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1.
Annu Rev Genet ; 48: 537-59, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25292357

RESUMO

mRNA degradation is an important mechanism for controlling gene expression in bacterial cells. This process involves the orderly action of a battery of cellular endonucleases and exonucleases, some universal and others present only in certain species. These ribonucleases function with the assistance of ancillary enzymes that covalently modify the 5' or 3' end of RNA or unwind base-paired regions. Triggered by initiating events at either the 5' terminus or an internal site, mRNA decay occurs at diverse rates that are transcript specific and governed by RNA sequence and structure, translating ribosomes, and bound sRNAs or proteins. In response to environmental cues, bacteria are able to orchestrate widespread changes in mRNA lifetimes by modulating the concentration or specific activity of cellular ribonucleases or by unmasking the mRNA-degrading activity of cellular toxins.


Assuntos
Escherichia coli/genética , Estabilidade de RNA/genética , Ribonucleases/genética , Ribossomos/genética , Endorribonucleases/genética , Regulação Bacteriana da Expressão Gênica , Complexos Multienzimáticos/genética , Filogenia , Polirribonucleotídeo Nucleotidiltransferase/genética , RNA Helicases/genética , RNA Mensageiro/genética
2.
J Biol Chem ; 292(5): 1934-1950, 2017 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-27974459

RESUMO

RNA degradation is crucial for regulating gene expression in all organisms. Like the decapping of eukaryotic mRNAs, the conversion of the 5'-terminal triphosphate of bacterial transcripts to a monophosphate can trigger RNA decay by exposing the transcript to attack by 5'-monophosphate-dependent ribonucleases. In both biological realms, this deprotection step is catalyzed by members of the Nudix hydrolase family. The genome of the gastric pathogen Helicobacter pylori, a Gram-negative epsilonproteobacterium, encodes two proteins resembling Nudix enzymes. Here we present evidence that one of them, HP1228 (renamed HpRppH), is an RNA pyrophosphohydrolase that triggers RNA degradation in H. pylori, whereas the other, HP0507, lacks such activity. In vitro, HpRppH converts RNA 5'-triphosphates and diphosphates to monophosphates. It requires at least two unpaired nucleotides at the 5' end of its substrates and prefers three or more but has only modest sequence preferences. The influence of HpRppH on RNA degradation in vivo was examined by using RNA-seq to search the H. pylori transcriptome for RNAs whose 5'-phosphorylation state and cellular concentration are governed by this enzyme. Analysis of cDNA libraries specific for transcripts bearing a 5'-triphosphate and/or monophosphate revealed at least 63 potential HpRppH targets. These included mRNAs and sRNAs, several of which were validated individually by half-life measurements and quantification of their 5'-terminal phosphorylation state in wild-type and mutant cells. These findings demonstrate an important role for RppH in post-transcriptional gene regulation in pathogenic Epsilonproteobacteria and suggest a possible basis for the phenotypes of H. pylori mutants lacking this enzyme.


Assuntos
Hidrolases Anidrido Ácido/metabolismo , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica/fisiologia , Helicobacter pylori/metabolismo , Estabilidade de RNA/fisiologia , RNA Bacteriano/metabolismo , RNA Mensageiro/metabolismo , Hidrolases Anidrido Ácido/genética , Proteínas de Bactérias/genética , Helicobacter pylori/genética , Helicobacter pylori/patogenicidade , RNA Bacteriano/genética
3.
Nucleic Acids Res ; 43(1): 309-23, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25429978

RESUMO

In metazoans, cleavage by the endoribonuclease SMG6 is often the first degradative event in non-sense-mediated mRNA decay (NMD). However, the exact sites of SMG6 cleavage have yet to be determined for any endogenous targets, and most evidence as to the identity of SMG6 substrates is indirect. Here, we use Parallel Analysis of RNA Ends to specifically identify the 5' termini of decay intermediates whose production is dependent on SMG6 and the universal NMD factor UPF1. In this manner, the SMG6 cleavage sites in hundreds of endogenous NMD targets in human cells have been mapped at high resolution. In addition, a preferred sequence motif spanning most SMG6 cleavage sites has been discovered and validated by mutational analysis. For many SMG6 substrates, depletion of SMG6 resulted in the accumulation of decapped transcripts, an effect indicative of competition between SMG6-dependent and SMG6-independent NMD pathways. These findings provide key insights into the mechanisms by which mRNAs targeted by NMD are degraded.


Assuntos
Degradação do RNAm Mediada por Códon sem Sentido , Clivagem do RNA , RNA Mensageiro/química , Telomerase/metabolismo , Células HeLa , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Motivos de Nucleotídeos , RNA Mensageiro/metabolismo , Análise de Sequência de RNA
4.
J Biol Chem ; 290(15): 9478-86, 2015 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-25657006

RESUMO

Bacterial RNA degradation often begins with conversion of the 5'-terminal triphosphate to a monophosphate by the RNA pyrophosphohydrolase RppH, an event that triggers rapid ribonucleolytic attack. Besides its role as the master regulator of 5'-end-dependent mRNA decay, RppH is important for the ability of pathogenic bacteria to invade host cells, yet little is known about how it chooses its targets. Here, we show that Escherichia coli RppH (EcRppH) requires at least two unpaired nucleotides at the RNA 5' end and prefers three or more such nucleotides. It can tolerate any nucleotide at the first three positions but has a modest preference for A at the 5' terminus and either a G or A at the second position. Mutational analysis has identified EcRppH residues crucial for substrate recognition or catalysis. The promiscuity of EcRppH differentiates it from its Bacillus subtilis counterpart, which has a strict RNA sequence requirement. EcRppH orthologs likely to share its relaxed sequence specificity are widespread in all classes of Proteobacteria, except Deltaproteobacteria, and in flowering plants. By contrast, the phylogenetic range of recognizable B. subtilis RppH orthologs appears to be restricted to the order Bacillales. These findings help to explain the selective influence of RppH on bacterial mRNA decay and show that RppH-dependent degradation has diversified significantly during the course of evolution.


Assuntos
Hidrolases Anidrido Ácido/genética , Proteínas de Escherichia coli/genética , Evolução Molecular , RNA Bacteriano/genética , Hidrolases Anidrido Ácido/classificação , Hidrolases Anidrido Ácido/metabolismo , Sequência de Aminoácidos , Bacillus subtilis/enzimologia , Bacillus subtilis/genética , Sequência de Bases , Sítios de Ligação/genética , Biocatálise , Escherichia coli/enzimologia , Escherichia coli/genética , Proteínas de Escherichia coli/classificação , Proteínas de Escherichia coli/metabolismo , Dados de Sequência Molecular , Mutação , Nucleotídeos/genética , Nucleotídeos/metabolismo , Filogenia , Estabilidade de RNA/genética , RNA Bacteriano/metabolismo , Homologia de Sequência de Aminoácidos , Especificidade da Espécie , Especificidade por Substrato
5.
J Am Assoc Lab Anim Sci ; 62(1): 64-73, 2023 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-36755202

RESUMO

Tens of thousands of rodents are used each year in Rodent Health Monitoring programs. However, Environment Health Monitoring (EHM) could replace sentinel rodent use while maintaining or even improving diagnostic quality. Despite its advantages, widespread implementation of EHM appears to be relatively low. To better understand EHM's prevalence and factors influencing its use, we surveyed research animal professionals. Our hypotheses were (1) EHM prevalence would be low and (2) EHM use would be associated with beliefs and knowledge about EHM. Participants were recruited via online promotion. A total of 158 individuals completed a mixed-methods survey about current practices, beliefs, and knowledge about EHM. Qualitative data were coded using thematic analysis and analyzed using generalized linear models. Results showed that current EHM implementation was low; only 11% of institutions used EHM exclusively. Across the 111 institutions surveyed, over 20,000 soiled bedding sentinels were used each year. However, most participants believed EHM to be advantageous in replacing sentinel animals (78% of participants). Some participants believed EHM could save time (31%), cost less (27%), and be highly accurate (15%). Conversely, some participants believed EHM would be difficult to use due to their current caging type (40%), higher costs (21%), lower accuracy (16%), and personnel attitudes/expertise (14%). Overall, respondents with higher planned EHM use also had more positive attitudes, norms, and control of EHM. We also identified several factors that could promote the implementation of EHM. Communication efforts should emphasize that EHM is compatible with various types of caging, can provide cost savings, has high accuracy, and is consistent with the 3Rs as a replacement. Efforts should also focus on improving attitudes, encouraging peers, and providing resources to facilitate implementation. Implementation in just the surveyed institutions could eliminate the need for well over 20,000 rodents each year, consistent with 3Rs goals.


Assuntos
Benchmarking , Roedores , Animais , Estudos Transversais , Atitude , Monitoramento Ambiental
6.
J Bacteriol ; 194(22): 6233-9, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22984254

RESUMO

Many Escherichia coli mRNAs are degraded by a 5'-end-dependent mechanism in which RppH-catalyzed conversion of the 5'-terminal triphosphate to a monophosphate triggers rapid endonucleolytic cleavage by RNase E. However, little is understood about what governs the decay rates of these transcripts. We investigated the decay of three such messages--rpsT P1, yfcZ, and ydfG--to characterize the rate-determining step in their degradation. The steady-state ratio of monophosphorylated to triphosphorylated rpsT P1 and yfcZ mRNA indicates that their decay rate is limited by cleavage of the monophosphorylated intermediate, making RNase E critical for their rapid turnover. Conversely, the decay rate of ydfG is limited by generation of the monophosphorylated intermediate; therefore, either RNase E or its less abundant paralog RNase G is sufficient for rapid ydfG degradation. Although all three transcripts are stabilized when RppH is absent, overproducing RppH does not accelerate their decay, nor does RppH overproduction appear to influence the longevity of most other messages that it targets. The failure of excess RppH to hasten rpsT P1 and yfcZ degradation despite increasing the percentage of each that is monophosphorylated is consistent with the observation that pyrophosphate removal is not the rate-limiting step in their decay. In contrast, neither the ydfG decay rate nor the fraction of ydfG transcripts that are monophosphorylated increases when the cellular concentration of RppH is raised, suggesting that, for some RppH targets, the rate of formation of the monophosphorylated intermediate is limited by an ancillary factor or by a step that precedes pyrophosphate removal.


Assuntos
Escherichia coli/metabolismo , Regulação Bacteriana da Expressão Gênica/fisiologia , RNA Bacteriano/metabolismo , Hidrolases Anidrido Ácido/genética , Hidrolases Anidrido Ácido/metabolismo , Endorribonucleases/genética , Endorribonucleases/metabolismo , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Regulação Enzimológica da Expressão Gênica , Immunoblotting , Fosforilação , Plasmídeos , RNA Bacteriano/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
7.
BMC Gastroenterol ; 11: 76, 2011 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-21689439

RESUMO

BACKGROUND: Fully covered Self-Expanding metal stents (FCSEMS) have been shown efficacious in palliating malignant biliary obstructions. There is little data analyzing mucosal response to their temporary placement in the bile duct. METHODS: Ten mini pigs underwent endoscopic placement of a FCSEMS (Wallflex, Boston Scientific). FCSEMS were kept in place for three months. At the end of the 3 months, FCSEMS were removed endoscopically. Five pigs were euthanized and their bile ducts harvested. The other five were kept alive for another month post removal. A single pathologist, created a scoring system (to determine degree of inflammation, fibrosis, and epithelial injury), examined all specimens in a blinded fashion. RESULTS: Four FCSEMS spontaneously migrated in the duodenum. On post mortem examination, mild mucosal thickness was noted in three bile duct specimens while superficial inflammation of the bile duct was noted in five animals. Histologic examination of the bile duct revealed focal acute inflammation in both groups. For the 5 animals euthanized immediately after stent removal, there was a tendency to have superficial mucosal erosion and fibrosis. In contrast, increased chronic inflammation was more commonly seen in the animals 1 month post stent removal, with all animals in this group showing moderate degrees of mononuclear inflammatory cell mucosal infiltrates. No severe inflammatory or fibrotic duct injury was observed in any of the study animals, with degree of injury graded as mild to moderate. CONCLUSION: FCSEMS appear to induce minimal tissue overgrowth or fibrosis post placement. Ease of removability and no significant histologic injury are advantages noted with FCSEMS., however, further studies are needed to evaluate treating benign biliary strictures with FCSEMS in humans.


Assuntos
Ductos Biliares Extra-Hepáticos/cirurgia , Colangite/patologia , Materiais Revestidos Biocompatíveis , Duodenopatias/patologia , Migração de Corpo Estranho/patologia , Stents/efeitos adversos , Animais , Ductos Biliares Extra-Hepáticos/patologia , Colangiopancreatografia Retrógrada Endoscópica , Colangite/etiologia , Doença Crônica , Duodenopatias/etiologia , Feminino , Fibrose/etiologia , Seguimentos , Migração de Corpo Estranho/etiologia , Metais , Modelos Animais , Mucosa/patologia , Suínos , Porco Miniatura
8.
One Health ; 12: 100231, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33728370

RESUMO

Lessons learned from recent pandemics, such as SARS-CoV-2 have illustrated that education and training in a One Health approach, which recognizes the interdependency of the health of people, animals and the environment, are essential in improving preparations for and responses to disease outbreaks. For this reason and others, there is a critical need to provide One Health (OH) training to medical professionals early in their careers. 133 U.S. medical schools were surveyed for the incorporation of OH learning activities. Results showed that 56% of surveyed programs included OH-related subject matter, primarily in the context of preclinical classroom learning. This supports previous findings that OH education efforts in medical schools lag behind veterinary schools, with many veterinary schools already including OH as a central part of their curricula. A two week OH elective course for third year medical students was developed and implemented at Georgetown University School of Medicine. Topics such as emerging infectious diseases, zoonoses, vector-borne diseases, epidemiology, emergency preparedness, the human-animal bond, and effects of climate change on public health were discussed. The 21 participants were surveyed before and after the course regarding their knowledge and understanding of OH. Participation in the course enhanced the students' knowledge of OH and furthermore, the students' perception of the importance of incorporating OH within the curriculum and in their future careers changed significantly. This study provides clear evidence that successful integration of OH material is achievable at low cost through interdepartmental and interdisciplinary collaboration. A more holistic approach to health care that takes into consideration environmental, wildlife, and domestic animal factors, and introduction of concepts such as OH into the medical school curriculum, can help close the educational gaps identified in the surveys.

9.
Behav Brain Res ; 408: 113288, 2021 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-33836170

RESUMO

Increased perineuronal net (PNN) deposition has been observed in association with corticosteroid administration and stress in rodent models of depression. PNNs are a specialized form of extracellular matrix (ECM) that may enhance GABA-mediated inhibitory neurotransmission to potentially restrict the excitation and plasticity of pyramidal glutamatergic neurons. In contrast, antidepressant administration increases levels of the PNN-degrading enzyme matrix metalloproteinase-9 (MMP-9), which enhances glutamatergic plasticity and neurotransmission. In the present study, we compare pro-MMP-9 levels and measures of stress in females from two mouse strains, C57BL/6 J and BALB/cJ, in the presence or absence of tail grasping versus tunnel-associated cage transfers. Prior work suggests that C57BL/6 J mice show relatively enhanced neuroplasticity and stress resilience, while BALB/c mice demonstrate enhanced susceptibility to adverse effects of stress. Herein we observe that as compared to the C57BL/6 J strain, BALB/c mice demonstrate a higher level of baseline anxiety as determined by elevated plus maze (EPM) testing. Moreover, as determined by open field testing, anxiety is differentially reduced in BALB/c mice by a choice-driven tunnel-entry cage transfer technique. Additionally, as compared to tail-handled C57BL/6 J mice, tail-handled BALB/c mice have reduced brain levels of pro-MMP-9 and increased levels of its endogenous inhibitor, tissue inhibitor of metalloproteinase-1 (TIMP-1); however, tunnel-associated cage transfer increases pro-MMP-9 levels in BALB/c mice. BALB/c mice also show increases in Western blot immunoreactive bands for brevican, a constituent of PNNs. Together, these data support the possibility that MMP-9, an effector of PNN remodeling, contributes to the phenotype of strain and handling-associated differences in behavior.


Assuntos
Comportamento Animal/fisiologia , Manobra Psicológica , Metaloproteinase 9 da Matriz/metabolismo , Resiliência Psicológica , Estresse Psicológico/metabolismo , Estresse Psicológico/fisiopatologia , Animais , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL
10.
Biochem Biophys Res Commun ; 395(1): 42-7, 2010 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-20346349

RESUMO

The shape and compaction of the bacterial nucleoid may affect the accessibility of genetic material to the transcriptional machinery in natural and synthetic systems. To investigate this phenomenon, the nature and contribution of RNA and protein to the compaction of nucleoids that had been gently released from Escherichia coli cells were investigated using fluorescent and transmission electron microscopy. We propose that the removal of RNA from the bacterial nucleoid affects nucleoid compaction by altering the branching density and molecular weight of the nucleoid. We show that a common detergent in nucleoid preparations, Brij 58, plays a previously unrecognized role as a macromolecular crowding agent. RNA-free nucleoids adopt a compact structure similar in size to exponential-phase nucleoids when the concentration of Brij 58 is increased, consistent with our hypothesis. We present evidence that control and protein-free nucleoids behave similarly in solutions containing a macromolecular crowding agent. These results show that the contribution to DNA compaction by nucleoid-associated proteins is small when compared to macromolecular crowding effects.


Assuntos
Núcleo Celular/química , Cetomacrogol/química , Proteínas de Escherichia coli/química , Escherichia coli/química , Ribonucleases/química , Fracionamento Celular , DNA Bacteriano/química , Escherichia coli/ultraestrutura , RNA Bacteriano/química
11.
J Immunol ; 181(2): 1357-64, 2008 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-18606690

RESUMO

IL-23 and IL-17A regulate granulopoiesis through G-CSF, the main granulopoietic cytokine. IL-23 is secreted by activated macrophages and dendritic cells and promotes the expansion of three subsets of IL-17A-expressing neutrophil-regulatory T (Tn) cells; CD4(-)CD8(-)alphabeta(low), CD4(+)CD8(-)alphabeta(+) (Th17), and gammadelta(+) T cells. In this study, we investigate the effects of IL-17A on circulating neutrophil levels using IL-17R-deficient (Il17ra(-/-)) mice and Il17ra(-/-)Itgb2(-/-) mice that lack both IL-17R and all four beta(2) integrins. IL-17R deficiency conferred a reduction in neutrophil numbers and G-CSF levels, as did Ab blockade against IL-17A in wild-type mice. Bone marrow transplantation revealed that IL-17R expression on nonhemopoietic cells had the greatest effects on regulating blood neutrophil counts. Although circulating neutrophil numbers were reduced, IL-17A expression, secretion, and the number of IL-17A-producing Tn cells were elevated in Il17ra(-/-) and Il17ra(-/-)Itgb2(-/-) mice, suggesting a negative feedback effect through IL-17R. The negative regulation of IL-17A-producing T cells and IL-17A and IL-17F gene expression through the interactions of IL-17A or IL-17F with IL-17R was confirmed in splenocyte cultures in vitro. We conclude that IL-17A regulates blood neutrophil counts by inducing G-CSF production mainly in nonhemopoietic cells. IL-17A controls the expansion of IL-17A-producing Tn cell populations through IL-17R.


Assuntos
Fator Estimulador de Colônias de Granulócitos/metabolismo , Interleucina-17/metabolismo , Interleucina-23/metabolismo , Neutrófilos/fisiologia , Receptores de Interleucina-17/metabolismo , Subpopulações de Linfócitos T/imunologia , Animais , Polaridade Celular , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Fator Estimulador de Colônias de Granulócitos/imunologia , Interleucina-17/imunologia , Interleucina-23/imunologia , Contagem de Leucócitos , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/imunologia , Receptores de Interleucina-17/deficiência , Receptores de Interleucina-17/imunologia , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo
12.
Cell Death Differ ; 27(7): 2143-2157, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-31959914

RESUMO

Nonalcoholic fatty liver disease (NAFLD) and its evolution to inflammatory steatohepatitis (NASH) are the most common causes of chronic liver damage and transplantation that are reaching epidemic proportions due to the upraising incidence of metabolic syndrome, obesity, and diabetes. Currently, there is no approved treatment for NASH. The mitochondrial citrate carrier, Slc25a1, has been proposed to play an important role in lipid metabolism, suggesting a potential role for this protein in the pathogenesis of this disease. Here, we show that Slc25a1 inhibition with a specific inhibitor compound, CTPI-2, halts salient alterations of NASH reverting steatosis, preventing the evolution to steatohepatitis, reducing inflammatory macrophage infiltration in the liver and adipose tissue, while starkly mitigating obesity induced by a high-fat diet. These effects are differentially recapitulated by a global ablation of one copy of the Slc25a1 gene or by a liver-targeted Slc25a1 knockout, which unravel dose-dependent and tissue-specific functions of this protein. Mechanistically, through citrate-dependent activities, Slc25a1 inhibition rewires the lipogenic program, blunts signaling from peroxisome proliferator-activated receptor gamma, a key regulator of glucose and lipid metabolism, and inhibits the expression of gluconeogenic genes. The combination of these activities leads not only to inhibition of lipid anabolic processes, but also to a normalization of hyperglycemia and glucose intolerance as well. In summary, our data show for the first time that Slc25a1 serves as an important player in the pathogenesis of fatty liver disease and thus, provides a potentially exploitable and novel therapeutic target.


Assuntos
Proteínas de Transporte/antagonistas & inibidores , Intolerância à Glucose/complicações , Inflamação/complicações , Mitocôndrias/metabolismo , Hepatopatia Gordurosa não Alcoólica/complicações , Acetilcoenzima A/metabolismo , Animais , Glicemia/metabolismo , Proteínas de Transporte/metabolismo , Polaridade Celular , Ácido Cítrico/metabolismo , Dieta Hiperlipídica , Modelos Animais de Doenças , Regulação para Baixo , Jejum/sangue , Gluconeogênese , Intolerância à Glucose/sangue , Hepatomegalia/sangue , Hepatomegalia/complicações , Hepatomegalia/diagnóstico por imagem , Humanos , Hiperglicemia/sangue , Hiperglicemia/complicações , Inflamação/sangue , Resistência à Insulina , Interleucina-6/biossíntese , Lipogênese , Fígado/diagnóstico por imagem , Fígado/metabolismo , Fígado/patologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/sangue , Hepatopatia Gordurosa não Alcoólica/diagnóstico por imagem , Obesidade/sangue , Obesidade/complicações , Fenótipo , Fatores de Tempo , Triglicerídeos/metabolismo , Fator de Necrose Tumoral alfa/biossíntese
13.
Am J Med Genet A ; 149A(5): 997-1000, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19353645

RESUMO

Rubinstein-Taybi syndrome (RSTS) is a heterogeneous disorder with approximately 45-55% of patients showing mutations in the CREB binding protein and a further 3% of patients having mutations in EP300. We report a male child with a deletion of exons 3-8 of the EP300 gene who has RSTS. He has a milder skeletal phenotype, a finding that has been described in other cases with EP300 mutations. The mother suffered from pre-eclampsia and HELLP syndrome in the pregnancy. She subsequently developed a mullerian tumor of her cervix 6 years after the birth of her son.


Assuntos
Proteína p300 Associada a E1A/genética , Síndrome de Rubinstein-Taybi/genética , Criança , Cromossomos Humanos Par 22 , Análise Mutacional de DNA , Humanos , Masculino , Deleção de Sequência
14.
ILAR J ; 60(1): 50-57, 2019 12 31.
Artigo em Inglês | MEDLINE | ID: mdl-31361817

RESUMO

There have been recent efforts to reduce the administrative burden imposed on investigators. Although a complete and thorough review of proposed animal studies is an essential function of the Institutional Animal Care and Use Committee (IACUC), efforts to streamline and clarify this process may help investigators spend less time writing animal use protocols and responding to committee comments. The IACUC relies on well-written protocols for an efficient review process. A well-designed protocol form is also critical in guiding investigators through the process. However, it is ultimately the investigators' responsibility to ensure that the information they provide answers all the IACUC's questions with enough detail and quality for a fast and effective review. This article, aimed primarily for researchers but also IACUC administrators, provides an overview of the IACUC protocol review and approval process, the criteria that the IACUC uses for evaluations, and the type of information that should be included in the various sections of the protocol form. Some specific examples are also provided.


Assuntos
Experimentação Animal/normas , Pesquisa Biomédica/métodos , Comitês de Cuidado Animal , Bem-Estar do Animal/normas , Animais , Animais de Laboratório
15.
Comp Med ; 69(6): 468-489, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31822323

RESUMO

The use of effective regimens for mitigating pain remain underutilized in research rodents despite the general acceptance of both the ethical imperative and regulatory requirements intended to maximize animal welfare. Factors contributing to this gap between the need for and the actual use of analgesia include lack of sufficient evidence-based data on effective regimens, under-dosing due to labor required to dose analgesics at appropriate intervals, concerns that the use of analgesics may impact study outcomes, and beliefs that rodents recover quickly from invasive procedures and as such do not need analgesics. Fundamentally, any discussion of clinical management of pain in rodents must recognize that nociceptive pathways and pain signaling mechanisms are highly conserved across mammalian species, and that central processing of pain is largely equivalent in rodents and other larger research species such as dogs, cats, or primates. Other obstacles to effective pain management in rodents have been the lack of objective, science-driven data on pain assessment, and the availability of appropriate pharmacological tools for pain mitigation. To address this deficit, we have reviewed and summarized the available publications on pain management in rats, mice and guinea pigs. Different drug classes and specific pharmacokinetic profiles, recommended dosages, and routes of administration are discussed, and updated recommendations are provided. Nonpharmacologic tools for increasing the comfort and wellbeing of research animals are also discussed. The potential adverse effects of analgesics are also reviewed. While gaps still exist in our understanding of clinical pain management in rodents, effective pharmacologic and nonpharmacologic strategies are available that can and should be used to provide analgesia while minimizing adverse effects. The key to effective clinical management of pain is thoughtful planning that incorporates study needs and veterinary guidance, knowledge of the pharmacokinetics and mechanisms of action of drugs being considered, careful attention to individual differences, and establishing an institutional culture that commits to pain management for all species as a central component of animal welfare.


Assuntos
Experimentação Animal/ética , Cobaias , Camundongos , Manejo da Dor/veterinária , Medição da Dor/veterinária , Ratos , Analgesia/veterinária , Analgésicos/efeitos adversos , Analgésicos/farmacocinética , Analgésicos/farmacologia , Bem-Estar do Animal , Animais , Relação Dose-Resposta a Droga
16.
J Am Assoc Lab Anim Sci ; 57(5): 477-482, 2018 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-30092857

RESUMO

Testing sentinel animals exposed to soiled bedding from colony animals is the most common method used for health monitoring in rodent facilities. Although environmental sampling is being explored-and, in many cases, has been implemented-as an alternative, exhaust plenum sampling is not effective for all ventilated rack designs. This study evaluated PCR testing of filter paper from sentinel cages on ventilated racks. We hypothesized that testing filter paper from cages containing soiled bedding would be as effective as testing sentinel mice and that periodic shaking of cages would generate sufficient particulate movement to substitute for the presence of live animals. Three cages containing soiled bedding were maintained in each of 8 rooms; one cage contained 2 Cr:NIH(S) mice, one had no mice and was shaken twice weekly, and the remaining one had no mice and was left undisturbed. For 3 consecutive months, a piece of filter paper from the undersurface of the cage lid was tested monthly for adventitial agents and then replaced. A second piece remained on the cage undersurface for 3 mo. Fecal pellets and oral and fur swabs were collected from sentinel mice at months 1 and 3 and tested for the same agents. At month 3, serology was performed on the sentinel mice; feces and oral and fur swabs from colony animals were tested concurrently for comparison. Filter paper from cages without mice and shaken were at least as effective than all other methods in detecting the presence of endemic agents, including mouse norovirus, Helicobacter spp., Pasteurella pneumotropica, Entamoeba muris, and Spironucleus muris. For IVC systems where exhaust plenum testing is ineffective, PCR testing of IVC filter tops should be considered as an alternative to soiled bedding sentinels. Environmental sampling may provide increased reliability and reduce the number of rodents used for routine health surveillance.


Assuntos
Bactérias/isolamento & purificação , Filtração/instrumentação , Abrigo para Animais , Reação em Cadeia da Polimerase/veterinária , Animais , Bactérias/genética , Ciência dos Animais de Laboratório , Camundongos , Reprodutibilidade dos Testes
17.
Clin Dysmorphol ; 16(3): 177-179, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17551332

RESUMO

A case of Jacobsen syndrome, suspected antenatally on the grounds of trigonocephaly and hypoplastic left heart syndrome, is presented. Clinicians are reminded that a hypoplastic left heart should not be assumed to be an isolated malformation and that a careful search for associated malformations can facilitate the recognition of an underlying genetic syndrome.


Assuntos
Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/genética , Deleção Cromossômica , Cromossomos Humanos Par 11/genética , Doenças Fetais/diagnóstico , Doenças Fetais/genética , Diagnóstico Pré-Natal , Evolução Fatal , Feminino , Humanos , Monossomia , Gravidez , Síndrome
19.
Sci Rep ; 6: 29475, 2016 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-27378017

RESUMO

The postmenopausal period in women is associated with decreased circulating estrogen levels, which accelerate bone loss and increase the risk of fracture. Here, we gained novel insight into the molecular mechanisms mediating bone loss in ovariectomized (OVX) mice, a model of human menopause, using co-expression network analysis. Specifically, we generated a co-expression network consisting of 53 gene modules using expression profiles from intact and OVX mice from a panel of inbred strains. The expression of four modules was altered by OVX, including module 23 whose expression was decreased by OVX across all strains. Module 23 was enriched for genes involved in the response to oxidative stress, a process known to be involved in OVX-induced bone loss. Additionally, module 23 homologs were co-expressed in human bone marrow. Alpha synuclein (Snca) was one of the most highly connected "hub" genes in module 23. We characterized mice deficient in Snca and observed a 40% reduction in OVX-induced bone loss. Furthermore, protection was associated with the altered expression of specific network modules, including module 23. In summary, the results of this study suggest that Snca regulates bone network homeostasis and ovariectomy-induced bone loss.


Assuntos
Osteoporose Pós-Menopausa/metabolismo , Fraturas por Osteoporose/metabolismo , alfa-Sinucleína/metabolismo , Animais , Medula Óssea/metabolismo , Osso e Ossos/patologia , Estrogênios/deficiência , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Genótipo , Homeostase , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Ovariectomia , Estresse Oxidativo , Pós-Menopausa , Microtomografia por Raio-X
20.
Eur J Hum Genet ; 24(7): 976-84, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-26733289

RESUMO

Classical galactosaemia (OMIM #230400), a rare disorder of carbohydrate metabolism, is caused by a deficient activity of galactose-1-phosphate uridyltransferase (EC 2.7.7.12). The pathophysiology of the long-term complications, mainly cognitive, neurological and female fertility problems remains poorly understood. The lack of validated biomarkers to determine prognosis, monitor disease progression and responses to new therapies, pose a huge challenge. We report the detailed analysis of an automated robotic hydrophilic interaction ultra-performance liquid chromatography N-glycan analytical method of high glycan peak resolution applied to serum IgG. This has revealed specific N-glycan processing defects observed in 40 adult galactosaemia patients (adults and adolescents), in comparison with 81 matched healthy controls. We have identified a significant increase in core fucosylated neutral glycans (P<0.0001) and a significant decrease in core fucosylated (P<0.001), non-fucosylated (P<0.0001) bisected glycans and, of specific note, decreased N-linked mannose-5 glycans (P<0.0001), in galactosaemia patients. We also report the abnormal expression of a number of related relevant N-glycan biosynthesis genes in peripheral blood mononuclear cells from 32 adult galactosaemia patients. We have noted significant dysregulation of two key N-glycan biosynthesis genes: ALG9 upregulated (P<0.001) and MGAT1 downregulated (P<0.01) in galactosaemia patients, which may contribute to its ongoing pathophysiology. Our data suggest that the use of IgG N-glycosylation analysis with matched N-glycan biosynthesis gene profiles may provide useful biomarkers for monitoring response to therapy and interventions. They also indicate potential gene modifying steps in this N-glycan biosynthesis pathway, of relevance to galactosaemia and related N-glycan biosynthesis disorders.


Assuntos
Galactosemias/genética , Imunoglobulina G/metabolismo , Polissacarídeos/biossíntese , Processamento de Proteína Pós-Traducional , Adolescente , Adulto , Biomarcadores/sangue , Estudos de Casos e Controles , Feminino , Galactosemias/sangue , Galactosemias/patologia , Glicosilação , Humanos , Imunoglobulina G/sangue , Masculino , Manosiltransferases/genética , Manosiltransferases/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Monócitos/metabolismo , N-Acetilglucosaminiltransferases/genética , N-Acetilglucosaminiltransferases/metabolismo
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