RESUMO
Advances in genomic diagnostics hold promise for improved care of rare hematologic diseases. Here, we describe a novel targeted therapeutic approach for Ghosal hematodiaphyseal dysplasia, an autosomal recessive disease characterized by severe normocytic anemia and bone abnormalities due to loss-of-function mutations in thromboxane A synthase 1 (TBXAS1). TBXAS1 metabolizes prostaglandin H2 (PGH2), a cyclooxygenase (COX) product of arachidonic acid, into thromboxane A2. Loss-of-function mutations in TBXAS result in an increase in PGH2 availability for other PG synthases. The current treatment for Ghosal hematodiaphyseal dysplasia syndrome consists of corticosteroids. We hypothesize that nonsteroidal anti-inflammatory drugs (NSAIDs), which inhibit COX-1 and COX-2, could ameliorate the effects of TBXAS1 loss and improve hematologic function by reducing prostaglandin formation. We treated 2 patients with Ghosal hematodiaphyseal dysplasia syndrome, an adult and a child, with standard doses of NSAIDs (aspirin or ibuprofen). Both patients had rapid improvements concerning hematologic parameters and inflammatory markers without adverse events. Mass spectrometry analysis demonstrated that urinary PG metabolites were increased along with proinflammatory lipoxygenase (LOX) products 5-hydroxyeicosatetraenoic acid and leukotriene E4. Our data show that NSAIDs at standard doses surprisingly reduced both COX and LOX products, leading to the resolution of cytopenia, and should be considered for first-line treatment for Ghosal hematodiaphyseal dysplasia syndrome.
Assuntos
Anemia Refratária , Anemia , Pancitopenia , Adulto , Criança , Humanos , Anemia Refratária/tratamento farmacológico , Anemia Refratária/genética , Anti-Inflamatórios não Esteroides/uso terapêutico , Anemia/tratamento farmacológico , Prostaglandina H2 , Síndrome , Transtornos da Insuficiência da Medula ÓsseaRESUMO
Prostaglandin endoperoxide H synthase (PGHS) is a heme-enzyme responsible for the conversion of arachidonic acid (AA) to prostaglandin H2 (PGH2). PGHS have both oxygenase (COX) and peroxidase (POX) activities and is present in two isoforms (PGHS-1 and -2) expressed in different tissues and cell conditions. It has been reported that PGHS activity is inhibited by the nitrated form of AA, nitro-arachidonic acid (NO2AA), which in turn could be synthesized by PGHS under nitro-oxidative conditions. Specifically, NO2AA inhibits COX in PGHS-1 as well as POX in both PGHS-1 and -2, in a dose and time-dependent manner. NO2AA inhibition involves lowering the binding stability and displacing the heme group from the active site. However, the complete mechanism remains to be understood. This review describes the interactions of PGHS with NO2AA, focusing on mechanisms of inhibition and nitration. In addition, using a novel approach combining EPR-spin trapping and mass spectrometry, we described possible intermediates formed during PGHS-2 catalysis and inhibition. This literature revision as well as the results presented here strongly suggest a free radical-dependent inhibitory mechanism of PGHS-2 by NO2AA. This is of relevance towards understanding the underlying mechanism of inhibition of PGHS by NO2AA and its anti-inflammatory potential.
Assuntos
Anti-Inflamatórios/química , Ácido Araquidônico/química , Ciclo-Oxigenase 2/química , Inibidores Enzimáticos/química , Nitrocompostos/química , Prostaglandina H2/química , Anti-Inflamatórios/metabolismo , Anti-Inflamatórios/farmacologia , Ácido Araquidônico/metabolismo , Ácido Araquidônico/farmacologia , Biocatálise , Ciclo-Oxigenase 2/metabolismo , Espectroscopia de Ressonância de Spin Eletrônica , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Heme/química , Heme/metabolismo , Humanos , Espectrometria de Massas , Nitrocompostos/metabolismo , Nitrocompostos/farmacologia , Prostaglandina H2/antagonistas & inibidores , Prostaglandina H2/biossíntese , Ligação ProteicaRESUMO
AIMS: To determine the role of reactive oxygen species (ROS) on sodium nitroprusside (SNP)-induced tolerance. Additionally, we evaluated the role of ROS on NF-κB activation and pro-inflammatory cytokines production during SNP-induced tolerance. MAIN METHODS: To induce in vitro tolerance, endothelium-intact or -denuded aortic rings isolated from male Balb-c mice were incubated for 15, 30, 45 or 60min with SNP (10nmol/L). KEY FINDINGS: Tolerance to SNP was observed after incubation of endothelium-denuded, but not endothelium-intact aortas for 60min with this inorganic nitrate. Pre-incubation of denuded rings with tiron (superoxide anion (O2-) scavenger), and the NADPH oxidase inhibitors apocynin and atorvastatin reversed SNP-induced tolerance. l-NAME (non-selective NOS inhibitor) and l-arginine (NOS substrate) also prevented SNP-induced tolerance. Similarly, ibuprofen (non-selective cyclooxygenase (COX) inhibitor), nimesulide (selective COX-2 inhibitor), AH6809 (prostaglandin PGF2α receptor antagonist) or SQ29584 [PGH2/thromboxane TXA2 receptor antagonist] reversed SNP-induced tolerance. Increased ROS generation was detected in tolerant arteries and both tiron and atorvastatin reversed this response. Tiron prevented tolerance-induced increase on O2- and hydrogen peroxide (H2O2) levels. The increase onp65/NF-κB expression and TNF-α production in tolerant arteries was prevented by tiron. The major new finding of our study is that SNP-induced tolerance is mediated by NADPH-oxidase derived ROS and vasoconstrictor prostanoids derived from COX-2, which are capable of reducing the vasorelaxation induced by SNP. Additionally, we found that ROS mediate the activation of NF-κB and the production of TNF-α in tolerant arteries. SIGNIFICANCE: These findings identify putative molecular mechanisms whereby SNP induces tolerance in the vasculature.
Assuntos
Aorta/metabolismo , Ciclo-Oxigenase 2/metabolismo , Nitroprussiato/farmacologia , Prostaglandina H2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Vasodilatação/efeitos dos fármacosRESUMO
En las glomerulopatías, las Microproteínas Urinarias (MU) se eliminan por distintos mecanismos fisiopatológicos. El objetivo del trabajo fue correlacionar las MU con el daño histológico evaluado en la punción biopsia renal (PBR) de pacientes con diversas glomerulopatías. Se estudiaron 44 orinas espontáneas (33 mujeres y 11 varones) entre 18 y 71 años de edad, por el método de electroforesis bidimensional de uso clínico (2D UC). Las proteínas identificadas se dividieron en 5 grupos y se compararon con lesiones vasculares, glomerulares y túbulointersticiales; estas dos últimas se dividieron en crónicas y agudas. Las del grupo identificado como las "Tres Marías" (fragmento de 35 kDa de la proteína inhibidor de tripsina cadena pesada H4, Prostaglandina H2 sintasa y fragmento de 23 kDa del Perlecan) resultaron no ser marcadoras de daño tubular, sino de alteraciones glomerulares crónicas. La presencia de las mismas, con proteinuria, se observa antes de la caída de la filtración glomerular (<60 mL/min) y correlaciona con el 30% de glomérulos totalmente esclerosados (p<0,001). El grupo Triángulo, en la glomerulopatía, contiene a la Alfa-1 microglobulina (A1m) y a las cadenas livianas libres de Inmunoglobulinas (CLL), e indica lesión glomerular activa. Por lo tanto, las MU en glomerulopatías, responden a lesiones glomerulares activas y crónicas. Los perfiles proteicos urinarios hallados por la 2D UC permitieron conocer el grado de lesión en los distintos compartimentos renales.
In Glomerulopathies, urinary microproteins (UM) are eliminated by different pathophysiological mechanisms. The objective of this work was to correlate the UM with the histological damage evaluated in the renal biopsy through puncture of patients with various glomerular diseases. Forty-four urine samples (33 females, 11 males) aged 18 to 71 years old were studied by the method of two-dimensional electrophoresis of clinical use (2D UC). The identified proteins were divided into 5 groups and were compared with vascular injury, glomerular and tubulointerstitial injury, the latter in chronic and acute cases. The group identified as the "Three Marias" (fragment 35 kDa protein trypsin inhibitor heavy chain H4, prostaglandin H2 synthase and 23 kDa fragment of Perlecan), was not found as marker of tubular damage, but it was found in chronic glomerular disorders. The presence of this same group -with proteinuria- is seen before the collapse of the glomerular filtration rate (<60 mL/min) and it is correlated with 30% of fully sclerotic glomeruli (p<0.001). In glomerulopathy, the Triangle group: Alpha-1 microglobulin (A1m) and free Immunoglobulin light chains (FLC), indicates active glomerular injury. Therefore, the UM in glomerular diseases, respond to active and chronic glomerular lesions. Urinary protein profiles found by the 2D UC made it possible to know the degree of renal injury in different renal compartments.
Nas glomerulopatias, as Micro proteínas Urinárias (MU) são eliminadas através de diferentes mecanismos fisiológicos. O objetivo do trabalho foi relacionar as MU com o dano histológico avaliado na punção biópsia renal (PBR) de pacientes com diversas glomerulopatias. Foram estudadas 44 urinas espontâneas (33 mulheres e 11 homens entre 18 e 71 anos de idade), pelo método de eletroforese bidimensional de uso clínico (2D UC). As proteínas identificadas foram divididas em 5 grupos e comparadas com lesões vasculares, glomerulares e túbulo-intersticiais, estas duas últimas classificadas em crônicas e agudas. O grupo identificado como as "Três Marias" (fragmento de 35 kDa da proteína inibidora de tripsina cadeia pesada H4, Prostaglandina H2 sintase e fragmento de 23 kDa do Perlecam), resultaram não ser marcadoras de dano tubular, mas de alterações glomerulares crônicas. A presença de tais proteínas, com proteinúria, observa-se antes da queda da filtragem glomerular (<60 mL/min) e correlaciona com 30% de glomérulos totalmente esclerosados (p<0,001). O grupo Triângulo, na glomerulopatia, que contém a Alfa-1 microglobulina (A1m) e as cadeias leves livres de Imunoglobulinas (CLL) indicam lesão glomerular ativa. Portanto, as MU em glomerulopatias respondem a lesões glomerulares ativas e crônicas. Os perfis proteicos urinários encontrados pela 2D UC permitiram conhecer o grau de lesão nos diferentes compartimentos renais.
Assuntos
Humanos , Masculino , Feminino , Adolescente , Adulto , Pessoa de Meia-Idade , Idoso , Biópsia por Agulha , Rim , Proteínas , Coleta de Urina , Epidemiologia Descritiva , Ligases , Estudo Observacional , Prostaglandina H2RESUMO
AIMS: Na(+), K(+)-ATPase activity contributes to the regulation of vascular contractility and it has been suggested that vascular Na(+), K(+)-ATPase activity may be altered during the progression of diabetes; however the mechanisms involved in the altered Na(+), K(+)-ATPase activity changes remain unclear. Thus, the aim of the present study was to evaluate ouabain-sensitive Na(+), K(+)-ATPase activity and the mechanism(s) responsible for any alterations on this activity in aortas from 1- and 4-week streptozotocin-pretreated (50 mg kg(-1), i.v.) rats. MAIN METHODS: Aortic rings were used to evaluate the relaxation induced by KCl (1-10mM) in the presence and absence of ouabain (0.1 mmol/L) as an index of ouabain-sensitive Na(+), K(+)-ATPase activity. Protein expression of COX-2 and p-PKC-betaII in aortas were also investigated. KEY FINDINGS: Ouabain-sensitive Na(+), K(+)-ATPase activity was unaltered following 1-week of streptozotocin administration, but was increased in the 4-week diabetic aorta (27%). Endothelium removal or nitric oxide synthase inhibition with l-NAME decreased ouabain-sensitive Na(+), K(+)-ATPase activity only in control aortas. In denuded aortic rings, indomethacin, NS-398, ridogrel or Gö-6976 normalized ouabain-sensitive Na(+), K(+)-ATPase activity in 4-week diabetic rats. In addition, COX-2 (51%) and p-PKC-betaII (59%) protein expression were increased in 4-week diabetic aortas compared to controls. SIGNIFICANCE: In conclusion, diabetes led to a time-dependent increase in ouabain-sensitive Na(+), K(+)-ATPase activity. The main mechanism involved in this activation is the release of TxA(2)/PGH(2) by COX-2 in smooth muscle cells, linked to activation of the PKC pathway.