Regulation of intestinal micro ecology between raw and salt-processed Alpinia oxyphylla on renal injury rats.

Alpinia oxyphylla Fructus is one of the traditional Chinese medicine plants in the treatment of kidney injury. In clinical practice, crude Alpinia oxyphylla Fructus (CAOF) and salt-processed Alpinia oxyphylla Fructus (SAOF) are the two commonly used drugs specificated in the prevention and treatment of diabetic nephropathy (DN). However, the intestinal micro ecology regulation between CAOF and SAOF on DN has not been reported. In this paper, intestinal micro ecology regulation activities between CAOF and SAOF in DN rats were compared and analyzed by short-chain fatty acids (SCFAs) and intestinal flora analysis. The results showed that both SAOF and CAOF can regulate the intestinal flora metabolite SCFAs level in DN rats, reduce blood glucose concentration and improve inflammatory reaction. The intestinal flora analysis showed SAOF and CAOF could increase the intestinal bacterial diversity. The treatment of renal injury may be related to their increased intestinal bacterial diversity.

Adrenomedullin induces heme oxygenase-1 gene expression and cGMP formation in rat vascular smooth muscle cells.

Adrenomedullin (ADM) is a potent vasodilatory peptide. It regulates blood pressure by increasing cyclic adenosine monophosphate (cAMP) and guanosine-3',5'-monophosphate (cGMP). We sought to investigate the effect of ADM on heme oxygenase-1 (HO-1) gene expression and cGMP formation in cultured rat vascular smooth muscle cells (VSMCs). ADM treatment, 10(-9) and 10(-8) mol/L, increased cGMP production, and it increased the intracellular cGMP content of platelets coincubated with VSMCs. It increased cGMP content by 158.8% and 273.5%, respectively; increased HO-1 activity by 49.5% and 87%, respectively; augmented HO-1 protein levels by 66% and 126%, respectively; upregulated the steady-state level of HO-1 mRNA by 73% and 159%, respectively, and increased HO-1 mRNA transcription synthesis by four- and seven-fold, respectively. These results suggest that ADM induces HO-1 gene expression and cGMP formation in rat VSMCs.

Transcriptional regulation of cardiac sarcoplasmic reticulum calcium-ATPase gene during the progression of sepsis.

Changes in sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) gene expression in the rat heart during different phases of sepsis were studied. Sepsis was induced by cecal ligation and puncture (CLP). Septic rats were divided into two groups: the early hyperdynamic (9 h after CLP, early sepsis) and the late hypodynamic (18 h after CLP; late sepsis) groups. Western blot analyses reveal that SERCA2a protein level remained unaltered during early sepsis but was decreased by 59% during late sepsis. Northern blot analyses show that the steady-state level of SERCA2a mRNA stayed unchanged during the early phase but was decreased by 43% during the late phase of sepsis. Nuclear runoff assays show that the transcription rate of SERCA2a gene transcript remained unaffected during early sepsis but was decreased by 34% during late sepsis. The actinomycin D pulse-chase studies indicate that the half-life of SERCA2a mRNA was unaffected during the early and the late phases of sepsis. These findings demonstrate that during the early phase of sepsis, the protein level, the mRNA abundance, and the transcription rate of SERCA2a remained unaltered, whereas during the late phase of sepsis, the rate of transcription of SERCA2a was decreased, and the decreased transcription rate was associated with decreases in SERCA2a mRNA abundance and SERCA2a protein level in the rat heart. Based on these data, it is concluded that SERCA2a gene expression decreased during the late phase of sepsis in the rat heart and that the decreased expression was regulated at the transcriptional level.

G protein and adenylate cyclase complex-mediated signal transduction in the rat heart during sepsis.

Changes in the protein level of various subunits of GTP-binding protein and the activity of adenylate cyclase in the rat heart during different phases of sepsis were studied. Sepsis was induced by cecal ligation and puncture (CLP). Experiments were divided into three groups: control, early sepsis, and late sepsis. Early and late sepsis refers to those animals sacrificed at 9 and 18 h, respectively, after CLP. The protein levels of various subunits of GTP-binding protein were determined by Western blot analysis. The activity of adenylate cyclase was measured based on the rate of formation of cAMP from [alpha-32P]ATP. The results show that protein levels of G alphas and G beta remained stable during the early and the late phases of sepsis. The protein levels of G alpha i-2 and G alpha i-3 remained relatively unaltered during the early phase of sepsis, but they were increased by 46.5% (P < 0.05) and 61.3% (P < 0.01), respectively, during the late phase of sepsis. The basal adenylate cyclase activity remained unchanged during the early phase while it was decreased by 25.7% (P < 0.05) during the late phase of sepsis. The isoproterenol-stimulated adenylate cyclase activity was unchanged during early sepsis while it was decreased by 44.6% (P < 0.01) during late sepsis. These data demonstrate that during the late hypodynamic phase of sepsis, myocardial G alpha i-2 and G alpha i-3 protein levels were increased and the increases were coupled with a reduction in adenylate cyclase activity. Because GTP-binding proteins mediate sympathetic control of cardiac function, the present findings may have a pathophysiological significance in contributing to the understanding of the pathogenesis of cardiac dysfunction during the late stage of sepsis.

Altered phospholamban-calcium ATPase interaction in cardiac sarcoplasmic reticulum during the progression of sepsis.

The purpose of this study was to investigate alterations of phospholamban phosphorylation and its interaction with Ca2+ transport(Ca2+-ATPase activity and Ca2+ uptake) in sarcoplasmic reticulum (SR) during the progression of sepsis. Sepsis was induced by cecal ligation and puncture (CLP). Phospholamban phosphorylation was studied by the labeling of the myocardial ATP pool by perfusing isolated rat hearts with [32P]H3PO4 followed by identification of the phosphorylated phospholamban. Results show that phospholamban phosphorylation was increased by 153% during the early hyperdynamic phase (9 h after CLP), while it was decreased by 51% during the late hypodynamic phase (18 h after CLP) of sepsis. The increase in phospholamban phosphorylation during early sepsis was associated with increases in +dP/dt(max) and tissue cAMP content, while Ca2+ transport, left ventricular developed pressure (LVDP), and -dP/dt(max) remained unchanged. The decrease in phospholamban phosphorylation during late sepsis was accompanied by decreases in Ca2+ transport, LVDP, +/-dP/dt(max), and tissue cAMP content. When isoproterenol was present in the perfusion medium, all parameters measured were stimulated in all three experimental groups (control, early sepsis, and late sepsis) except that Ca2+-ATPase activity and SR Ca2+ uptake were unresponsive in the early and the late septic groups. These findings demonstrate that during the late hypodynamic phase of sepsis, the observed decrease in myocardial contractility was due to the decrease in phospholamban phosphorylation, which resulted in decreased Ca2+ transport across the SR. In contrast, during the early hyperdynamic phase of sepsis, the increase in phospholamban phosphorylation did not correlate with increases in Ca2+ uptake and Ca2+-ATPase activity. Thus, the interaction between phospholamban phosphorylation and Ca2+ transport across the SR was disrupted during the early phase of sepsis.

[Comparison of ryanodine binding to cardiac sarcoplasmic reticulum and nuclear envelope of rat].

AIM: The characteristics of ryanodine receptor in rat cardiac sarcoplasmic reticulum (SR) and nuclear envelope (NE) were studied. METHODS: Velocity and isopyknic gradient centrifugation was employed to fractionate rat SR and NE. Ryanodine receptor was assayed with [3H] ryanodine saturate binding to the preparations. RESULTS: The maximal binding (Bmax) and dissociating constant (Kd) of ryanodine receptor in rat cardiac NE were, 1.7% and 60% of those in SR respectively. Phosphorylation in vitro by PKA and PKC increased Bmax of the receptors in SR by 372% and 121%, and augmented those in NE by 221% and 306%, without any effects on Kd. CONCLUSION: Ryanodine receptors were present in rat myocardial NE, with lower density and higher affinity than those located in SR, which can be activated by PKA and PKC.

Group II Phospholipase A(2) Activity and Its Molecular Regulation in Rat Heart during Sepsis.

Changes of activity and content of myocardial group II phospholipase A(2) and its mRNA transcription and stability during rat sepsis were investigated. Results showed that, compared with control group,myocardial group II phospholipase A(2) activity in early and late sepsis decreased by 25.0%(P<0.05) and increased by 47.6%(P<0.01),respectively group II phospholipase A(2) protein concentration reduced by 27.0% and augmented by 48.0%(P<0.01),respectively. Myocardial group II phospholipase A(2) mRNA transcription rate and content showed similar two-phases changes. The mRNA transcription rate during early and late sepsis decreased by 45.0% and increased by 70.0%(P<0.01),respectively. The mRNA content decreased by 34.1% in early sepsis and increased by 157.0% in late sepsis(P<0.01), respectively. The half-life of group II phospholipase A(2) mRNA remained unchanged notably during early and late stage of sepsis. These data suggest that myocardial group II phospholipase A(2) activity decreased in early stage of sepsis and increased in its late stage, and these changes were regulated transcriptionally.