallel as heterotypic complexes to regulate expression of specific subordinate genes.

The distribution of chondroitin sulfates in articular and growth cartilages of human bones.

The relative contents of chondroitin 4- and 6-sulfates in cartilages of different human bones are reported. Articular and vertebral body cartilages contain almost exclusively chondroitin 6-sulfate, whereas growth and subarticular cartilages contain nearly equal amounts of chondroitin 4-sulfate and chondroitin 6-sulfate. Adult cartilages, where the calcification process is complete, contain only chondroitin 6-sulfate. These results that chondroitin 4-sulfate may be an important component for the calcification process, whereas chondroitin 6-sulfate seems to be related to the integrity of the articular surfaces. A chemical defect of chondroitin 6-sulfate in a new mucopolysaccharidosis, characterized by platyspondyly and irregularities of articular surfaces, is in agreement with these results.