Neuropeptide Y reverses chronic stress-induced baroreflex hypersensitivity in rats

Chronic stress, as a risk factor for cardiovascular diseases, has been reported to result in elevated plasma neuropeptide Y (NPY) and be highly associated with abnormal cardiac autonomic function. This study aimed to explore the effect of NPY on the chronic stress-induced abnormal baroreceptor reflex sensitivity (BRS). Seven types of recognized stressors were used to develop chronic stress rat model. Subcutaneously implanting ALZET mini-osmotic pumps containing NPY were used to evaluate the action of NPY on the stressed male rats. We found that chronic stress showed no influence on baseline systolic blood pressure (SBP) and heart rate (HR), whereas NPY (85 μg for 30 days) could elevate baseline SBP and induce bradycardia in rats intervened by various stimuli. NPY pretreatment could preserve chronic stress-induced decreases in left ventricular systolic pressure (LVSP) and the maximum rate of change in left ventricular pressure in the isovolumic contraction period (+dp/dt(max)) but has shown no effect on left ventricular end diastolic pressure (LVEDP) and the maximum rate of change in left ventricular pressure in the isovolumic relaxation period (-dp/dt(max)). Notably, chronic stress led to baroreflex oversensitivity indicated by the elevated ratio of Δheart rate (HR)/ Δmean arterial blood pressure (MABP) in rats followed by vasoconstrictor (phenylephrine, PE) or vasodilator (sodium nitroprusside, SNP) administration, which was almost completely reversed by NPY pretreatment. The expressions of substance P (SP) and gamma aminobutyric acid A receptor (GABA(A)R) in nucleus tractus solitarius were increased in chronic stress rats, which were counteracted by NPY pretreatment. We conclude that chronic stress-induced baroreflex hypersensitivity could be blocked by NPY pretreatment. Furthermore, the altered expressions of neurotransmitters and receptors in the brainstem might contribute to this process.     

A novel anti-Cyr61 antibody inhibits breast cancer growth and metastasis in vivo

Cysteine-rich protein 61(CCN1/Cyr61) has been implicated as an important mediator in proliferation and metastasis of breast cancer, which indicated that blockage of Cyr61 might be a potent target for breast cancer treatment. However, the antitumor effect of anti-Cyr61 antibodies on breast cancer in vivo has not been reported so far. In this study, we reported the effect and likely mechanism of generated anti-human Cyr61 monoclonal antibodies (mAb) on Cyr61 high expression line MDA-MB-231, known as a highly malignant and invasive human breast cancer cell line, at aspects of proliferation and migration in vitro and in vivo. We found the mAb, denoted as 093G9, revealed inhibitory effects on MDA-MB-231 cell proliferation, migration, and invasion through downregulation of both AKT and ERK phosphorylation in vitro compared with its isotype control. 093G9 also showed significant efficacy on suppressing primary tumor growth and spontaneous lymph node metastasis in in vivo mouse model. The specific epitope recognized by 093G9 was identified to be ¹⁴⁰LPNLGCP¹⁴⁶, adjacent to the VWC domain of Cyr61 by Ph.D.-C7C phage library display system. Our study provides direct evidence that Cyr61 can be a potent therapeutic target for patients who bear high Cyr61 expression breast cancer. Furthermore, the mAb, 093G9 developed in our laboratory, has shown a promising therapeutic characteristic in breast cancer.     

Caveolin-1-mediated negative signaling plays a critical role in the induction of regulatory dendritic cells by DNA and protein coimmunization

Induction of Ag-specific regulatory T cells (iTregs) by vaccination is a promising strategy for treating autoimmune diseases. We previously demonstrated that DNA and protein covaccination converted naive T cells to Ag-specific iTregs by inducing CD11c(+)CD40(low)IL-10(+) regulatory dendritic cells (DCregs). However, it is unclear how coimmunization induces the DCregs. In this paper, we report that the event is initiated by coentry of sequence-matched DNA and protein immunogens into the same DC via caveolae-mediated endocytosis, which leads to inhibition of phosphorylation of caveolin-1 (Cav-1), the main component of caveolae, and upregulation of Tollip. This triggers downstream signaling that upregulates suppressor of cytokine signaling 1 and downregulates NF-κB and STAT-1α. Silencing either Cav-1 or Tollip blocks the negative signaling, leading to upregulated expression of CD40, downregulated production of IL-10, and loss of iTreg-inducing function. We further show that DCregs can be induced in culture from primary DCs and JAWS II DC lines by feeding them sequence-matched DNA and protein immunogens. The in vitro-generated DCregs are effective in ameliorating autoimmune and inflammatory diseases in several mouse models. Our study thus suggests that DNA and protein coimmunization induces DCregs through Cav-1- and Tollip-mediated negative signaling. It also describes a novel method for generating therapeutic DCregs in vitro.     

Structure-activity relationship of a cold-adapted purine nucleoside phosphorylase by site-directed mutagenesis

Purine nucleoside phosphorylase can be expressed in Escherichia coli and the intact cells can be used as a catalyst for the biosynthesis of nucleosides. The purine nucleoside phosphorylases from E. coli (EcPNP) and Pseudoalteromonas sp. XM2107 (PsPNP) have been purified. In order to improve the catalytic efficiency, the model of three-dimensional structure of PsPNP was constructed, and then 9 active/binding-site mutants were constructed by one-step site-directed mutagenesis and characterized by steady-state kinetics. Double mutations exhibited the largest change of catalytic activity. The T90R:T156S mutant revealed 1000 fold enhancements in k(cat)/K(m) for inosine phosphorolysis. However, the T90A:T156A mutant revealed 500 fold reduction in catalytic activity when compared with wild-type one. These results in combination with the predicted locations of Thr90 and Thr156 side chains by homology modeling suggested that: (i) a complete hydrophobic pocket played an important role in the catalytic function of PsPNP; (ii) a potential transition state structure was present in hydrogen bond between the carboxyl groups of Thr90 in the phosphate binding site. Therefore, the application of site-directed mutagenesis will be benefit to further improve catalytic efficiency of PsPNP during the enzymatic synthesis of antivirus drug ribavirin.     

Biaxial vasoactivity of porcine coronary artery

The passive mechanical properties of blood vessel mainly stem from the interaction of collagen and elastin fibers, but vessel constriction is attributed to smooth muscle cell (SMC) contraction. Although the passive properties of coronary arteries have been well characterized, the active biaxial stress-strain relationship is not known. Here, we carry out biaxial (inflation and axial extension) mechanical tests in right coronary arteries that provide the active coronary stress-strain relationship in circumferential and axial directions. Based on the measurements, a biaxial active strain energy function is proposed to quantify the constitutive stress-strain relationship in the physiological range of loading. The strain energy is expressed as a Gauss error function in the physiological pressure range. In K(+)-induced vasoconstriction, the mean ± SE values of outer diameters at transmural pressure of 80 mmHg were 3.41 ± 0.17 and 3.28 ± 0.24 mm at axial stretch ratios of 1.3 and 1.5, respectively, which were significantly smaller than those in Ca(2+)-free-induced vasodilated state (i.e., 4.01 ± 0.16 and 3.75 ± 0.20 mm, respectively). The mean ± SE values of the inner and outer diameters in no-load state and the opening angles in zero-stress state were 1.69 ± 0.04 mm and 2.25 ± 0.08 mm and 126 ± 22°, respectively. The active stresses have a maximal value at the passive pressure of 80-100 mmHg and at the active pressure of 140-160 mmHg. Moreover, a mechanical analysis shows a significant reduction of mean stress and strain (averaged through the vessel wall). These findings have important implications for understanding SMC mechanics.     

Unusual carbon fixation gives rise to diverse polyketide extender units

Polyketides are structurally diverse and medically important natural products that have various biological activities. During biosynthesis, chain elongation uses activated dicarboxylic acid building blocks, and their availability therefore limits side chain variation in polyketides. Recently, the crotonyl-CoA carboxylase-reductase (CCR) class of enzymes was identified in primary metabolism and was found to be involved in extender-unit biosynthesis of polyketides. These enzymes are, in theory, capable of forming dicarboxylic acids that show any side chain from the respective unsaturated fatty acid precursor. To our knowledge, we here report the first crystal structure of a CCR, the hexylmalonyl-CoA synthase from Streptomyces sp. JS360, in complex with its substrate. Structural analysis and biochemical characterization of the enzyme, including active site mutations, are reported. Our analysis reveals how primary metabolic CCRs can evolve to produce various dicarboxylic acid building blocks, setting the stage to use CCRs for the production of unique extender units and, consequently, altered polyketides.     

Modeling Impacts of Alternative Practices on Net Global Warming Potential and Greenhouse Gas Intensity from Rice–Wheat Annual Rotation in China

Background

Evaluating the net exchange of greenhouse gas (GHG) emissions in conjunction with soil carbon sequestration may give a comprehensive insight on the role of agricultural production in global warming.

Materials and Methods

Measured data of methane (CH₄) and nitrous oxide (N₂O) were utilized to test the applicability of the Denitrification and Decomposition (DNDC) model to a winter wheat – single rice rotation system in southern China. Six alternative scenarios were simulated against the baseline scenario to evaluate their long-term (45-year) impacts on net global warming potential (GWP) and greenhouse gas intensity (GHGI).

Principal Results

The simulated cumulative CH₄ emissions fell within the statistical deviation ranges of the field data, with the exception of N₂O emissions during rice-growing season and both gases from the control treatment. Sensitivity tests showed that both CH₄ and N₂O emissions were significantly affected by changes in both environmental factors and management practices. Compared with the baseline scenario, the long-term simulation had the following results: (1) high straw return and manure amendment scenarios greatly increased CH₄ emissions, while other scenarios had similar CH₄ emissions, (2) high inorganic N fertilizer increased N₂O emissions while manure amendment and reduced inorganic N fertilizer scenarios decreased N₂O emissions, (3) the mean annual soil organic carbon sequestration rates (SOCSR) under manure amendment, high straw return, and no-tillage scenarios averaged 0.20 t C ha⁻¹ yr⁻¹, being greater than other scenarios, and (4) the reduced inorganic N fertilizer scenario produced the least N loss from the system, while all the scenarios produced comparable grain yields.

Conclusions

In terms of net GWP and GHGI for the comprehensive assessment of climate change and crop production, reduced inorganic N fertilizer scenario followed by no-tillage scenario would be advocated for this specified cropping system.