RESUMO
It has been demonstrated that supplementation with the two main omega 3 polyunsaturated fatty acids (ω3 FAs), docosahexaenoic acid (DHA), and eicosapentaenoic acid (EPA), leads to modifications in the cardiac physiology. ω3 FAs can affect the membrane's lipid composition, as well as proteins' location and/or function. The Na+/H+ exchanger (NHE1) is an integral membrane protein involved in the maintenance of intracellular pH and its hyperactivity has been associated with the development of various cardiovascular diseases such as cardiac hypertrophy. Our aim was to determine the effect of ω3 FAs on systolic blood pressure (SBP), lipid profiles, NHE1 activity, and cardiac function in spontaneously hypertensive rats (SHR) using Wistar rats (W) as normotensive control. After weaning, the rats received orally ω3 FAs (200 mg/kg body mass/day/ 4 months). We measured SBP, lipid profiles, and different echocardiography parameters, which were used to calculate cardiac hypertrophy index, systolic function, and ventricular geometry. The rats were sacrificed, and ventricular cardiomyocytes were obtained to measure NHE1 activity. While the treatment with ω3 FAs did not affect the SBP, lipid analysis of plasma revealed a significant decrease in omega-6/omega-3 ratio, correlated with a significant reduction in left ventricular mass index in SHR. The NHE1 activity was significantly higher in SHR compared with W. While in W the NHE1 activity was similar in both groups, a significant decrease in NHE1 activity was detected in SHRs supplemented with ω3 FAs, reaching values comparable with W. Altogether, these findings revealed that diet supplementation with ω3 FAs since early age prevents the development of cardiac hypertrophy in SHR, perhaps by decreasing NHE1 activity, without altering hemodynamic overload.
Assuntos
Ácidos Graxos Ômega-3 , Ratos , Animais , Ratos Wistar , Ácidos Graxos Ômega-3/farmacologia , Ácidos Docosa-Hexaenoicos , Ácido Eicosapentaenoico , Ratos Endogâmicos SHR , Cardiomegalia/tratamento farmacológico , Cardiomegalia/prevenção & controleRESUMO
The Membrane Attack Complex-Perforin (MACPF) family is ubiquitously found in all kingdoms. They have diverse cellular roles, however MACPFs with pore-forming toxic function in venoms and poisons are very rare in animals. Here we present the structure of PmPV2, a MACPF toxin from the poisonous apple snail eggs, that can affect the digestive and nervous systems of potential predators. We report the three-dimensional structure of PmPV2, at 17.2 Å resolution determined by negative-stain electron microscopy and its solution structure by small angle X-ray scattering (SAXS). We found that PV2s differ from nearly all MACPFs in two respects: it is a dimer in solution and protomers combine two immune proteins into an AB toxin. The MACPF chain is linked by a single disulfide bond to a tachylectin chain, and two heterodimers are arranged head-to-tail by non-covalent forces in the native protein. MACPF domain is fused with a putative new Ct-accessory domain exclusive to invertebrates. The tachylectin is a six-bladed ß-propeller, similar to animal tectonins. We experimentally validated the predicted functions of both subunits and demonstrated for the first time that PV2s are true pore-forming toxins. The tachylectin "B" delivery subunit would bind to target membranes, and then the MACPF "A" toxic subunit would disrupt lipid bilayers forming large pores altering the plasma membrane conductance. These results indicate that PV2s toxicity evolved by linking two immune proteins where their combined preexisting functions gave rise to a new toxic entity with a novel role in defense against predation. This structure is an unparalleled example of protein exaptation.
Assuntos
Complexo de Ataque à Membrana do Sistema Complemento/ultraestrutura , Lectinas/ultraestrutura , Perforina/ultraestrutura , Conformação Proteica , Sequência de Aminoácidos/genética , Animais , Membrana Celular/química , Membrana Celular/ultraestrutura , Complexo de Ataque à Membrana do Sistema Complemento/química , Complexo de Ataque à Membrana do Sistema Complemento/imunologia , Cristalografia por Raios X , Dimerização , Lectinas/química , Lectinas/imunologia , Modelos Moleculares , Perforina/química , Perforina/imunologia , Subunidades Proteicas/genética , Espalhamento a Baixo Ângulo , Caramujos/ultraestrutura , Difração de Raios XRESUMO
This study was performed to determine whether fatty acids incorporated into liver cell nuclei phosphatidylcholine (PtdCho) could be remodeled in the isolated nuclear. For this reason, rat liver cell nuclei were incubated in vitro with [1-14C]20:4n-6-CoA. PtdCho molecular species with the highest specific activity had an unsaturated fatty acid at sn-1 and sn-2 positions (20:4-20:4>18:2-20:4>18:1-20:4). 16:0-20:4 and 18:0-20:4 PtdChos showed a minor specific activity. When labeled nuclei were reincubated in the absence of labeled substrate with the addition of cytosol, ATP and CoA, the specific activity of 20:4-20:4, 18:2-20:4 and 18:1-20:4 species decreased, while that of 16:0-20:4 and 18:0-20:4 increased. In conclusion, the asymmetric fatty acid distribution of saturated fatty acids at sn-1 position, and unsaturated fatty acids at sn-2 position of nuclear PtdCho molecular species was re-established by an acyl-CoA-dependent remodeling process.
Assuntos
Núcleo Celular/metabolismo , Hepatócitos/citologia , Hepatócitos/metabolismo , Fosfatidilcolinas/metabolismo , Animais , Ácido Araquidônico/metabolismo , Núcleo Celular/química , Citosol/química , Citosol/metabolismo , Masculino , Fosfatidilcolinas/química , Ratos , Ratos WistarRESUMO
Liver nuclear incorporation of stearic (18:0), linoleic (18:2n-6), and arachidonic (20:4n-6) acids was studied by incubation in vitro of the [1-14C] fatty acids with nuclei, with or without the cytosol fraction at different times. The [1-14C] fatty acids were incorporated into the nuclei as free fatty acids in the following order: 18:0 > 20:4n-6 >> 18:2n-6, and esterified into nuclear lipids by an acyl-CoA pathway. All [1-14C] fatty acids were esterified mainly to phospholipids and triacylglycerols and in a minor proportion to diacylglycerols. Only [1-14C]18:2n-6-CoA was incorporated into cholesterol esters. The incorporation was not modified by cytosol addition. The incorporation of 20:4n-6 into nuclear phosphatidylcholine (PC) pools was also studied by incubation of liver nuclei in vitro with [1-14C]20:4n-6-CoA, and nuclear labeled PC molecular species were determined. From the 15 PC nuclear molecular species determined, five were labeled with [1-14C]20:4n-6-CoA: 18:0-20:4, 16:0-20:4, 18:1-20:4, 18:2-20:4, and 20:4-20:4. The highest specific radioactivity was found in 20:4-20:4 PC, which is a minor species. In conclusion, liver cell nuclei possess the necessary enzymes to incorporate exogenous saturated and unsaturated fatty acids into lipids by an acyl-CoA pathway, showing specificity for each fatty acid. Liver cell nuclei also utilize exogenous 20:4n-6-CoA to synthesize the major molecular species of PC with 20:4n-6 at the sn-2 position. However, the most actively synthesized is 20:4-20:4 PC, which is a quantitatively minor component. The labeling pattern of 20:4-20:4 PC would indicate that this molecular species is synthesized mainly by the de novo pathway.