Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 8 de 8
Filtrar
1.
Mol Cell Biol ; 15(1): 235-45, 1995 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-7799930

RESUMO

Human insulin-like growth factor II (IGF-II) mRNAs are subject to site-specific endonucleolytic cleavage in the 3' untranslated region, leading to an unstable 5' cleavage product containing the IGF-II coding region and a very stable 3' cleavage product of 1.8 kb. This endonucleolytic cleavage is most probably the first and rate-limiting step in degradation of IGF-II mRNAs. Two sequence elements within the 3' untranslated region are required for cleavage: element I, located approximately 2 kb upstream of the cleavage site, and element II, encompassing the cleavage site itself. We have identified a stable double-stranded RNA stem structure (delta G = -100 kcal/mol [418.4 kJ/mol]) that can be formed between element I and a region downstream of the cleavage site in element II. This structure is conserved among human, rat, and mouse mRNAs. Detailed analysis of the requirements for cleavage shows that the relative position of the elements is not essential for cleavage. Furthermore, the distance between the coding region and the cleavage site does not affect the cleavage reaction. Mutational analysis of the long-range RNA-RNA interaction shows that not only the double-stranded character but also the sequence of the stable RNA stem is important for cleavage.


Assuntos
Endonucleases/metabolismo , Fator de Crescimento Insulin-Like II/genética , RNA Mensageiro/metabolismo , Animais , Sequência de Bases , Regulação da Expressão Gênica , Humanos , Camundongos , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Processamento Pós-Transcricional do RNA , RNA Mensageiro/química , Ratos , Relação Estrutura-Atividade
2.
Biochim Biophys Acta ; 873(3): 367-71, 1986 Oct 17.
Artigo em Inglês | MEDLINE | ID: mdl-3756185

RESUMO

Kinetic constants for the transesterification of eight dinucleoside phosphates CpX and UpX by bovine and turtle pancreatic ribonuclease were determined. Both ribonucleases have a preference for purine nucleotides at the position X. However, bovine ribonuclease, like other mammalian ribonucleases, prefers 6-amino bases at this site, while turtle ribonuclease prefers 6-keto bases. This difference in specificity at the B2 site may be explained by the substitution of glutamic acid at position 111 by valine in turtle ribonuclease. These results have been confirmed by inhibition studies with the four nucleoside triphosphates. Inhibition studies with pT and pTp showed that a cationic binding group (P0) for the 5'-phosphate of the pyrimidine nucleotides bound at the primary B1 site is present in turtle ribonuclease, although lysine at position 66 in bovine ribonuclease is absent in turtle ribonuclease. However, the side chain of lysine 122 in turtle ribonuclease is probably located in the correct position to take over the role as cationic P0 site.


Assuntos
Pâncreas/enzimologia , Ribonucleases/metabolismo , Animais , Sequência de Bases , Sítios de Ligação , Bovinos , Esterificação , Cinética , Modelos Moleculares , Ribonuclease Pancreático/metabolismo , Especificidade da Espécie , Especificidade por Substrato , Tartarugas
3.
J Endocrinol ; 144(3): 491-502, 1995 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-7537791

RESUMO

In order to determine the effects of IGF-II overexpression on growth of mice, transgenic mice were produced carrying one of three different H-2Kb human IGF-II minigenes in which different non-coding exons (exon 5, truncated exon 5 or exon 6) preceded the coding exons 7, 8 and 9. These were spaced by truncated introns and for proper polyadenylation an SV40 polyadenylation signal was incorporated. The highest levels of IGF-II minigene mRNA expression were found in lines containing the truncated exon 5 construct (II5'). Those containing exon 6 (II6) had less expression and 5 constructs (II5) gave only moderate levels of mRNA expression. In general mRNA expression was highest in thymus and spleen, low in liver and kidney and absent in the brain. In addition, one II5' line showed expression in the brain. Serum IGF-II levels at 8 weeks of age were increased 7- to 8-fold in homozygous transgenic lines with construct II5' without brain expression and 2- to 3-fold in the one that showed expression in the brain; serum IGF-I levels were unchanged. Serum IGFs in the lines containing the constructs II5 and II6 were not different from those of the controls. In all cases body length and weight as well as the weight of several organs such as brain, liver, kidneys, heart and spleen when expressed as a function of age did not differ from controls. Only the thymus showed a significant increase in weight in the transgenics II5'. Inbreeding of 2 lines containing construct II5' with pituitary deficient Snell dwarf mice did not influence body length or weight despite increased serum IGF-II levels. Again the thymus showed a marked increase in growth. The biological activity of the IGF-II peptide was further demonstrated by increased serum IGF-binding protein-3 in the transgenic dwarf mice, as shown by Western ligand blotting. In summary, overexpression of IGF-II in transgenic normal and dwarf mice does not affect overall body growth, but causes increased growth of the thymus. This suggests a role for IGF-II in thymic development by paracrine/autocrine action.


Assuntos
Fator de Crescimento Insulin-Like II/metabolismo , Camundongos Transgênicos/metabolismo , Timo/crescimento & desenvolvimento , Animais , Sequência de Bases , Northern Blotting , Proteínas de Transporte/metabolismo , Expressão Gênica , Engenharia Genética , Inibidores do Crescimento/metabolismo , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina , Fator de Crescimento Insulin-Like II/genética , Camundongos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Somatomedinas/metabolismo
4.
J Pharm Sci ; 86(3): 335-9, 1997 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-9050802

RESUMO

Complexation of carbamazepine with 2-hydroxypropyl-beta-cyclodextrin was performed to provide improved formulations of this widely used antiepileptic drug. Based on this approach, liquid dosage forms were configured for both parenteral and oral use. Intravenous administration of an aqueous carbamazepine x 2-hydroxypropyl-beta-cyclodextrin (CBZ x HPbetaCD) complex at a CBZ dose of 20 mg/kg was well tolerated and generated high initial drug levels that fell monoexponentially as a function of time, yielding a plasma elimination half-life of 38 min. Oral studies were completed with three preparations: a commercially available tablet and suspension, as well as a CBZ x HPbetaCD oral solution. Oral administration of tablets gave erratic and slow absorption, leading to maximum CBZ concentrations (C(max)) of <2 microg/mL, which were manifested only at 2.5 h after drug dosing. The absolute bioavailability of CBZ from the tablets was approximately 25%. Both the suspension and CBZ x HPbetaCD solution gave a significantly improved profile. Thus, the liquid oral dosage forms approximately doubled the oral bioavailability of CBZ compared with the tablets.


Assuntos
Anticonvulsivantes/farmacocinética , Carbamazepina/farmacocinética , Ciclodextrinas , beta-Ciclodextrinas , 2-Hidroxipropil-beta-Ciclodextrina , Administração Oral , Animais , Anticonvulsivantes/administração & dosagem , Disponibilidade Biológica , Carbamazepina/administração & dosagem , Estudos Cross-Over , Cães , Infusões Intravenosas , Suspensões , Comprimidos
6.
Biochem Biophys Res Commun ; 179(3): 1509-16, 1991 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-1656956

RESUMO

Expression of the human insulin-like growth factor II (IGF-II) gene gives rise to a family of mRNAs which are expressed in a tissue- and developmental-specific manner. Recently, an IGF-II RNA species of 1.8 kb was detected which consists of the 3' terminal untranslated region of the last IGF-II exon. We have investigated the mechanism of formation of this RNA employing IGF-II minigenes. Analysis of the expression of these genes has revealed that the 1.8-kb RNA does not result from transcription activation, but arises by specific cleavage of IGF-II transcripts.


Assuntos
Endonucleases/metabolismo , Fator de Crescimento Insulin-Like II/genética , RNA Mensageiro/metabolismo , Animais , Sequência de Bases , Northern Blotting , Calorimetria , Linhagem Celular , Clonagem Molecular , Éxons , Genes , Humanos , Insulina/genética , Camundongos , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Plasmídeos , RNA Mensageiro/genética , Ratos , Homologia de Sequência do Ácido Nucleico , Transfecção
7.
Nucleic Acids Res ; 20(19): 5003-9, 1992 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-1408818

RESUMO

The human insulin-like growth factor II (IGF-II) gene constitutes a complex transcriptional unit that contains nine exons and four promoters. Expression of the IGF-II gene yields a family of mRNAs that all encode prepro-IGF-II. In addition, a stable 1.8 kb RNA is formed that is derived from the 3' untranslated region of exon 9. Recently, we have shown that this RNA species arises by site-specific endonucleolytic cleavage of IGF-II mRNAs and not by transcription from a separate promoter. In the present study we establish that two widely separated sequence elements of approximately 300 nucleotides, both located within exon 9, are required for this cleavage reaction. The first element encompasses about 200 nucleotides upstream and 100 nucleotides downstream of the cleavage site, while the second element is located within a region of 330 nucleotides about 2 kb upstream of the cleavage site. Interestingly, site-specific cleavage also occurred when a fragment from exon 9 of the IGF-II gene containing these two elements was inserted into the 3' untranslated part of the beta-globin gene. Apparently, the expressed hybrid beta-globin-IGF-II mRNA contains all the regulatory elements to confer site-specific endonucleolytic cleavage.


Assuntos
Processamento Alternativo , Éxons , Fator de Crescimento Insulin-Like II/genética , Regiões Promotoras Genéticas , RNA Mensageiro/metabolismo , Sequência de Bases , Linhagem Celular , Clonagem Molecular , Globinas/biossíntese , Globinas/genética , Humanos , Fator de Crescimento Insulin-Like II/biossíntese , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Plasmídeos , RNA Mensageiro/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Mapeamento por Restrição , Transfecção
8.
Antimicrob Agents Chemother ; 41(1): 122-8, 1997 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-8980767

RESUMO

AIDS encephalopathy is an insidious complication of human immunodeficiency virus infection which is difficult to treat because of the poor uptake of many potentially useful antiretroviral drugs through the blood-brain barrier. A chemical delivery system (CDS) for zidovudine (AZT) based on redox trapping within the brain has been prepared and tested in several animal models to circumvent this limitation. The behavior of the AZT-CDS in the dog was considered. Parenteral administration of AZT resulted in rapid systemic elimination and poor uptake by the central nervous system. Ratios of the area under the concentration-time curve of AZT for cerebrospinal fluid to that for blood were 0.32, and ratios of the area under the concentration-time curve of AZT for brain to that for blood were approximately 0.25. Administration of an aqueous formulation of the AZT-CDS resulted in rapid tissue uptake and conversion of the CDS to the corresponding quaternary salt with the subsequent production of AZT. Delivered in this way, the levels of AZT in brain were 1.75- to 3.3-fold higher than those associated with conventional AZT administration. In addition, the levels of AZT in blood were 46% lower than those associated with AZT administration. The higher concentrations in brain and lower concentration in blood combined to significantly increase the ratio of the concentration of AZT in the brain to that in blood after AZT-CDS administration compared to that after AZT dosing.


Assuntos
Fármacos Anti-HIV/administração & dosagem , Química Encefálica/efeitos dos fármacos , Sistemas de Liberação de Medicamentos , Zidovudina/administração & dosagem , Animais , Fármacos Anti-HIV/sangue , Fármacos Anti-HIV/líquido cefalorraquidiano , Fármacos Anti-HIV/farmacocinética , Cerebelo/química , Cerebelo/efeitos dos fármacos , Córtex Cerebral/química , Córtex Cerebral/efeitos dos fármacos , Estudos Cross-Over , Cães , Relação Dose-Resposta a Droga , Distribuição Tecidual , Zidovudina/líquido cefalorraquidiano , Zidovudina/farmacocinética
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA