RESUMO
OBJECTIVE: Extracellular inorganic pyrophosphate (ePPi) plays a key role in the regulation of normal and pathologic mineralization. The purpose of this work was to evaluate the role of P1 and P2 purine receptors in modulating ePPi production by articular chondrocytes. METHODS: Porcine cartilage explants and chondrocyte monolayers were cultured in the presence of P1 agonists, or a P2 agonist or antagonist and inhibitors of P2 signaling. Ambient media ePPi concentrations were measured after 48-96h. RESULTS: The P1 agonists NECA and CGS 21680 significantly decreased ePPi concentrations surrounding chondrocytes and cartilage explants. The P2 agonist, ADP, increased ePPi levels, and the P2 antagonist, suramin, decreased ePPi concentrations. Thapsigargin and 1,2 bis-(2-aminophenoxy)ethane-N,N,N'N'-tetra acetic acid (BAPTA), which dampen Ca(2+)-related P2 signaling, suppressed the response to ADP. CONCLUSIONS: Purine receptors are important regulators of ePPi production by chondrocytes. P1 receptor stimulation diminishes and P2 receptor stimulation enhances ePPi production. Alterations in receptor signaling or aberrations of extracellular purine nucleotide metabolism resulting in abnormal quantities or proportions of P1 and P2 receptor ligands could foster changes in ePPi production that in turn affect mineralization. We propose a homeostatic role for extracellular purine nucleotides and purine receptors in stabilizing ePPi concentrations.
Assuntos
Cartilagem Articular/metabolismo , Condrócitos/metabolismo , Difosfatos/metabolismo , Agonistas do Receptor Purinérgico P1/farmacologia , Agonistas do Receptor Purinérgico P2/farmacologia , Receptores Purinérgicos/efeitos dos fármacos , Difosfato de Adenosina/farmacologia , Animais , Células Cultivadas , AMP Cíclico/metabolismo , Antagonistas do Receptor Purinérgico P2/farmacologia , RNA Mensageiro/metabolismo , Suramina/farmacologia , SuínosRESUMO
Classical diagnostics for structural equation models are based on aggregate forms of the data and are ill suited for checking distributional or linearity assumptions. We extend recently developed goodness-of-fit tests for correlated data based on subject-specific residuals to structural equation models with latent variables. The proposed tests lend themselves to graphical displays and are designed to detect misspecified distributional or linearity assumptions. To complement graphical displays, test statistics are defined; the null distributions of the test statistics are approximated using computationally efficient simulation techniques. The properties of the proposed tests are examined via simulation studies. We illustrate the methods using data from a study of in utero lead exposure.
Assuntos
Biometria/métodos , Interpretação Estatística de Dados , Modelos Teóricos , Simulação por Computador , Feminino , Humanos , Chumbo , Exposição Materna , GravidezRESUMO
BACKGROUND: Little is known about mouse allergen exposure in home environments and the development of wheezing, asthma and atopy in childhood. OBJECTIVE: To examine the relation between mouse allergen exposure and wheezing, atopy, and asthma in the first 7 years of life. METHODS: Prospective study of 498 children with parental history of allergy or asthma followed from birth to age 7 years, with longitudinal questionnaire ascertainment of reported mouse exposure and dust sample mouse urinary protein allergen levels measured at age 2-3 months. RESULTS: Parental report of mouse exposure in the first year of life was associated with increased risk of transient wheeze and wheezing in early life. Current report of mouse exposure was also significantly associated with current wheeze throughout the first 7 years of life in the longitudinal analysis (P = 0.03 for overall relation of current mouse to current wheeze). However, early life mouse exposure did not predict asthma, eczema or allergic rhinitis at age 7 years. Exposure to detectable levels of mouse urinary protein in house dust samples collected at age 2-3 months was associated with a twofold increase in the odds of atopy (sensitization to >=1 allergen) at school age (95% confidence interval for odds ratio = 1.1-3.7; P = 0.03 in a multivariate analysis. CONCLUSIONS: Among children with parental history of asthma or allergies, current mouse exposure is associated with increased risk of wheeze during the first 7 years of life. Early mouse exposure was associated with early wheeze and atopy later in life.
Assuntos
Alérgenos/imunologia , Dermatite Atópica/imunologia , Poeira/imunologia , Camundongos/imunologia , Sons Respiratórios/imunologia , Adulto , Poluição do Ar em Ambientes Fechados , Animais , Criança , Pré-Escolar , Feminino , Seguimentos , Humanos , Lactente , Masculino , Testes Cutâneos , Inquéritos e QuestionáriosRESUMO
Previous studies have shown increased nucleotide pyrophosphohydrolase (EC 3.6.1.8) (NTPPHase) activity in detergent extracts of degenerated human cartilage containing calcium pyrophosphate dihydrate (CPPD) crystals relative to those from osteoarthritis or normal cartilage. NTPPHase was later shown to be an ectoenzyme and its activity was increased in synovial fluid from patients with CPPD crystal deposits relative to fluids from other types of arthritis. We have purified a soluble 61-kD NTPPHase from conditioned media of organ-cultured porcine articular cartilage to electrophoretic homogeneity. Its NH2-terminal sequence through 26 cycles showed < 30% homology to any previously reported protein sequence. An antibody raised to a synthetic peptide corresponding to this sequence reacted with denatured but not native enzyme. This antibody reacted against a sedimentable vesicle-associated 127-kD protein in conditioned media from cultured articular cartilage or from chondrocytes in primary monolayer culture and against a series of soluble proteins in conditioned media supernatant, including a 61-kD protein representing our original isolate. No reactivity was found in 1% SDS extracts of washed cultured chondrocytes, although these contained greater NTPPHase activity than the conditioned media. Antibody to PC-1, another ectoNTPPHase, reacted with 1% SDS extracts of whole chondrocytes but not against those chromatographic fractions containing the major portion of NTPPHase activity. Release of the vesicle-associated 127-kD enzyme into conditioned medium was stimulated three- to sevenfold by TGF beta 1. The antibody also reacted with a series of soluble proteins and with 127-kD sedimentable protein in human synovial fluid. Kinetic studies supported the existence of a unique vesicle-associated NTPPHase; apparent Km (mM) of chondrocyte membrane NTPPHase was 1.5 and 3.0 at pH 7.3 and 9.88, respectively; apparent Km (mM) of vesicle associated NTPPHase was 0.83 and 1.28 at pH 7.3 and 9.88. The data suggest the existence of a unique ecto-NTPPHase associated with vesicles derived from normal articular cartilage.
Assuntos
Cartilagem Articular/enzimologia , Cartilagem/enzimologia , Pirofosfatases/metabolismo , Sequência de Aminoácidos , Animais , Anticorpos , Western Blotting , Membrana Celular/enzimologia , Células Cultivadas , Cromatografia de Afinidade , Meios de Cultivo Condicionados , Eletroforese em Gel de Poliacrilamida , Humanos , Cinética , Dados de Sequência Molecular , Peso Molecular , Técnicas de Cultura de Órgãos , Osteoartrite/enzimologia , Pirofosfatases/química , Pirofosfatases/isolamento & purificação , Valores de Referência , SuínosRESUMO
In calcium pyrophosphate dihydrate (CPPD) crystal deposition disease, metabolic abnormalities favoring extracellular inorganic pyrophosphate (PPi) accumulation have been suspected. Elevations of intracellular PPi in cultured skin fibroblasts from a single French kindred with familial CPPD deposition (19) and elevated nucleoside triphosphate pyrophosphohydrolase activity (NTPPPH), which generates PPi in extracts of CPPD crystal-containing cartilages (14) favor this suspicion. To determine whether NTPPPH activity or PPi content of cells might be a disease marker expressed in extraarticular cells, human skin-derived fibroblasts were obtained from control donors and patients affected with the sporadic and familial varieties of CPPD (CPPD-S and CPPD-F) deposition. Intracellular PPi was elevated in both CPPD-S (P less than 0.05) and CPPD-F (P less than 0.01) fibroblasts compared with control fibroblasts. Ecto-NTPPPH activity was elevated in CPPD-S (P less than 0.01) but not CPPD-F. Intracellular PPi correlated with ecto-NTPPPH (P less than 0.01). Elevated PPi levels in skin fibroblasts may serve as a biochemical marker for patients with familial or sporadic CPPD crystal deposition disease; ecto-NTPPPH activity further separates the sporadic and familial disease types. Expression of these biochemical abnormalities in nonarticular cells implies a generalized metabolic abnormality.
Assuntos
Pirofosfato de Cálcio/metabolismo , Difosfatos/análise , Difosfatos/metabolismo , Distúrbios do Metabolismo do Fósforo/metabolismo , Pirofosfatases/metabolismo , Pele/metabolismo , Células Cultivadas , Cristalização , Fibroblastos/metabolismo , HumanosRESUMO
Hypophosphatasia features selective deficiency of activity of the tissue-nonspecific (liver/bone/kidney) alkaline phosphatase (ALP) isoenzyme (TNSALP); placental and intestinal ALP isoenzyme (PALP and IALP, respectively) activity is not reduced. Three phosphocompounds (phosphoethanolamine [PEA], inorganic pyrophosphate [PPi], and pyridoxal 5'-phosphate [PLP]) accumulate endogenously and appear, therefore, to be natural substrates for TNSALP. Carriers for hypophosphatasia may have decreased serum ALP activity and elevated substrate levels. To test whether human PALP and TNSALP are physiologically active toward the same substrates, we studied PEA, PPi, and PLP levels during and after pregnancy in three women who are carriers for hypophosphatasia. Hypophosphatasemia corrected during the third trimester because of PALP in maternal blood. Blood or urine concentrations of PEA, PPi, and PLP diminished substantially during that time. After childbirth, maternal circulating levels of PALP decreased, and PEA, PPi, and PLP levels abruptly increased. In serum, unremarkable concentrations of IALP and low levels of TNSALP did not change during the study period. We conclude that PALP, like TNSALP, is physiologically active toward PEA, PPi, and PLP in humans. We speculate from molecular/crystallographic information, indicating significant similarity of structure of the substrate-binding site of ALPs throughout nature, that all ALP isoenzymes recognize these same three phosphocompound substrates.
Assuntos
Fosfatase Alcalina/metabolismo , Hipofosfatasia/enzimologia , Isoenzimas/metabolismo , Gravidez/fisiologia , Difosfatos/metabolismo , Etanolaminas/metabolismo , Feminino , Heterozigoto , Humanos , Hipofosfatasia/genética , Placenta/enzimologia , Estudos Prospectivos , Fosfato de Piridoxal/metabolismo , Especificidade por SubstratoRESUMO
Associations between patient characteristics and survival were investigated in 432 patients with hepatocellular carcinoma. Those patients were prospectively studied by the Eastern Cooperative Oncology Group, and each had his or her diagnosis reconfirmed by a pathology review panel. There were 301 North American and 131 South African patients. Sixty-nine % of the North American patients and 82% of the South African patients were male. There were 187 Black patients, 62 of whom were from North America. The study population is unique among hepatocellular carcinoma patients in that eligibility, evaluability, and endpoint definitions were standardized, and patients from both North America and South Africa received similar treatments at a similar time. Factors with the most significant adverse effect on survival are impaired performance status, male sex, older age, and disease symptoms (jaundice and reduced appetite). There is no apparent difference in survival between White and Black patients within North America, but North American patients survived longer than South African patients. Among the different therapies, p.o. 5-fluorouracil was associated with the poorest median survival time (6 wk), and i.v. 5-fluorouracil plus semustine with the best median survival time (24 wk).
Assuntos
Carcinoma Hepatocelular/mortalidade , Neoplasias Hepáticas/mortalidade , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , PrognósticoRESUMO
PURPOSE: This three-armed phase III study in adults with advanced soft tissue sarcomas was planned as a comparison of objective regression rates, toxicity, and survival of patients receiving doxorubicin alone, ifosfamide plus doxorubicin, and mitomycin plus doxorubicin plus cisplatin. PATIENTS AND METHODS: Between December 1987 and July 1990, 279 patients with histologically confirmed sarcomas were enrolled to receive treatment A (doxorubicin 80 mg/m2), treatment B (ifosfamide 7.5 g/m2 plus doxorubicin 60 mg/m2), or treatment C (mitomycin 8 mg/m2 plus doxorubicin 40 mg/m2 plus cisplatin 60 mg/m2). RESULTS: Of 262 assessable patients, 74 (29%) achieved objective tumor regression. Objective regression occurred in 20% of the 90 patients who received doxorubicin alone (complete remission [CR] rate, 2%), in 34% of the 88 who received ifosfamide plus doxorubicin (CR rate, 3%), and in 32% of the 84 who received mitomycin plus doxorubicin plus cisplatin (CR rate, 7%). With grade 3 or greater myelosuppression in 53% of group A, 80% of group B, and 55% of group C, regimen B was significantly more myelosuppressive than either regimen A or C (P = .01) with two, three, and one treatment-related deaths, respectively. Synovial sarcomas were responsive to ifosfamide plus doxorubicin, especially among patients younger than 40 years of age. CONCLUSION: Ifosfamide plus doxorubicin produced a significantly higher regression rate (P = .03) than did doxorubicin alone; however, this was achieved at a level of myelosuppression significantly more intense than that produced by the single agent or by the three-drug combination. Mitomycin, doxorubicin, and cisplatin also appeared to be more active than the single agent; however, at a myelosuppression level similar to that of doxorubicin alone, this trend (P = .07) did not attain the usual level for significance. No significant survival differences were observed.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Doxorrubicina/uso terapêutico , Sarcoma/tratamento farmacológico , Neoplasias de Tecidos Moles/tratamento farmacológico , Adulto , Idoso , Idoso de 80 Anos ou mais , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Doenças da Medula Óssea/induzido quimicamente , Cisplatino/administração & dosagem , Doxorrubicina/administração & dosagem , Doxorrubicina/efeitos adversos , Feminino , Humanos , Ifosfamida/administração & dosagem , Masculino , Pessoa de Meia-Idade , Mitomicinas/administração & dosagem , Análise de Sobrevida , Resultado do TratamentoRESUMO
PURPOSE: To investigate mechanism-directed regimens in maximizing the efficacy of fluorouracil (5-FU) in advanced colorected cancer. PATIENTS AND METHODS: Based on promising phase II data, a randomized comparison of various methods for the biochemical modulation of 5-FU was undertaken in patients with advanced colorectal cancer. The control group received single-agent 5-FU as a 24-hour infusion weekly. Patients (N = 1,120) with no prior chemotherapy for metastatic disease were randomized to one of the following arms: arm A, 5-FU 2,600 mg/m2 by 24-hour infusion, weekly; arm B, N-phosphonoacetyl-l-aspartic acid 250 mg/m2 day l, 5-FU 2,600 mg/m2 by 24-hour infusion day 2, weekly; arm C, 5-FU 600 mg/m2 with oral leucovorin (LV) 125 mg/m2 hourly for the preceding 4 hours, weekly; arm D, 5-FU 600 mg/m2 with intravenous (IV) LV 600 mg/m2, weekly; arm E, 5-FU 750 mg/m2/d IV by continuous infusion for 5 days, then 750 mg/m2 weekly, and recombinant interferon alfa-2a 9 million units subcutaneously three times weekly. Median follow-up was 4.8 years. RESULTS: Of the 1,098 assessable patients, 57% had measurable disease. The toxicity of all the regimens was tolerable. Grade 4 or worse toxicity occurred in 11%, 11%, 30%, 24%, and 22% on each arm, respectively; diarrhea was the most common adverse effect. These toxicity patterns favored significantly (P <.001) the 24-hour infusion arms. Median survival (months) by arm was A, 14.8; B, 11.9; C, 13.5; D, 13.6; and E, 15.2. These survival durations did not differ significantly. CONCLUSION: We conclude that a weekly infusion regimen of 5-FU is significantly less toxic than and as effective as 5-FU bolus regimens modulated by either LV or interferon in patients with metastatic colorectal cancer.
Assuntos
Antineoplásicos/administração & dosagem , Ácido Aspártico/análogos & derivados , Ácido Aspártico/administração & dosagem , Neoplasias Colorretais/tratamento farmacológico , Fluoruracila/administração & dosagem , Interferon-alfa/administração & dosagem , Leucovorina/administração & dosagem , Ácido Fosfonoacéticos/análogos & derivados , Ácido Fosfonoacéticos/administração & dosagem , Administração Oral , Idoso , Neoplasias Colorretais/mortalidade , Feminino , Fluoruracila/efeitos adversos , Humanos , Infusões Intravenosas , Interferon alfa-2 , Masculino , Pessoa de Meia-Idade , Prognóstico , Proteínas RecombinantesRESUMO
Chorionic villus sampling (CVS) is a valued method of prenatal diagnosis that is often preferred over amniocentesis because it can be performed earlier, but which has also raised concern over a possible association with increased risk of terminal transverse limb deficiency (TTLD). We present and apply a meta-analytic method for estimating a combined dose-response effect from a series of case-control and cohort studies in which the exposure variable is interval-censored. Assuming coarsening at random for the interval-censoring, and calling upon the familiar result of Cornfield to pool case-control and cohort information on the association between a rare binary outcome and a multilevel exposure variable, we form a likelihood-based model to assess the effect of gestational age at the time of CVS on the presence or absence of a rare birth defect. Effect estimates are computed with a variant of the EM algorithm termed the method of weights, which enables the use of standard weighted regression software. Our findings suggest that CVS exposure at early gestational age leads to an increased risk of TTLD.
RESUMO
Hypophosphatasia (HPP) often leads to premature loss of deciduous teeth, due to disturbed cementum formation. We addressed the question to what extent cementum and dentin are similarly affected. To this end, we compared teeth from children with HPP with those from matched controls and analyzed them microscopically and chemically. It was observed that both acellular and cellular cementum formation was affected. For dentin, however, no differences in mineral content were recorded. To explain the dissimilar effects on cementum and dentin in HPP, we assessed pyrophosphate (an inhibitor of mineralization) and the expression/activity of enzymes related to pyrophosphate metabolism in both the periodontal ligament and the pulp of normal teeth. Expression of nucleotide pyrophosphatase phosphodiesterase 1 (NPP1) in pulp proved to be significantly lower than in the periodontal ligament. Also, the activity of NPP1 was less in pulp, as was the concentration of pyrophosphate. Our findings suggest that mineralization of dentin is less likely to be under the influence of the inhibitory action of pyrophosphate than mineralization of cementum.
Assuntos
Cemento Dentário/patologia , Dentina/patologia , Hipofosfatasia/patologia , Adolescente , Adulto , Estudos de Casos e Controles , Cementogênese/fisiologia , Criança , Pré-Escolar , Cemento Dentário/química , Polpa Dentária/enzimologia , Dentina/química , Dentinogênese/fisiologia , Difosfatos/análise , Humanos , Hipofosfatasia/metabolismo , Hipofosfatasia/fisiopatologia , Lactente , Microrradiografia , Minerais/análise , Ligamento Periodontal/enzimologia , Diester Fosfórico Hidrolases/análise , Pirofosfatases/análise , Calcificação de Dente/fisiologiaRESUMO
The aim of this study was to identify changes in cartilage intermediate layer protein/nucleotide pyrophosphohydrolase (CILP/NTPPH) expression in articular cartilage during aging. Adult (3-4 years old) and young (7-10 days old) porcine articular hyaline cartilage and fibrocartilage were studied by Northern blot analysis, in situ hybridization, and immunohistochemistry using a complementary DNA (cDNA) probe encoding porcine CILP/NTPPH and antibody to a synthetic peptide corresponding to a CILP/NTPPH sequence. Northern blot analysis of chondrocytes showed lower expression of CILP/NTPPH messenger RNA (mRNA) in young cartilage than in adult cartilage. In adult cartilage, extracellular matrix from the surface to the middeep zone was immunoreactive for CILP/NTPPH, especially in the pericellular matrix surrounding the middeep zone chondrocytes. In young cartilage, chondrocytes were moderately immunoreactive for CILP/NTPPH throughout all zones except the calcified zone. The matrix of young cartilage was negative except in the superficial zone. In young cartilage, CILP/NTPPH mRNA expression was undetectable. In adult cartilage, chondrocytes showed strong mRNA expression for CILP/NTPPH throughout middeep zones. Protein and mRNA signals were not detectable below the tidemark. CILP/NTPPH secretion into matrix around chondrocytes increases with aging. In this extracellular site it may generate inorganic pyrophosphate and contribute to age-related calcium pyrophosphate dihydrate crystal deposition disease.
Assuntos
Envelhecimento/metabolismo , Condrócitos/enzimologia , Proteínas da Matriz Extracelular/metabolismo , Pirofosfatases/metabolismo , Animais , Northern Blotting/métodos , Cartilagem Articular/citologia , Cartilagem Articular/enzimologia , Proteínas da Matriz Extracelular/genética , Expressão Gênica , Hialina , Pirofosfatases/genética , SuínosRESUMO
Hypophosphatasia is an inborn error of metabolism characterized by deficient activity of the tissue-nonspecific isoenzyme of alkaline phosphatase (TNSALP) and skeletal disease due to impaired mineralization of cartilage and bone matrix. We investigated two independently generated TNSALP gene knock-out mouse strains as potential models for hypophosphatasia. Homozygous mice (-/-) had < 1% of wild-type plasma TNSALP activity; heterozygotes had the predicted mean of approximately 50%. Phosphoethanolamine, inorganic pyrophosphate, and pyridoxal 5'-phosphate are putative natural substrates for TNSALP and all were increased endogenously in the knock-out mice. Skeletal disease first appeared radiographically at approximately 10 days of age and featured worsening rachitic changes, osteopenia, and fracture. Histologic studies revealed developmental arrest of chondrocyte differentiation in epiphyses and in growth plates with diminished or absent hypertrophic zones. Progressive osteoidosis from defective skeletal matrix mineralization was noted but not associated with features of secondary hyperparathyroidism. Plasma and urine calcium and phosphate levels were unremarkable. Our findings demonstrate that TNSALP knock-out mice are a good model for the infantile form of hypophosphatasia and provide compelling evidence for an important role for TNSALP in postnatal development and mineralization of the murine skeleton.
Assuntos
Fosfatase Alcalina/genética , Hipofosfatasia/genética , Fatores Etários , Fosfatase Alcalina/sangue , Animais , Animais Recém-Nascidos , Peso Corporal/genética , Modelos Animais de Doenças , Heterozigoto , Membro Posterior/diagnóstico por imagem , Membro Posterior/crescimento & desenvolvimento , Histocitoquímica , Homozigoto , Hipofosfatasia/diagnóstico por imagem , Hipofosfatasia/metabolismo , Camundongos , Camundongos Knockout , Fosfatos/urina , Fosfatidiletanolaminas/urina , Fosfato de Piridoxal/sangue , Radiografia , Tíbia/diagnóstico por imagem , Tíbia/crescimento & desenvolvimentoRESUMO
Two patients with linear scleroderma (en coup de sabre) developed systemic lupus erythematosus (SLE). This association has been well documented in only one previous case. The presence of high titer antibodies to ribonucleoprotein (RNP) initially led to the diagnosis of the mixed connective tissue disease. Development of more serious clinical involvement and antibodies to Sm (case 1) or native deoxyribonucleic acid (nDNA) (case 2) helped establish a diagnosis of SLE. Use of these studies in the differential diagnosis of systemic rheumatic diseases is disucssed briefly. The presence of anti-RNAP antibodies in patients with localized scleroderma may herald a more serious rheumatic disease.
Assuntos
Lúpus Eritematoso Sistêmico/complicações , Escleroderma Sistêmico/complicações , Adulto , Anticorpos Antinucleares , Biópsia , Criança , Diagnóstico Diferencial , Feminino , Humanos , Lúpus Eritematoso Sistêmico/diagnóstico , Lúpus Eritematoso Sistêmico/imunologia , Doença Mista do Tecido Conjuntivo/imunologia , Ribonucleoproteínas/imunologia , Escleroderma Sistêmico/imunologiaRESUMO
The porcine 127-kDa nucleotide pyrophosphohydrolase (NTPPHase) had been previously purified from the conditioned culture media of porcine articular cartilage. Protein sequencing of an internal 61-kDa proteolytic fragment of NTPPHase (61-kDa NTPPHase) determined the 26 N-terminal amino acids. This sequence was used to amplify a DNA fragment, which was used as a probe to clone the gene encoding the 61-kDa NTPPHase from a porcine chondrocyte cDNA library. DNA sequence analysis showed the cDNA insert to be 2509 bp, corresponding to a predicted open reading frame (ORF) encoding 599 amino acids. The 26 N-terminal amino acids of the 61-kDa NTPPHase were located within the ORF immediately downstream of a putative protease recognition region, RRKRR. This is consistent with this cDNA insert representing an internal proteolytic fragment of the full length 127-kDa NTPPHase. BLAST and FASTA analysis confirmed that the deduced amino acid sequence of 61-kDa NTPPHase was unique and did not possess a high degree of homology to sequence in the non-redundant protein and nucleotide databases. Proteins that possess limited homology (< 17%) with the 61-kDa NTTPPHase include several prokaryotic and eukaryotic ATP pyrophosphate-lyases (adenylate cyclase). Northern blot analysis of porcine chondrocyte RNA showed that the DNA encoding the 61-kDa NTPPHase hybridized to a single 4.0-kb RNA transcript. This DNA probe also hybridized to a single species of human chondrocyte RNA. Expression of a 61-kDa protein was detected by coupled in-vitro transcription/translation. Western blot analysis of this in-vitro transcription/translation reaction detected a 61-kDa protein, using an antibody raised against the peptide sequence that was originally used to clone the 61-kDa NTPPHase. These data indicate the successful in-vitro cloning and expression of the porcine chondrocyte 61-kDa NTPPHase. Future studies that utilize the gene encoding the 61-kDa NTPPHase may allow the characterization of the role of NTPPHase in calcium pyrophosphate dihydrate (CPPD) crystal deposition disease.
Assuntos
Condrócitos/enzimologia , Regulação Enzimológica da Expressão Gênica/genética , Pirofosfatases/genética , Sequência de Aminoácidos , Aminoácidos/análise , Animais , Sequência de Bases , Cartilagem Articular/enzimologia , Clonagem Molecular , DNA Complementar/genética , Biblioteca Gênica , Humanos , Dados de Sequência Molecular , Peso Molecular , Osteoartrite , Biossíntese de Proteínas , Pirofosfatases/química , RNA Mensageiro/análise , Análise de Sequência de DNA , Especificidade da Espécie , Suínos , Transcrição GênicaRESUMO
Advanced age is the most common risk factor for the development of calcium pyrophosphate dihydrate (CPPD) crystal-associated arthritis. However, the link between ageing and CPPD crystal formation in cartilage remains unexplained. In CPPD deposition disease, excess extracellular inorganic pyrophosphate (ePPi), generated by articular chondrocytes, accumulates in affected joints and contributes to CPPD crystallogenesis. Transforming growth factor beta 1 (TGF beta 1) is the first known physiologic stimulant of ePPi elaboration by adult porcine and human cartilage. We hypothesized that sensitivity of articular cartilage to the ePPi-stimulatory effects of TGF beta 1 may increase with ageing. Accordingly, we compared the effects of TGF beta 1 on cartilage ePPi elaboration from juvenile, young adult, and old adult pigs. Cartilage organ cultures from old animals increased ePPi elaboration in response to TGF beta 1 to a greater extent than did cartilage from juvenile and young adult animals. Similar results were seen in chondrocyte monolayers. Concurrent exposure to epidermal growth factor (EGF) augmented, but was not necessary for TGF beta 1-induced ePPi elaboration by adult cartilage. In contrast, in juvenile cartilage, concurrent exposure to EGF was required to permit TGF beta 1-induced ePPi elaboration. Thus, increased cartilage responsiveness to the ePPi-stimulatory effects of TGF beta 1 occurs with ageing, and may explain the link between advanced age and CPPD deposition disease.
Assuntos
Envelhecimento/metabolismo , Cartilagem Articular/metabolismo , Difosfatos/metabolismo , Substâncias de Crescimento/farmacologia , Animais , Meios de Cultura/metabolismo , Fator de Crescimento Epidérmico/farmacologia , Prolina/metabolismo , Pirofosfatases/metabolismo , Suínos , Timidina/farmacocinética , Fator de Crescimento Transformador beta/farmacologiaRESUMO
A case of acute erosive, reactive arthritis following Campylobacter jejuni-induced ulcerative colitis is presented. This is the 12th such case reported in the literature and the first in which destructive lesions of periarticular bone are demonstrated. A review of the literature suggests that reactive arthritis associated with C. jejuni infection is similar to that following other invasive types of bacterial diarrhea and is often associated with HLA-B27 lymphocyte antigen.
Assuntos
Artrite/complicações , Infecções por Campylobacter/complicações , Colite Ulcerativa/complicações , Doença Aguda , Artrite/diagnóstico por imagem , Artrografia , Campylobacter fetus , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
A patient with paroxysmal nocturnal hemoglobinuria, who required many blood transfusions for hemolytic episodes, had a persistent hyperchloremic metabolic acidosis. Bicarbonate infusion demonstrated a large fractional excretion of bicarbonate (28.6 per cent at a plasma bicarbonate level of 23 meq/liter) which was consistent with proximal renal tubular acidosis. Generalized aminoaciduria and decreased tubular reabsorption of phosphate were also present. Marked deposition of iron in renal proximal tubules was associated with these functional abnormalities. We believe that, as systemic acidosis can promote hemolysis in patients with paroxysmal nocturnal hemoglobinuria, hemolysis can lead, by way of iron deposition in renal tubules, to further acidosis. This cycle should be interrupted with appropriate doses of bicarbonate.
Assuntos
Acidose Tubular Renal/etiologia , Hemoglobinúria Paroxística/complicações , Adulto , Sangue , Hemólise , Humanos , Concentração de Íons de Hidrogênio , MasculinoRESUMO
Collagen fibers in synovial fluid sediment were described a decade ago. Since then, tissue-specific collagen molecules (types) have been characterized. Techniques were devised to identify the collagen types in joint fluid sediment. Collagens were found in 12 of 17 pellets prepared from fluid aspirates from 17 knee joints of patients with various forms of arthritis. Collagen types I and III and polypeptide chains A and B (basement membrane collagen) were specifically identified in four of seven fluids from patients with active systemic lupus erythematosus (SLE) and in a single fluid from a patient with severe septic arthritis. This "collagen profile" was identical to that of rheumatoid synovium. Type II collagen, characteristic of hyaline articular cartilage, was found in two of six fluids from osteoarthritic joints. The presence of sufficient collagen (about 5 micrograms) to permit typing was correlated with roentgenographic evidence of joint space narrowing; the presence of the "synovial" collagen profile was correlated with decreased joint fluid pH.
Assuntos
Artrite/metabolismo , Colágeno/análise , Líquido Sinovial/análise , Artrite Infecciosa/metabolismo , Artrite Reumatoide/metabolismo , Membrana Basal , Cartilagem Articular , Colágeno/imunologia , Humanos , Imunidade Celular , Lúpus Eritematoso Sistêmico/metabolismo , Osteoartrite/metabolismoRESUMO
Sixty-four patients were evaluated prospectively for a reflex sympathetic dystrophy syndrome (RSDS), using quantitative clinical measurements, high-resolution roentgenography and scintigraphy. Five separate groups were identified by their clinical features, allowing us to distinguish patients with definite or incomplete forms of the RSDS as well as 16 patients with other disorders. Scintigraphy was found to be a useful diagnostic study that may also provide a method of predicting therapeutic response. Systemic corticosteroid therapy proved to be a highly effective mode of treatment for up to 90 percent of the patients with the RSDS.