RESUMO
Statins may confer renal protection in a variety of glomerular diseases, including diabetic nephropathy (DN). However, various glomerular lesions have different etiologies and may have different responses to statins. This study was performed to determine the differential effects of simvastatin (SMV) on glomerular pathology including mesangial expansion and podocyte injury in a mouse model of early stage type 2 diabetes mellitus (DM). Type 2 DM was induced in male C57BL/6 mice by feeding a high fat diet (HF; 45 kcal% fat). After 22 weeks, one group of HF mice was treated with SMV (HF-SMV; 7 mug/day/g BW) and another group was treated with vehicle (HF-vehicle) for 4 weeks via osmotic mini-pump. A third group served as age-matched normal diet vehicle controls (ND-vehicle; 10 kcal% fat). At the end of treatment, glomerular morphology was evaluated in a blind manner to determine the progression of DN. Body weight, blood glucose, insulin, HDL-cholesterol and triglycerides, but not LDL-cholesterol, were increased in HF mice. Over the course of treatment, the 24-hour urinary albumin excretion (UAE) was unchanged in ND-vehicle. HF mice exhibited elevated UAE, which decreased with SMV, but was unchanged with vehicle. The absolute mesangial volume and the relative mesangial volume per glomerular volume increased in HF-vehicle and remained elevated with SMV treatment. The immuno-staining of nephrin, a protein marker of the integrity of podocyte slit diaphragms, was decreased in HF-vehicle; however, the nephrin quantity of the HF-SMV group was not different from ND-vehicle. It is concluded that SMV reverses podocyte damage, but does not affect mesangial expansion in the kidneys of early stage proteinuria of type 2 DM.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Mesângio Glomerular/efeitos dos fármacos , Podócitos/efeitos dos fármacos , Sinvastatina/farmacologia , Análise de Variância , Animais , Glicemia/análise , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 2/patologia , Modelos Animais de Doenças , Mesângio Glomerular/patologia , Imuno-Histoquímica , Testes de Função Renal , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica , Tamanho do Órgão , Podócitos/patologia , Probabilidade , Distribuição Aleatória , Fatores de Risco , Índice de Gravidade de DoençaRESUMO
Pretreatment of rats with insulin-like growth factor I (IGF-I) ameliorates the course of acute ischemic renal injury. Differential display PCR was used to identify genes that are expressed in kidney after induction of acute ischemic renal injury in rats pretreated with vehicle or IGF-I. One amplification product that showed enhanced expression in kidneys of rats rendered ischemic compared to kidneys of sham-operated rats was identified as osteopontin. Sequence analysis of full-length complementary DNAs revealed a single species. Renal tissue was obtained for study 12 h and 1, 2, 3, 5, 7, 14, and 28 days postinjury. Levels of whole kidney osteopontin messenger RNA (mRNA) in rats rendered ischemic 1 day previously were elevated approximately 18-fold compared to levels measured in sham-operated controls, as determined by Northern analysis. No differences were noted 12 h postinjury. Levels of osteopontin mRNA remained elevated for 14 days after ischemia, but were no longer elevated at 28 days. IGF-I pretreatment resulted in enhanced levels of osteopontin mRNA 12 h, 1 day, and 5 days postinjury. In situ hybridization demonstrated that the elevated expression of osteopontin 1 day postinjury was localized predominantly to cells in the distal tubule and medullary thick ascending limb of Henle's loop. Immunostaining showed an identical localization for elevated protein expression. Five days postinjury, osteopontin peptide and mRNA were clearly detected in regenerating proximal tubules in addition to distal tubule and medullary thick ascending limb. We propose that endogenous osteopontin serves to promote tissue regeneration and tissue remodeling within 1 day after acute ischemic injury of kidney. IGF-I enhanced expression of osteopontin at an earlier time postischemia may ameliorate the course of injury.
Assuntos
Expressão Gênica , Fator de Crescimento Insulin-Like I/farmacologia , Isquemia/complicações , Nefropatias/etiologia , Rim/irrigação sanguínea , Rim/metabolismo , Sialoglicoproteínas/genética , Animais , Sequência de Bases , Northern Blotting , Hibridização In Situ , Rim/química , Nefropatias/metabolismo , Nefropatias/terapia , Masculino , Dados de Sequência Molecular , Osteopontina , Reação em Cadeia da Polimerase , RNA Mensageiro/análise , Ratos , Ratos Sprague-Dawley , Distribuição TecidualRESUMO
BACKGROUND: We have previously reported that the expression of the receptor for activated C kinase (RACK1) is induced post-ischemia/preperfusion injury to the kidney, and activation of protein kinase C (PKC) protects renal cells from hypoxic injury. This study was done to determine whether the induced expression of RACK1 is accompanied by changes in the level of expression and subcellular distribution of PKC isozymes. METHODS: Ischemia/reperfusion injury resulting in acute renal failure was induced by 60 minutes of bilateral renal artery clamping in rats. The expression levels and translocation of various PKC isozymes between soluble and particulate fractions in whole kidney homogenates were demonstrated by immunoblot analysis. The expression pattern of the various PKC isozymes in the kidney postinjury was performed by immunohistochemistry. RESULTS: PKC alpha, beta II, and zeta were induced and translocated from the soluble fraction to the particulate fraction post-injury. Immunolocalization showed PKC alpha, beta II, and zeta expression to be induced in the proximal tubule epithelial cell (PTEC) at 0 to 30 minutes post-ischemia/reperfusion injury (IRI). At one-day postinjury, the alpha isozyme was translocated to the plasma membrane of the undamaged PTEC, while it was translocated to the nucleus in damaged PTEC. PKC beta II expression was along the basal and lateral side of the undamaged PTEC, while it was distributed in the cytoplasm of sloughed cells in the damaged PTEC. PKC zeta expression at one day was along the apical side of the damaged PTEC. At seven-days postinjury, the expressions of the alpha and zeta isozymes were localized to the plasma membrane of the regenerating PTEC and the expression of PKC beta II isozyme to certain interstitial cells. CONCLUSION: The induced expression, translocation, and the intracellular spatial distributions of the enzymes suggest that they may mediate multiple processes during IRI.
Assuntos
Isquemia/enzimologia , Rim/irrigação sanguínea , Rim/enzimologia , Proteína Quinase C/biossíntese , Injúria Renal Aguda/enzimologia , Injúria Renal Aguda/patologia , Animais , Indução Enzimática , Isquemia/patologia , Isoenzimas/biossíntese , Isoenzimas/metabolismo , Rim/lesões , Túbulos Renais Proximais/enzimologia , Cinética , Masculino , Peptídeos/metabolismo , Proteína Quinase C/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Quinase C Ativada , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Frações Subcelulares/enzimologiaRESUMO
An AG dinucleotide is an invariant feature of all acceptor splice sites, and deletion or substitution of (one of) these nucleotides will result in abnormal processing of the beta-globin mRNA. Restriction endonuclease mapping of DNA from an American black patient with Hb S-beta zero-thalassemia failed to detect any deletion in the beta 0-globin gene region, but cloning and sequencing of the beta 0-globin gene showed a point mutation (A----C) in the highly conserved dinucleotide AG of the acceptor splice junction of the IVS-2. Blot hybridization analysis of RNA prepared from the erythroid cells of the patient showed only RNA of normal size. The patient and her daughter, who has the same condition, have high levels of Hb F (27%-35%); the mechanism responsible for the greatly increased gamma chain production remains unclear.
Assuntos
DNA/genética , Globinas/genética , Talassemia/genética , Adulto , Sequência de Bases , População Negra , Feminino , Haploidia , Humanos , Pessoa de Meia-Idade , Hibridização de Ácido Nucleico , RNA/genética , Estados UnidosRESUMO
Differential display-polymerase chain reaction (DD-PCR) was used to identify genes that are expressed in kidney following induction of acute ischemic renal injury. The receptor for activated C kinase (RACK1) mRNA expression in kidneys obtained from rats 12 h following ischemia is enhanced twofold compared with sham-operated rats. The maximal enhancement of expression (3.3-fold) is at 7 days following reperfusion. Expression remains elevated at 14 days. RACK1 transcripts and protein are localized to the damaged and regenerating segments of proximal tubules. At 1 day following injury, RACK1 protein is present in the epithelial cells of the damaged S3 segment and in cells sloughed into the tubular lumen. By 5 days following injury, RACK1 protein expression is enhanced in the regenerating cells relining the injured tubules of the S3 segment and in papillary proliferations within regenerating tubules. Increased expression of RACK1 could enhance the activity of PKC and, in so doing, regulate the process of regeneration of the proximal tubule following ischemic renal injury.
Assuntos
Isquemia/metabolismo , Rim/metabolismo , Peptídeos/metabolismo , Circulação Renal , Animais , Masculino , Peptídeos/genética , Reação em Cadeia da Polimerase , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Quinase C Ativada , Fatores de Tempo , Distribuição TecidualRESUMO
We have cloned and sequenced a cDNA encoding a novel zinc finger protein (Fzf-1) containing two tandem repeats of zinc finger motifs of the C2H2 type. The cDNA is 3.0 Kb long and has an open reading frame which codes for a protein of 789 amino acids. The expression pattern of the zinc finger gene was studied in chick embryonic skin and feathers by in situ hybridization. The expression of the gene is found to be temporally and spatially regulated. In stage 38 chick embryos, the transcripts are localized to the epidermis but in 10-day-old embryos, the signal is localized to the forming dermis. In 12-day-old chick, the transcripts are localized to the mesenchymal region of the elongated feather buds. Reverse transcription followed by Polymerase Chain Reaction (RT-PCR) did not detect the transcripts in any other tissues.
Assuntos
Plumas/metabolismo , Genes , Pele/metabolismo , Dedos de Zinco/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Embrião de Galinha , Galinhas , Clonagem Molecular , Primers do DNA/genética , DNA Complementar/genética , Plumas/embriologia , Plumas/crescimento & desenvolvimento , Hibridização In Situ , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Pele/embriologia , Pele/crescimento & desenvolvimentoRESUMO
Acute renal failure continues to have an unacceptably high mortality rate. Ischemic renal injury is the most common cause of acute renal failure. Understanding the molecular mechanisms of cell death and regeneration is important for designing future therapeutic strategies. Recent interest in our laboratory has focused on molecular response after ischemic renal injury and, in particular, genes that are important in cell death and repair after ischemia. The identification of genes that are differentially expressed after ischemia has led to new information regarding the identity of possible mediators of cell death and regeneration in renal tubular epithelial cells.
Assuntos
Regulação da Expressão Gênica , Isquemia , Nefropatias , Regeneração/genética , Animais , Morte Celular/genética , Humanos , Isquemia/genética , Isquemia/patologia , Isquemia/fisiopatologia , Rim/irrigação sanguínea , Rim/patologia , Rim/fisiopatologia , Nefropatias/genética , Nefropatias/patologia , Nefropatias/fisiopatologiaRESUMO
The morphogenetic mechanisms involved in shaping the embyro are largely unknown. Previous studies from this laboratory suggest that the mesonephros promotes limb outgrowth in ovo in the chicken embryo and might be involved in early limb morphogenesis, since damage to the mesonephros results in truncated limbs. In limb bud organ cultures, the presence of the mesonephros promotes cartilage formation. This effect can be reproduced by exogenous IGF-I or prevented by blocking antibody to IGF-I. In order to examine the hypothesis that mesonephros-derived IGF-I is involved in the early morphogenesis of the limb, we examined the spatial and temporal expression of IGF-I and type I receptor for IGF by in situ hybridization at stages when the onset of limb development occurs. The results show that neither transcript is detected at stage 13, prior to the appearance of the limb bud; but both transcripts are detected in the mesonephros at stage 14, an early stage in limb outgrowth. The hybridization signal in the mesonephros for both transcripts increases with development and signal was codistributed as well. At stage 18 the level of receptor transcripts detected in the flank relative to the limb decreased. Thus, the temporal and spatial patterns of expression of IGF-I and its receptor are consistent with their involvement in the initiation of limb outgrowth and support the model that localized expression of a growth factor and its receptor can be involved in shaping the embryo.
Assuntos
Extremidades/embriologia , Fator de Crescimento Insulin-Like I/análise , Mesonefro/química , Receptor IGF Tipo 1/análise , Animais , Embrião de Galinha , Regulação da Expressão Gênica , Hibridização In Situ , MorfogêneseRESUMO
Genes differentially expressed after acute renal ischemic injury were identified using differential display-polymerase chain reaction (DD-PCR). Messenger RNA for calcyclin, a member of the S100 family of calcium-binding proteins, is increased in kidneys by 6 h following ischemic injury to rats compared with sham surgery. The level of calcyclin mRNA is increased 10-fold by 1 day postinjury and declines thereafter. In situ hybridization demonstrates little calcyclin mRNA in kidneys of sham-operated rats. However, calcyclin protein is present in glomeruli and distal tubules (DT). Compared with kidneys from sham-operated controls, both calcyclin mRNA and protein expression are increased at 1-3 days following ischemic injury in the thick ascending limb of Henle, the DT, and in damaged regenerating segments of proximal tubules. By 7 days postischemia there is a reduction in mRNA and protein expression. Calcyclin could play a role in the regulation of renal cell proliferation and regeneration in the recovery process after acute ischemic injury.
Assuntos
Proteínas de Ligação ao Cálcio/biossíntese , Proteínas de Ciclo Celular , Isquemia/metabolismo , Rim/irrigação sanguínea , Proteínas S100 , Transcrição Gênica , Animais , Clonagem Molecular , Fator de Crescimento Epidérmico/biossíntese , Hibridização In Situ , Isquemia/patologia , Rim/metabolismo , Rim/patologia , Masculino , Reação em Cadeia da Polimerase , RNA Mensageiro/biossíntese , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/biossíntese , Proteína A6 Ligante de Cálcio S100RESUMO
Mice deficient for B cell leukemia/lymphoma gene 2 [bcl-2(-/-) mice] manifest congenital renal hypoplasia and develop multicystic kidney disease and renal failure postnatally. To characterize postpartum renal development, to identify the cellular origin of the cysts, and to provide insight into the role that bcl-2 deficiency plays in the cystogenic process, we examined the morphology of kidneys from bcl-2 (-/-) mice and wild-type littermates [bcl-2 (+/+)] from birth (P0) to postpartum day 28 (P28), determined whether abnormalities of cellular proliferation and apoptosis accompany cyst development, and characterized expression of the bcl-2-related protein, bax. Between P0 and P7, kidneys from bcl-2 (-/-) and bcl-2 (+/+) mice undergo a comparable increase in weight and have similar histological appearances. However, during the next 2 wk of life, weight gain in kidneys from bcl-2 (-/-) mice is reduced compared with that in kidneys from bcl-2 (+/+) animals, and cysts develop in tubules with staining characteristics of proximal tubule, distal tubule/medullary thick ascending limb of Henle's loop, and collecting duct. Unaffected glomeruli and proximal tubules in kidneys of bcl-2 (-/-) mice undergo compensatory growth. Cystogenesis is accompanied by enhanced incorporation of 5-bromo-2'-deoxyuridine in cells within cortex and medulla and apoptosis of cells within cysts and in the renal interstitium. Bax protein is expressed in the distal tubule in kidneys of bcl-2 (+/+) and bcl-2 (-/-) mice and in some, but not all cysts. We conclude that abnormal regulation of DNA synthesis and apoptosis accompany cystogenesis in bcl-2 (-/-) mice during postpartum kidney development. Continued expression of bax could enhance apoptotic cell death.
Assuntos
Animais Recém-Nascidos/crescimento & desenvolvimento , Deleção de Genes , Rim/crescimento & desenvolvimento , Doenças Renais Policísticas/genética , Proteínas Proto-Oncogênicas/deficiência , Proteínas Proto-Oncogênicas/genética , Animais , Apoptose , Bromodesoxiuridina/metabolismo , Divisão Celular , Rim/patologia , Camundongos , Doenças Renais Policísticas/metabolismo , Doenças Renais Policísticas/patologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2 , Valores de ReferênciaRESUMO
Restriction endonuclease mapping defined a partial deletion of about 1.35 kb in the beta-globin gene of a black American patient with hemoglobin S-beta zero-thalassemia and in his uncle with a beta zero-thalassemia trait. The 5' endpoint of the deletion is about 600 bases upstream from the cap site, and the 3' endpoint lies within about 500 bases from the 5' splice junction of the second intervening sequence. The deletion is different from that of a previously reported Indian beta zero-thalassemia allele, where 0.6 kb is deleted at the 3' end of the beta-globin gene.
Assuntos
Deleção Cromossômica , Globinas/genética , Talassemia/genética , Adolescente , Adulto , População Negra , Mapeamento Cromossômico , Enzimas de Restrição do DNA/metabolismo , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Studies identifying genes that are differentially expressed following induction of acute ischemic injury have been useful in delineating the pathophysiology of acute renal failure. METHODS: A differential cDNA library screening technique was used to identify genes that are differentially expressed in rat kidney following induction of acute ischemic renal injury. RESULTS: Levels of mRNA with a high homology to that coding for Siva, a human proapoptotic protein, were increased approximately 4.5-fold in kidneys obtained from rats within 12 hours following ischemia, compared to kidneys from sham-operated rats. A partial cDNA sequence for the rat protein (rat Siva) was determined that overlaps 92% of the human open reading frame. The cDNA sequence predicts a protein 177 amino acids in length with 76% homology to human Siva. Levels of rat Siva in kidneys were elevated at one, five and seven days post-ischemia were not different from those in kidneys from sham-operated controls. In situ hybridization demonstrated that rat Siva mRNA was expressed in cells lining damaged sections in the S3 segment of the proximal tubule at 12 hours and one day post-ischemia. At five and seven days, Siva mRNA was located in epithelial cells of regenerating tubules including in papillary proliferations. TdT-mediated dUTP-biotin nick end-labeling (TUNEL)-positive cells colocalized with cells containing Siva mRNA. CD27, the receptor for Siva was localized by immunohistochemistry to sloughed cells in the lumens of damaged S3 segments at 12 hours post-ischemia and to cells within papillary proliferations at five days post-injury. CONCLUSIONS: Siva that is produced within the kidney could be a mediator of apoptosis post-ischemia via an interaction with CD27.
Assuntos
Proteínas de Transporte/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular , Isquemia/metabolismo , Rim/metabolismo , Circulação Renal , Traumatismo por Reperfusão/metabolismo , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Sequência de Aminoácidos/genética , Animais , Proteínas Reguladoras de Apoptose , Sequência de Bases/genética , Proteínas de Transporte/genética , Humanos , Masculino , Dados de Sequência Molecular , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Valores de Referência , Circulação Renal/fisiologiaRESUMO
Here we describe the isolation and characterization of a new chicken homeobox-containing gene, G-Hox 7, which is related to Drosophila msh. The deduced amino acid sequence of the cDNA shows greater than 96% homology to the homeo domain of other vertebrate msh-like genes. As for other species, the amino and carboxy termini of the protein are, however, greatly divergent when compared phylogenetically. In situ hybridization studies revealed the early and wide-spread expression of G-Hox 7 during chick development. This includes its expression in the primitive streak and extraembryonic cells undergoing epiboly, and its expression along the neural axis, including the forebrain. Expression was also observed in the neural crest, neural crest-derived facial and branchial structures, the otocyst, limb, and heart valves. This widespread and recurrent expression of the transcript suggests that the gene may play an essential role at multiple sites during the initiation of new developmental pathways.
Assuntos
Galinhas/genética , Proteínas de Ligação a DNA/genética , Genes Homeobox , Proteínas de Homeodomínio , Sequência de Aminoácidos , Animais , Proteínas Aviárias , Sequência de Bases , Northern Blotting , Embrião de Galinha , Clonagem Molecular , DNA , Drosophila/genética , Olho/embriologia , Olho/metabolismo , Regulação da Expressão Gênica , Coração/embriologia , Fator de Transcrição MSX1 , Dados de Sequência Molecular , Miocárdio/metabolismo , Crista Neural/metabolismo , Hibridização de Ácido Nucleico , Mapeamento por Restrição , Alinhamento de SequênciaRESUMO
A human craniofacial cDNA library was screened with a degenerate oligonucleotide probe based on the conserved third helix of homeobox genes. From this screening, we identified a homeobox gene, H6, which shared only 57-65% amino acid identity to previously reported homeodomains. H6 was physically mapped to the 4p16.1 region by using somatic cell hybrids containing specific deletions of human chromosome 4. Linkage data from a single-stranded conformational polymorphism derived from the 3' untranslated region of the H6 cDNA placed this homeobox gene more than 20 centimorgans proximal of the previously mapped HOX7 gene on chromosome 4. Identity comparisons of the H6 homeodomain with previously reported homeodomains reveal the highest identities to be with the Nk class of homeobox genes in Drosophila melanogaster.
Assuntos
Cromossomos Humanos Par 4 , Genes Homeobox , Sequência de Aminoácidos , Animais , Sequência de Bases , Galinhas , Mapeamento Cromossômico , Sequência Consenso , Ligação Genética , Humanos , Camundongos , Dados de Sequência Molecular , Oligodesoxirribonucleotídeos/química , Linhagem , Reação em Cadeia da Polimerase , Polimorfismo Genético , Alinhamento de SequênciaRESUMO
AIMS/HYPOTHESIS: Type 2 diabetes often results in diabetic nephropathy, which is preceded by an elevated glomerular filtration rate (GFR). This study was designed to develop a mouse model of Type 2 diabetes and to elucidate the glomerular events in the early stages of diabetic nephropathy. METHODS: Four-week-old mice were fed a normal or high-fat (42% of total calories from fat) diet, and body weight, blood glucose, insulin, leptin, lipids and GFR were monitored from 9 to 21 weeks or longer after the feeding programme. Mesangial cell dedifferentiation was accessed by alpha-smooth muscle actin staining. Glomerular hypertrophy was determined using image analysis with haematoxylin-eosin staining. Matrix deposition was determined by type IV collagen staining. RESULTS: After 9 weeks, mice fed a high-fat diet weighed more than mice fed a normal diet (30.5+/-1.2 vs 22.3+/-0.5 g, p<0.05), and mice fed a high-fat diet were hyperinsulinaemic (283.9+/-69.7 vs 102.9+/-36.4 pmol/l, p<0.05), hyperglycaemic (8.0+/-0.6 vs 6.5+/-0.2 mmol/l, p<0.05) and their leptin levels were increased six-fold (1.48+/-0.45 vs 0.25+/-0.03 ng/ml, p<0.05). After 13 weeks, mice fed a high-fat diet showed hyperfiltration (GFR; 440+/-60 vs 210+/-10 microl/min, p<0.05). During the early stages of diabetic nephropathy, mesangial cell dedifferentiation was evident, shown by increased expression of alpha-smooth muscle actin in the glomeruli. After 9 weeks, mice fed a high-fat diet already demonstrated increased type IV collagen deposition. After 13 weeks, they developed enlarged glomerular tufts compared with those of their age-matched controls. CONCLUSIONS/INTERPRETATION: The results of this study suggest that collagen IV deposition precedes the hyperfiltration and enlargement of glomeruli in early-stage diabetic nephropathy. Dedifferentiation of mesangial cells may be associated with collagen IV deposition.
Assuntos
Diabetes Mellitus Tipo 2/patologia , Gorduras na Dieta , Glomérulos Renais/patologia , Animais , Glicemia/metabolismo , Peso Corporal , Diferenciação Celular , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/fisiopatologia , Modelos Animais de Doenças , Insulina/sangue , Glomérulos Renais/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Aumento de PesoRESUMO
The enzyme, poly(ADP-ribose) polymerase (PARP), effects repair of DNA after ischemia-reperfusion (I/R) injury to cells in nerve and muscle tissue. However, its activation in severely damaged cells can lead to ATP depletion and death. We show that PARP expression is enhanced in damaged renal proximal tubules beginning at 6-12 h after I/R injury. Intraperitoneal administration of PARP inhibitors, benzamide or 3-amino benzamide, after I/R injury accelerates the recovery of normal renal function, as assessed by monitoring the levels of plasma creatinine and blood urea nitrogen during 6 days postischemia. PARP inhibition leads to increased cell proliferation at 1 day postinjury as assessed by proliferating cell nuclear antigen and improves the histopathological appearance of kidneys examined at 7 days postinjury. Furthermore, inhibition of PARP increases levels of ATP measured at 24 h postischemia compared with those in vehicle-treated animals. Our data indicate that PARP activation is a part of the cascade of molecular events that occurs after I/R injury in the kidney. Although caution is advised, transient inhibition of PARP postischemia may constitute a novel therapy for acute renal failure.
Assuntos
Rim/irrigação sanguínea , Inibidores de Poli(ADP-Ribose) Polimerases , Traumatismo por Reperfusão/prevenção & controle , Trifosfato de Adenosina/metabolismo , Animais , Benzamidas/farmacologia , Western Blotting , Reparo do DNA , Inibidores Enzimáticos/farmacologia , Imuno-Histoquímica , Rim/química , Masculino , Antígeno Nuclear de Célula em Proliferação/análise , Ratos , Ratos Sprague-DawleyRESUMO
De novo CD44 and ligand expression at wound margins accompanies cellular proliferation and migration that effect repair of injured mucosal and vascular endothelial tissues. To determine whether CD44 could play a role in recovery from acute ischemic renal injury, we characterized its renal expression and those of two of its ligands, hyaluronic acid and osteopontin. Although no expression is detectable in nonischemic kidneys, several mRNAs for CD44 are present within 1 day after injury. CD44 mRNA is expressed in proximal tubules undergoing repair. CD44 peptide is present in basal and lateral cell membranes. Hyaluronic acid is normally expressed in the interstitium of the renal papilla only. By 1 day postischemia, hyaluronic acid can be detected, in addition, in the interstitium surrounding regenerating tubules. Osteopontin, not normally expressed in the renal proximal tubule, is expressed in regenerating tubules by 3 days after induction of acute ischemic injury. Immunoreactive osteopontin peptide continues to be localized in those tubules still undergoing repair for as long as 7 days after the injury. Our data are consistent with a role for CD44-ligand interactions in the regenerating proximal tubule participating in the process of recovery after ischemic injury.
Assuntos
Receptores de Hialuronatos/metabolismo , Isquemia/metabolismo , Rim/metabolismo , Circulação Renal , Doença Aguda , Animais , Receptores de Hialuronatos/genética , Ácido Hialurônico/metabolismo , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/fisiopatologia , Masculino , Osteopontina , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Regeneração , Sialoglicoproteínas/metabolismo , Fatores de Tempo , Distribuição TecidualRESUMO
Hematological and clinical data are presented for a young Malay patient with a homozygous (delta beta)zero-thalassemic condition. His red blood cells contained 100% fetal hemoglobin with alpha and G gamma chains only. Detailed gene mapping defined a large deletion with a 5' end between the Aha III and Apa I sites, some 200-400 bp 5' to the A gamma globin gene and a 3' end beyond sequences 17-18 kb 3' to the beta globin gene. This G gamma (A gamma delta beta)zero-type of thalassemia is different from all the other six types described before. Comparison of the hematological data of this patient with those of homozygotes for either the Sicilian or Spanish types of G gamma A gamma (delta beta)zero-thalassemia showed no differences; all homozygotes have a moderate anemia which is accentuated by the relatively high oxygen affinity of the Hb F containing erythrocytes.
Assuntos
Talassemia/genética , Criança , Deleção Cromossômica , Mapeamento Cromossômico , Enzimas de Restrição do DNA , Hemoglobina Fetal/genética , Globinas/genética , Homozigoto , Humanos , Masculino , Talassemia/sangueRESUMO
Renal expression of osteopontin is enhanced in the setting of acute ischemic injury. Because of the parallels that exist between recovery from renal ischemia and renal development, we characterized the role that osteopontin plays during metanephrogenesis in the rat. Osteopontin mRNA is present in kidneys obtained from rat embryos as early as embryonic day 13 (E13). Immunohistochemical staining of metanephroi obtained from E16 rat embryos and metanephroi obtained from E13 embryos and cultured for 3 days in vitro demonstrated that osteopontin is expressed both in the developing nephron and in the ureteric bud. Addition of anti-osteopontin antibodies to metanephric organ cultures results in failure of the metanephric blastema to undergo normal tubulogenesis. Addition of the arginine-glycine-aspartic acid-containing peptide, cyclo-RGDfV, or the anti-alpha(v)beta3-integrin antibody, LM609, to cultures has a similar effect. These findings establish that osteopontin is produced within the rat metanephros during development in vivo and suggest that the binding of osteopontin to the alpha(v)beta3-integrin is required for tubulogenesis to occur in vitro. Blastemal cells within metanephroi cultured in the presence of OP199 manifest increased apoptosis compared with controls. It is possible that osteopontin plays an important anti-apoptotic role during the process of metanephric blastema condensation that is a prerequisite for the formation of nephrons in vivo.