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Osteoporosis and other metabolic bone diseases are prevalent in the aging population. While bone has the capacity to regenerate throughout life, bone formation rates decline with age and contribute to reduced bone density and strength. Identifying mechanisms and pathways that increase bone accrual in adults could prevent fractures and accelerate healing. G protein-gated inwardly rectifying K+ (GIRK) channels are key effectors of G protein-coupled receptor signaling. Girk3 was recently shown to regulate endochondral ossification. Here, we demonstrate that deletion of Girk3 increases bone mass after 18 weeks of age. Male 24-week-old Girk3 -/- mice have greater trabecular bone mineral density and bone volume fraction than wildtype (WT) mice. Osteoblast activity is moderately increased in 24-week-old Girk3 -/- mice compared to WT mice. In vitro, Girk3-/- bone marrow stromal cells (BMSCs) are more proliferative than WT BMSCs. Calvarial osteoblasts and BMSCs from Girk3 -/- mice are also more osteogenic than WT cells, with altered expression of genes that regulate the wingless-related integration site (Wnt) family. Wnt inhibition via Dickkopf-1 (Dkk1) or ß-catenin inhibition via XAV939 prevents enhanced mineralization, but not proliferation, in Girk3 -/- BMSCs and slows these processes in WT cells. Finally, selective ablation of Girk3 from cells expressing Cre recombinase from the 2.3 kb-Col1a1 promoter, including osteoblasts and osteocytes, is sufficient to increase bone mass and bone strength in male mice at 24 weeks of age. Taken together, these data demonstrate that Girk3 regulates progenitor cell proliferation, osteoblast differentiation, and bone mass accrual in adult male mice.
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OBJECTIVE: The purpose of this study was to investigate the effect of age and oxidative stress on regulation of nuclear factor erythroid-2-related factor 2 (Nrf2) in young, old, and osteoarthritic (OA) human articular chondrocytes. DESIGN: Levels of Nrf2 in primary human chondrocytes isolated from young, old, and OA donors were measured by immunoblotting, qPCR, and immunohistochemistry. Effects on levels of Nrf2, antioxidant proteins regulated by Nrf2, as well as p65, and the anabolic response to insulin-like growth factor-1 (IGF-1) were evaluated after induction of oxidative stress with menadione, Nrf2 knockdown with siRNA, and/or Nrf2 activation with RTA-408. RESULTS: Nrf2 protein levels were significantly lower in older adult chondrocytes (â¼0.59 fold; p = 0.034) and OA chondrocytes (â¼0.50 fold; p = 0.016) compared to younger cells. Menadione significantly increased Nrf2 protein levels in young chondrocytes by just under four-fold without changes in old chondrocytes. Nrf2 knockdown and activation differentially regulated levels of anti-oxidant proteins including sulfiredoxin and NAD(P)H quinone dehydrogenase 1. Nrf2 activation with RTA-408 also decreased basal p65 phosphorylation, increased aggrecan and type II collagen gene expression, and increased production of proteoglycans in OA chondrocytes treated with IGF-1. CONCLUSIONS: Targeted therapeutic strategies aimed at maintaining Nrf2 activity could be useful in maintaining chondrocyte homeostasis through maintenance of intracellular antioxidant function and redox balance.
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Cartílago Articular , Factor 2 Relacionado con NF-E2 , Osteoartritis , Anciano , Humanos , Antioxidantes/farmacología , Cartílago Articular/metabolismo , Células Cultivadas , Condrocitos/metabolismo , Homeostasis , Factor I del Crecimiento Similar a la Insulina/farmacología , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Osteoartritis/metabolismo , Estrés Oxidativo/fisiología , Vitamina K 3/metabolismo , Vitamina K 3/farmacologíaRESUMEN
Long bones are formed and repaired through the process of endochondral ossification. Activation of G protein-coupled receptor (GPCR) signaling pathways is crucial for skeletal development and long bone growth. G protein-gated inwardly-rectifying K+ (GIRK) channel genes are key functional components and effectors of GPCR signaling pathways in excitable cells of the heart and brain, but their roles in non-excitable cells that directly contribute to endochondral bone formation have not been studied. In this study, we analyzed skeletal phenotypes of Girk2-/-, Girk3-/- and Girk2/3-/- mice. Bones from 12-week-old Girk2-/- mice were normal in length, but femurs and tibiae from Girk3-/- and Girk2/3-/- mice were longer than age-matched controls at 12-weeks-old. Epiphyseal chondrocytes from 5-day-old Girk3-/- mice expressed higher levels of genes involved in collagen chain trimerization and collagen fibril assembly, lower levels of genes encoding VEGF receptors, and produced larger micromasses than wildtype chondrocytes in vitro. Girk3-/- chondrocytes were also more responsive to the kappa opioid receptor (KOR) ligand dynorphin, as evidenced by greater pCREB expression, greater cAMP and GAG production, and upregulation of Col2a1 and Sox9 transcripts. Imaging studies showed that Kdr (Vegfr2) and endomucin expression was dramatically reduced in bones from young Girk3-/- mice, supporting a role for delayed vasculogenesis and extended postnatal endochondral bone growth. Together these data indicate that GIRK3 controls several processes involved in bone lengthening.
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Alargamiento Óseo , Canales de Potasio Rectificados Internamente Asociados a la Proteína G , Analgésicos Opioides/metabolismo , Animales , Encéfalo/metabolismo , Condrocitos/metabolismo , Canales de Potasio Rectificados Internamente Asociados a la Proteína G/genética , Canales de Potasio Rectificados Internamente Asociados a la Proteína G/metabolismo , RatonesRESUMEN
Endochondral ossification is tightly controlled by a coordinated network of signaling cascades including parathyroid hormone (PTH). Pleckstrin homology (PH) domain and leucine rich repeat phosphatase 1 (Phlpp1) affects endochondral ossification by suppressing chondrocyte proliferation in the growth plate, longitudinal bone growth, and bone mineralization. As such, Phlpp1-/- mice have shorter long bones, thicker growth plates, and proportionally larger growth plate proliferative zones. The goal of this study was to determine how Phlpp1 deficiency affects PTH signaling during bone growth. Transcriptomic analysis revealed greater PTH receptor 1 (Pth1r) expression and enrichment of histone 3 lysine 27 acetylation (H3K27ac) at the Pth1r promoter in Phlpp1-deficient chondrocytes. PTH (1-34) enhanced and PTH (7-34) attenuated cell proliferation, cAMP signaling, cAMP response element-binding protein (CREB) phosphorylation, and cell metabolic activity in Phlpp1-inhibited chondrocytes. To understand the role of Pth1r action in the endochondral phenotypes of Phlpp1-deficient mice, Phlpp1-/- mice were injected with Pth1r ligand PTH (7-34) daily for the first 4 weeks of life. PTH (7-34) reversed the abnormal growth plate and long-bone growth phenotypes of Phlpp1-/- mice but did not rescue deficits in bone mineral density or trabecular number. These results show that elevated Pth1r expression and signaling contributes to increased proliferation in Phlpp1-/- chondrocytes and shorter bones in Phlpp1-deficient mice. Our data reveal a novel molecular relationship between Phlpp1 and Pth1r in chondrocytes during growth plate development and longitudinal bone growth. © 2021 American Society for Bone and Mineral Research (ASBMR).
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Monoéster Fosfórico Hidrolasas , Receptor de Hormona Paratiroídea Tipo 1 , Animales , Proteínas Sanguíneas , Desarrollo Óseo , Condrocitos , Factor-23 de Crecimiento de Fibroblastos , Leucina , Ratones , Ratones Noqueados , Hormona Paratiroidea , Fosfoproteínas Fosfatasas , Fosfoproteínas , Receptor de Hormona Paratiroídea Tipo 1/genéticaRESUMEN
Primary bone tumors are rare cancers that cause significant morbidity and mortality. The recent identification of recurrent mutations in histone genes H3F3A and H3F3B within specific bone cancers, namely, chondroblastomas and giant cell tumors of bone (GCTB), has provided insights into the cellular and molecular origins of these neoplasms and enhanced understanding of how histone variants control chromatin function. Somatic mutations in H3F3A and H3F3B produce oncohistones, H3.3G34W and H3.3K36M, in more than nine of ten GCTB and chondroblastomas, respectively. Incorporation of the mutant histones into nucleosomes inhibits histone methyltransferases NSD2 and SETD2 to alter the chromatin landscape and change gene expression patterns that control cell proliferation, survival, and differentiation, as well as DNA repair and chromosome stability. The discovery of these histone mutations has facilitated more accurate diagnoses of these diseases and stratification of malignant tumors from benign tumors so that appropriate care can be delivered. The broad-scale epigenomic and transcriptomic changes that arise from incorporation of mutant histones into chromatin provide opportunities to develop new and disease-specific therapies. In this chapter, we review how mutant histones inhibit SETD2 and NSD2 function in bone tumors and discuss how this information could lead to better treatments for these cancers.
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Neoplasias Óseas , Condroblastoma , Tumor Óseo de Células Gigantes , Histonas/genética , Mutación , Neoplasias Óseas/genética , Condroblastoma/genética , Tumor Óseo de Células Gigantes/genética , N-Metiltransferasa de Histona-Lisina , Humanos , Proteínas RepresorasRESUMEN
Long bone formation is a complex process that requires precise transcriptional control of gene expression programs in mesenchymal progenitor cells. Histone deacetylases (Hdacs) coordinate chromatin structure and gene expression by enzymatically removing acetyl groups from histones and other proteins. Hdac inhibitors are used clinically to manage mood disorders, cancers, and other conditions but are teratogenic to the developing skeleton and increase fracture risk in adults. In this study, the functions of Hdac3, one of the enzymes blocked by current Hdac inhibitor therapies, in skeletal mesenchymal progenitor cells were determined. Homozygous deletion of Hdac3 in Prrx1-expressing cells prevented limb lengthening, altered pathways associated with endochondral and intramembranous bone development, caused perinatal lethality, and slowed chondrocyte and osteoblast differentiation in vitro. Transcriptomic analysis revealed that Hdac3 regulates vastly different pathways in mesenchymal cells expressing the Prxx1-Cre driver than those expressing the Col2-CreERT driver. Notably, Fgf21 was elevated in Hdac3-CKOPrrx1 limbs as well as in chondrogenic cells exposed to Hdac3 inhibitors. Elevated expression of Mmp3 and Mmp10 transcripts was also observed. In conclusion, Hdac3 regulates distinct pathways in mesenchymal cell populations and is required for mesenchymal progenitor cell differentiation and long bone development. © 2017 American Society for Bone and Mineral Research.
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Desarrollo Óseo , Eliminación de Gen , Histona Desacetilasas/metabolismo , Células Madre Mesenquimatosas/metabolismo , Animales , Apoptosis , Huesos/patología , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Condrocitos/metabolismo , Factores de Crecimiento de Fibroblastos/metabolismo , Regulación de la Expresión Génica , Placa de Crecimiento/patología , Mesodermo/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Osteoblastos/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Cráneo/patologíaRESUMEN
OBJECTIVE: Spinocerebellar ataxia (SCA) leads to abnormal ocular motility and alignment. The objective of this study was to quantitatively assess vision, ocular motility and alignment and its impact on vision related quality of life (VRQOL) in SCA. METHODS: Nineteen genetically diagnosed SCA subjects (11 SCA type 3, 3 SCA type 1 and 5 SCA type 6) participated at two university centers. All subjects completed the National Eye Institute Visual Function Questionnaire (NEI-VFQ), 10-Item Neuro-Ophthalmic Supplement (NOS), scale for assessment and rating of ataxia (SARA) and ophthalmic examination. Twelve subjects seen at one of the 2 sites underwent quantitative ocular motility and alignment assessment. RESULTS: Composite scores for NEI-VFQ (mean 76.3±13) and NOS (mean 65.2±16.8) were significantly decreased in SCA subjects. NEI-VFQ subscale scores were decreased for general, near, distance and peripheral vision and driving. SCA patients had decreased low contrast sensitivity, stereoacuity and multiple ocular motility defects which included gaze limitation (9/12), nystagmus (5/12), distance esophoria (11/12), near exophoria (12/12) and receded near point of convergence. A significant negative correlation was noted between composite scores and distance convergence fusional amplitude. CONCLUSION: VRQOL is significantly decreased in SCA compared to normal population. All SCA patients should be screened for visual disability and referred for neuro-ophthalmic assessment promptly.
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Trastornos de la Motilidad Ocular/fisiopatología , Calidad de Vida , Ataxias Espinocerebelosas/fisiopatología , Trastornos de la Visión/fisiopatología , Agudeza Visual/fisiología , Adulto , Anciano , Sensibilidad de Contraste/fisiología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Trastornos de la Motilidad Ocular/etiología , Ataxias Espinocerebelosas/complicaciones , Trastornos de la Visión/etiologíaRESUMEN
Familial occurrence of myasthenia gravis is uncommon and reports of maternal transmission of muscle-specific tyrosine kinase (MuSK) antibody myasthenia are rarer still. We report two families with maternal transmission of MuSK antibody myasthenia gravis to the offspring by different mechanisms. The first family demonstrates transmission genetic susceptibility of inheriting myasthenia gravis from MuSK antibodies, whereas the second one demonstrates transplacental transmission of MuSK antibodies at birth.
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Inmunidad Materno-Adquirida , Intercambio Materno-Fetal , Miastenia Gravis/genética , Proteínas Tirosina Quinasas Receptoras/genética , Receptores Colinérgicos/genética , Adulto , Autoanticuerpos/administración & dosificación , Autoanticuerpos/biosíntesis , Niño , Femenino , Humanos , Inmunidad Materno-Adquirida/genética , Recién Nacido , Masculino , Intercambio Materno-Fetal/genética , Miastenia Gravis/inmunología , Miastenia Gravis/metabolismo , Embarazo , Proteínas Tirosina Quinasas Receptoras/administración & dosificación , Receptores Colinérgicos/administración & dosificación , Adulto JovenRESUMEN
Studies have documented the transmission of HIV in incarcerated populations resulting from injection drug use or sexual activity. Less than 1% of the jails and prisons in the United States allow inmates access to condoms, and none allows access to needles. Results of a survey to measure the acceptability of a condom distribution program at the Washington, DC. Central Detention Facility, where condoms are available to inmates, are presented here. Three hundred seven inmates and 100 correctional officers were surveyed from October 2000 through October 2001. The surveys demonstrate that the program is generally supported and thought to be important by inmates and correctional staff. The program has not resulted in any major security infractions and could be replicated in other correctional settings.
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Actitud Frente a la Salud , Condones/provisión & distribución , Infecciones por VIH/prevención & control , Prisiones , Adulto , Recolección de Datos , District of Columbia , Infecciones por VIH/transmisión , Accesibilidad a los Servicios de Salud , Humanos , Masculino , Conducta SexualRESUMEN
A sensitive and highly reproducible method has been used to show that Aquaporin 3 (AQP(3)) mRNA is present in the ovine placenta and chorion from at least 60 days of gestation (term=145-150d) with levels increasing substantially (>16 fold) at 100 days, and remaining constant thereafter. By immuno- and hybridization histochemistry, the epithelial cells expressing AQP(3)were found to be the trophoblast cells. Some AQP(3)was expressed in fibroblasts of the amnion and allantois but none was expressed in the epithelia of these membranes. AQP(1)was expressed in endothelial cells of fetal and maternal blood vessels but not in any epithelial cell of the ovine placenta and fetal membranes. The level of AQP(3)expression is consistent with known ovine placental permeabilities to water, glycerol and urea.
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Acuaporinas/metabolismo , Membranas Extraembrionarias/metabolismo , Placenta/metabolismo , Secuencia de Aminoácidos , Animales , Acuaporina 1 , Acuaporina 3 , Acuaporinas/genética , Secuencia de Bases , Cartilla de ADN/genética , ADN Complementario/genética , Membranas Extraembrionarias/crecimiento & desarrollo , Femenino , Regulación del Desarrollo de la Expresión Génica , Edad Gestacional , Hibridación in Situ , Datos de Secuencia Molecular , Permeabilidad , Placentación , Embarazo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , OvinosRESUMEN
The aim of this study was to test the hypothesis that the relative insensitivity of the ovine fetal kidney to arginine vasopressin (AVP) is due to low levels of expression of the gene for aquaporin-2 (AQP2) which encodes the AVP-regulated water channel. We report the cloning of the cDNA for the ovine AQP2 which has a major transcript at 4.2 kilobases (kb) and a minor transcript at 1.5 kb, resembling the human gene transcripts. At 40-60 days' (term = 145-150 days'), mRNA levels are very low, detectable only by reverse transcription-polymerase chain reaction (RT-PCR). By Northern blot analysis AQP2 mRNA is detectable at 75 days'. The ratio of AQP2/glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA increases approximately 2.4-fold between 100 and 140 days' when it is about 41% of adult values. Both glucocorticoids and the renin-angiotensin system are involved in maturation of renal function. When fetuses at 75 or 85 days of gestation were exposed to high levels of dexamethasone for 2-3 days, mRNAs for both GAPDH and AQP2 doubled, but the ratio was unchanged. Angiotensin I, infused for 3 days at 115-120 days' gestation, increased the AQP2/GAPDH mRNA ratios by twofold (major transcript) and sixfold (minor transcript), which were highly significant (P<0.001). The increasing sensitivity of the ovine fetal kidney to AVP, from 100-140 days of gestation, is largely due to increasing AQP2 gene expression over this period.
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Acuaporinas/genética , ADN Complementario/genética , Riñón/fisiología , Angiotensina I/farmacología , Animales , Acuaporina 2 , Acuaporina 6 , Arginina Vasopresina/farmacología , Northern Blotting , Clonación Molecular , Feto , Gliceraldehído-3-Fosfato Deshidrogenasas/genética , Humanos , Inmunohistoquímica , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , ARN Mensajero/análisis , OvinosRESUMEN
1. The earliest form of the kidney, the pronephros, does not really occur in the ovine embryo; instead, a giant glomerulus forms at the anterior end of the mesonephros. 2. In the sheep, the mesonephros is present from 11-38% of total gestation (150 days) and produces a dilute urine, as well as expressing the genes for erythropoietin, renin, angiotensinogen, angiotensin-converting enzyme and the angiotensin II (AngII) receptors AT1 and AT2. 3. The ovine metanephros begins to develop at 18% of gestation and nephrogenesis is complete several weeks before birth. All components of the renin-angiotensin system (RAS) are expressed from at least 27% of gestation. 4. Both AT1 and AT2 receptors are expressed by the adrenocortical cells early in gestation but, at mid-gestation, exogenous AngII does not stimulate aldosterone secretion in vivo. 5. Preliminary results suggest that AngII has important roles in renal development in the ovine foetus but the role(s), if any, in adrenal development, remains to be investigated.
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Glándulas Suprarrenales/embriología , Riñón/embriología , Sistema Renina-Angiotensina/fisiología , Ovinos/embriología , Glándulas Suprarrenales/patología , Angiotensina II/fisiología , Animales , Desarrollo Embrionario y Fetal/fisiología , Humanos , Riñón/patologíaRESUMEN
The cDNA for the ovine aquaporin 1 (AQP1) was obtained and found to be 97%, 88%, and 85%, respectively, homologous to the bovine, human, and rat AQP1 cDNA. The level of total kidney mRNA expressed as a ratio to glyceraldehyde-3-phosphate dehydrogenase increased sevenfold from 60 days to 140 days of gestation (term=150 days) and reached adult values by 6 weeks after birth. Treatment of pregnant ewes (and their fetuses) at 64 and 74 days of gestation with dexamethasone (0.76 mg/h for 48 h) resulted in a small but statistically significant increase in AQP1 mRNA only in the 74-day fetuses. By immunohistochemistry, it was shown that the increase in AQP1 mRNA with dexamethasone resulted largely from an increase in maturity of the inner zone of the fetal renal cortex (i.e., more tubules) as well as stronger expression of AQP1 in proximal tubules and thin descending limbs of loops of Henle. A similar effect occurred in fetuses infused for 3 days with angiotensin I (6.7 microg/h) in the last third of gestation.
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Acuaporinas/genética , ADN Complementario/genética , Regulación del Desarrollo de la Expresión Génica/fisiología , Riñón/crecimiento & desarrollo , Riñón/metabolismo , Adulto , Secuencia de Aminoácidos , Animales , Acuaporina 1 , Acuaporinas/biosíntesis , Secuencia de Bases , Antígenos de Grupos Sanguíneos , Northern Blotting , Bovinos , Clonación Molecular , Cartilla de ADN , ADN Complementario/biosíntesis , Dexametasona/farmacología , Femenino , Glucocorticoides/farmacología , Humanos , Inmunohistoquímica , Riñón/efectos de los fármacos , Riñón/embriología , Datos de Secuencia Molecular , Embarazo , ARN Mensajero/biosíntesis , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , OvinosRESUMEN
The 16S rRNA gene sequences of Salmonella typhi and Salmonella typhimurium were amplified by PCR, cloned, and sequenced. These sequences were analyzed by comparison with reference organisms from the family Enterobacteriaceae. Both S. typhi and S. typhimurium belong to the gamma subdivision of the class Proteobacteria.
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ADN Bacteriano/análisis , ARN Ribosómico 16S/análisis , Salmonella typhi/genética , Salmonella typhimurium/genética , Datos de Secuencia Molecular , Filogenia , Salmonella typhi/clasificación , Salmonella typhimurium/clasificaciónRESUMEN
The distributions of aquaporin-1 mRNA and protein were studied by hybridization histochemistry with a homologous riboprobe and immunohistochemistry, in the adult sheep kidney. Heaviest labelling occurred in the thin descending limb (DTL) of the loop of Henle in the inner stripe of the outer medulla, with apparent decreasing expression in the inner medulla, outer stripe of the outer medulla and cortex, but no quantitation was performed. Only proximal tubules (PT) (convoluted and straight) and DTL labelled. The glomerulus showed no labelling, consistent with the pattern in the rat but different to that in the human. During ontogeny, no labelling occurred in the mesonephros at 27 or 41 days of gestation (term = 145-150 days) but other structures did label at 27 days (heart, lung bud, blood vessels surrounding developing spinal cord). Labelling first occurred faintly in the metanephros at 41 days of gestation and increased throughout gestation consistent with morphological development of nephrons.
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Acuaporinas , Canales Iónicos/análisis , Canales Iónicos/genética , Médula Renal/química , Médula Renal/embriología , Factores de Edad , Animales , Acuaporina 1 , Feto/fisiología , Regulación del Desarrollo de la Expresión Génica , Immunoblotting , Inmunohistoquímica , Hibridación in Situ , Sondas ARN , ARN Mensajero/análisis , OvinosRESUMEN
cDNAs encoding three phospholipase A2 (PLA2) isoforms in Naja naja sputatrix were cloned and characterized. One of them encoded an acidic PLA2 (APLA) while the others encoded neutral PLA2 (NPLA-1 and NPLA-2). The specific characteristics of APLA and NPLA were attributed to mutations at nt139 and nt328 from G to C and G to A, respectively, resulting in amino acid substitutions from Asp20 and 83 in APLA to His20 and Asn83 in NPLA. Amino acid sequencing of purified protein also showed the presence of this Asp20 and His20 in APLA and NPLA, respectively. The cDNA encoding one of the PLA2 (NAJPLA-2A), when expressed in Escherichia coli, yielded a protein that exhibited PLA2 activity.
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ADN Complementario/aislamiento & purificación , Venenos Elapídicos/enzimología , Isoenzimas/química , Isoenzimas/genética , Fosfolipasas A/química , Fosfolipasas A/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , ADN Complementario/química , Venenos Elapídicos/química , Venenos Elapídicos/genética , Elapidae , Regulación de la Expresión Génica , Genes , Isoenzimas/aislamiento & purificación , Datos de Secuencia Molecular , Fosfolipasas A/aislamiento & purificación , Fosfolipasas A2 , Proteínas Recombinantes/metabolismoRESUMEN
In the ovine fetus at 41 days of gestation (term is 150 days), there are two sets of kidneys, mesonephrol and metanephrol. We have examined the expression of the erythropoietin (Epo) gene in both types of kidneys by competitive reverse transcriptase-polymerase chain reaction and hybridization histochemistry and compared the expression to that of the 60-day fetal metanephros. At 41 days, the Epo gene was expressed in both mesonephros and metanephros, as well as in the fetal liver. The cells expressing the Epo gene in the mesonephros were interstitial cells in the vicinity of the proximal tubules.