RESUMO
In vertebrates, connexins (Cxs) and pannexins (Panxs) are proteins that form gap junction channels and/or hemichannels located at cell-cell interfaces and cell surface, respectively. Similar channel types are formed by innexins in invertebrate cells. These channels serve as pathways for cellular communication that coordinate diverse physiologic processes. However, it is known that many acquired and inherited diseases deregulate Cx and/or Panx channels, condition that frequently worsens the pathological state of vertebrates. Recent evidences suggest that Cx and/or Panx hemichannels play a relevant role in bacterial and viral infections. Nonetheless, little is known about the role of Cx- and Panx-based channels in parasitic infections of vertebrates. In this review, available data on changes in Cx and gap junction channel changes induced by parasitic infections are summarized. Additionally, we describe recent findings that suggest possible roles of hemichannels in parasitic infections. Finally, the possibility of new therapeutic designs based on hemichannel blokers is presented.
Assuntos
Conexinas/metabolismo , Junções Comunicantes/metabolismo , Junções Comunicantes/parasitologia , Doenças Parasitárias/metabolismo , Animais , Infecções Bacterianas/metabolismo , Infecções Bacterianas/patologia , Junções Comunicantes/microbiologia , Junções Comunicantes/patologia , Junções Comunicantes/virologia , Humanos , Doenças Parasitárias/patologia , Viroses/metabolismo , Viroses/patologiaRESUMO
Antecedentes: En la isquemia/reperfusión (IR) miocárdica es relevante la pérdida de cardiomiocitos por apoptosis. En estos, los hemicanales (HC) permiten el ingreso de sustancias proapoptóticas durante la IR. Boldina (B), compuesto extraído del Peumus boldus, ha demostrado ser antioxidante y bloquear los HC. Objetivo: Determinar el efecto de boldina sobre la apoptosis de cardiomiocitos de ratas sometidas a IR. Métodos: Ratas macho de 200 g de peso se sometieron a ligadura reversible de la arteria coronaria izquierda por 30 minutos (I) y posterior reperfusión (R) por 24 horas post I. Un subgrupo de estos animales recibió una dosis de boldina intraventricular (IR+B, 40 mg/Kg) y luego dos dosis vía gavage (75 mg/Kg) a los 30 y 60 minutos post-R. Como controles se usaron ratas sham con operación ficticia, que recibieron igual tratamiento. Se determinaron las masas corporal (MC) y cardiaca relativa (MCR) y presión arterial sistólica (PAS). Porcentaje de cardiomiocitos apoptóticos (CMAP), otras células apoptóticas (OCAP) y total de células apoptóticas (TCAP) se determinó por TUNEL. La activación de metaloproteinasas (MMPs) 2 y 9 se determinó por zimografía y el mRNA de MCP-1 por RT-PCR. Resultados: La boldina no modificó la MC y la MCR. Sin embargo, disminuyó significativamente la PAS así como el por cientoCMAP y el por cientoTCAP en el grupo IR+B versus IR (CMAP 69 +/- 1,5 vs 44 ± 0,4, p=0,016, TCAP 71 +/- 2,4 vs 57 +/- 1,5, p=0,016). No se encontraron diferencias en el OCAP, actividad de MMPs y en los niveles de mRNA de MCP-1. Conclusiones: Boldina disminuyó la PAS y la apoptosis de cardiomiocitos post IR. Su efecto no es mediado por modificaciones en la actividad de MMPs y expresión génica de MCP-1.
Background: Ischemia / reperfusion (IR) is relevant in the myocardial loss of cardiomyocytes through apoptosis. During IR, hemi channels (HC) allow the entry of proapoptotic substances to the cell. Boldine, a compound extracted from Peumus boldus, has proven to be antioxidant and to block HC. Objective: To determine the effect of boldine on cardiomyocyte apoptosis in rats subjected to IR. Methods: Male rats, body weight (BW) 200 g, were subjected to reversible ligation of the left coronary artery for 30 minutes (I) and subsequent reperfusion (R) for 24 hours. A subset of these animals (IR+B) received an intraventricular dose of boldine (40mg/kg) and then two doses via gavage (75 mg/kg) at 30 and 60 minutes post-R. Sham operated rats (S) receiving the same treatment were used as controls. We determined body weight (BW), relative heart mass (RHM) and systolic blood pressure (SBP). Percentage of apoptotic cardiomyocytes (CMAP), other apoptotic cells (OCAP) and total apoptotic cells (TCAP) were determined by TUNEL. Activation of metalloproteinases (MMPs) 2 and 9 was determined by zymography and MCP-1 mRNA levels by RT-PCR. Results: Compared to IR alone, IR+Boldine did not change BW or RCM, but significantly decreased PAS, TCAP (71 +/- 2.4 vs 57 +/- 1.5, p=0.016) and CMAP (69 +/- 1.5 vs 44 +/- 0.4, p=0.016). No difference was observed in the OCAP, MMPs activity and MCP-1 mRNA levels. Conclusions: Boldine decreased SBP and post-IR cardiomyocyte apoptosis without effect on other cells. This effect was not mediated by MMPs activity or MCP-1 gene expression.
Assuntos
Masculino , Animais , Ratos , Antioxidantes/farmacologia , Apoptose , Aporfinas/farmacologia , Miócitos Cardíacos , Ratos Sprague-DawleyRESUMO
ATP released from cells is known to activate plasma membrane P2X (ionotropic) or P2Y (metabotropic) receptors. In skeletal muscle cells, depolarizing stimuli induce both a fast calcium signal associated with contraction and a slow signal that regulates gene expression. Here we show that nucleotides released to the extracellular medium by electrical stimulation are partly involved in the fast component and are largely responsible for the slow signals. In rat skeletal myotubes, a tetanic stimulus (45 Hz, 400 1-ms pulses) rapidly increased extracellular levels of ATP, ADP, and AMP after 15 s to 3 min. Exogenous ATP induced an increase in intracellular free Ca(2+) concentration, with an EC(50) value of 7.8 +/- 3.1 microm. Exogenous ADP, UTP, and UDP also promoted calcium transients. Both fast and slow calcium signals evoked by tetanic stimulation were inhibited by either 100 mum suramin or 2 units/ml apyrase. Apyrase also reduced fast and slow calcium signals evoked by tetanus (45 Hz, 400 0.3-ms pulses) in isolated mouse adult skeletal fibers. A likely candidate for the ATP release pathway is the pannexin-1 hemichannel; its blockers inhibited both calcium transients and ATP release. The dihydropyridine receptor co-precipitated with both the P2Y(2) receptor and pannexin-1. As reported previously for electrical stimulation, 500 mum ATP significantly increased mRNA expression for both c-fos and interleukin 6. Our results suggest that nucleotides released during skeletal muscle activity through pannexin-1 hemichannels act through P2X and P2Y receptors to modulate both Ca(2+) homeostasis and muscle physiology.
Assuntos
Trifosfato de Adenosina/metabolismo , Sinalização do Cálcio/fisiologia , Cálcio/metabolismo , Expressão Gênica , Músculo Esquelético/fisiologia , Animais , Apirase/farmacologia , Canais de Cálcio Tipo L/metabolismo , Linhagem Celular , Conexinas/genética , Conexinas/metabolismo , Estimulação Elétrica , Interleucina-6/genética , Interleucina-6/metabolismo , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Camundongos , Músculo Esquelético/citologia , Músculo Esquelético/efeitos dos fármacos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Cloreto de Potássio/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ratos , Receptores Purinérgicos P2/genética , Receptores Purinérgicos P2/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Suramina/farmacologiaRESUMO
A methodology is described and applied for performing carbon mass balances across cellulase enzyme production processes using both soluble sugar and insoluble cellulose substrates. The fungus Trichoderma reesei was grown on either glucose, lactose, or cellulose in aerobic batch mode, and the evolution of the main carbonaceous components (cell mass, cellulose, soluble protein, adsorbed protein, sugars, and carbon dioxide) was followed. A variety of analytical techniques were utilized to measure these components, including (i) gravimetric analysis, (ii) near-infrared spectroscopy, (iii) bicinchoninic acid based soluble protein measurement, (iv) gas mass spectrometry and flow rate, (v) CHNS/O elemental analyses, and (vi) high-performance liquid chromatography. The combined set of measurements allowed carbon mass balances across the cellulase production process to be assessed to determine the consistency of the underlying kinetic data. Results demonstrate the capability to determine the levels and distribution of all major carbonaceous components during the cellulase production process on both soluble and insoluble substrates. Average carbon mass balance closures were near 100% during early stages (<72 h) of the cultivations using glucose, lactose, or cellulose as the substrates, but carbon mass closures trended high later in the cultivation. Analysis of carbon allocation results suggests that an error in the gas mass flow rate measurement was the primary cause for carbon mass balance closures to exceed 110% late in the process.