Possible role of the CYC2c gene in the cornflower-like ray floret phenotype of Gaillardia cultivars

Journal of Plant Research - Gaillardia plants have been widely cultivated in China and have become an important component of garden landscaping. Different from the common ligulate ray floret, the...     

Interplay between formation of photosynthetic complexes and expression of genes for iron–sulfur cluster assembly in Rhodobacter sphaeroides?

Photosynthesis Research - Formation of photosynthetic complexes leads to a higher demand for Fe–S clusters. We hypothesized that in the facultative phototrophic alpha-proteobacterium...     

TNF-α induces upregulation of EGFR expression and signaling in human colonic myofibroblasts

The myofibroblast has recently been identified as an important mediator of tumor necrosis factor-α (TNF-α)-associated colitis and cancer, but the mechanism(s) involved remains incompletely understood. Recent evidence suggests that TNF-α is a central regulator of multiple inflammatory signaling cascades. One important target of TNF-α may be the signaling pathway downstream of the epidermal growth factor receptor (EGFR), which has been associated with many human cancers. Here, we show that long-term exposure of 18Co cells, a model of human colonic myofibroblasts, with TNF-α led to a striking increase in cell surface EGFR expression, an effect that was completely inhibited by cycloheximide. Subsequent EGFR binding by EGF and heparin binding (HB)-EGF was associated with enhanced EGFR tyrosine kinase activity, prolonged ERK activation, and a significant increase in cyclooxygenase-2 (COX-2) expression compared with 18Co cells treated with EGF and HB-EGF alone. TNF-α also increased EGFR expression and signaling in primary myofibroblasts isolated from human colon tissue. TNF-α-induced upregulation of EGFR may be a plausible mechanism to explain the exaggerated cellular responsiveness that characterizes inflammatory bowel disease and that may contribute to a microenvironment that predisposes to colitis-associated cancer through enhanced COX-2 expression.     

Impaired thymic export and apoptosis contribute to regulatory T-cell defects in patients with chronic heart failure

Objective

Animal studies suggest that regulatory T (T_reg) cells play a beneficial role in ventricular remodeling and our previous data have demonstrated defects of T_reg cells in patients with chronic heart failure (CHF). However, the mechanisms behind T_reg-cell defects remained unknown. We here sought to elucidate the mechanism of T_reg-cell defects in CHF patients.

Methods and Results

We performed flow cytometry analysis and demonstrated reduced numbers of peripheral blood CD4⁺CD25⁺FOXP3⁺CD45RO⁻CD45RA⁺ naïve T_reg (nT_reg) cells and CD4⁺CD25⁺FOXP3⁺CD45RO⁺CD45RA⁻ memory T_reg (mT_reg) cells in CHF patients as compared with non-CHF controls. Moreover, the nT_reg/mT_reg ratio (p<0.01), CD4⁺CD25⁺FOXP3⁺CD45RO⁻ CD45RA⁺CD31⁺ recent thymic emigrant T_reg cell (RTE-T_reg) frequency (p<0.01), and T-cell receptor excision circle levels in T_reg cells (p<0.01) were lower in CHF patients than in non-CHF controls. Combined annexin-V and 7-AAD staining showed that peripheral T_reg cells from CHF patients exhibited increased spontaneous apoptosis and were more prone to interleukin (IL)-2 deprivation- and CD95 ligand-mediated apoptosis than those from non-CHF individuals. Furthermore, analyses by both flow cytometry and real-time polymerase chain reaction showed that T_reg-cell frequency in the mediastinal lymph nodes or Foxp3 expression in hearts of CHF patients was no higher than that of the non-CHF controls.

Conclusion

Our data suggested that the T_reg-cell defects of CHF patients were likely caused by decreased thymic output of nascent T_reg cells and increased susceptibility to apoptosis in the periphery.     

Magnetostructural correlations in μ-pyrazolato-μ-acetato dinickel(II) complexes: Subtle effects of acetate tilting

A series of dinickel acetato complexes [LNi₂(OAc)(solvent)_x](ClO₄)₂ (1-3, 5) as well as related complexes [LNi₂(OAc)₂](OAc) (4) and [LNi₂(OAc)(NO₃)₂] (6), all derived from three pyrazolate-based binucleating ligands, have been prepared and characterized by X-ray crystallography. The solid state structures reveal different acetate binding modes (μ_1,3-bridging and bidentate chelating) plus severe twisting and tilting of the μ_1,3-acetate bridges with respect to their bimetallic scaffolds, reflecting the great flexibility of carboxylate coordination. Magnetic properties of all six complexes have been investigated, and the strength of antiferromagnetic coupling is discussed in the light of the structural differences, suggesting a magnetostructural correlation for acetato-bridged complexes.     

Low-Dose Prospectively Electrocardiogram-Gated Axial Dual-Source CT Angiography in Patients with Pulsatile Bilateral Bidirectional Glenn Shunt: An Alternative Noninvasive Method for Postoperative Morphological Estimation

Objective

To explore the clinical value of low-dose prospectively electrocardiogram-gated axial dual-source CT angiography (low-dose PGA scanning, CTA) in patients with pulsatile bilateral bidirectional Glenn shunt (bBDG) as an alternative noninvasive method for postoperative morphological estimation.

Methods

Twenty patients with pulsatile bBDG (mean age 4.2±1.6 years) underwent both low-dose PGA scanning and conventional cardiac angiography (CCA) for the morphological changes. The morphological evaluation included the anatomy of superior vena cava (SVC) and pulmonary artery (PA), the anastomotic location, thrombosis, aorto-pulmonary collateral circulation, pulmonary arteriovenous malformations, etc. Objective and subjective image quality was assessed. Bland–Altman analysis and linear regression analyses were used to evaluate the correlation on measurements between CTA and CCA. Effective radiation dose of both modalities was calculated.

Results

The CT attenuation value of bilateral SVC and PA was higher than 300 HU. The average subjective image quality score was 4.05±0.69. The morphology of bilateral SVC and PA was displayed completely and intuitively by CTA images. There were 24 SVC above PA and 15 SVC beside PA. Thrombosis was found in 1 patient. Collateral vessels were detected in 13 patients. No pulmonary arteriovenous malformation was found in our study. A strong correlation (R²>0.8, P<0.001) was observed between the measurements on CTA images and on CCA images. Bland–Altman analysis demonstrated a systematic overestimation of the measurements by CTA (the mean value of bias>0).The mean effective dose of CTA and CCA was 0.50±0.17 mSv and 4.85±1.34 mSv respectively.

Conclusion

CT angiography with a low-dose PGA scanning is an accurate and reliable noninvasive examination in the assessment of morphological changes in patients with pulsatile bBDG.     

Cell cycle-dependent expression of volume-activated chloride currents in nasopharyngeal carcinoma cells

Patch-clamping and cell imageanalysis techniques were used to study the expression of thevolume-activated Cl current,I_Cl(vol), and regulatory volume decrease (RVD)capacity in the cell cycle in nasopharyngeal carcinoma cells (CNE-2Z). Hypotonic challenge caused CNE-2Z cells to swell and activated aCl current with a linear conductance, negligibletime-dependent inactivation, and a reversal potential close to theCl equilibrium potential. The sequence of anionpermeability was I > Br > Cl > gluconate. The Cl channelblockers tamoxifen, 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB),and ATP inhibited I_Cl(vol). Synchronous cultures of cells were obtained by the mitotic shake-off technique and by adouble chemical-block (thymidine and hydroxyurea) technique. Theexpression of I_Cl(vol) was cell cycle dependent,being high in G₁ phase, downregulated in S phase, butincreasing again in M phase. Hypotonic solution activated RVD, whichwas cell cycle dependent and inhibited by the Cl channelblockers NPPB, tamoxifen, and ATP. The expression of I_Cl(vol) was closely correlated with the RVDcapacity in the cell cycle, suggesting a functional relationship.Inhibition of I_Cl(vol) by NPPB (100 µM)arrested cells in G₀/G₁. The data also suggest that expression of I_Cl(vol) and RVD capacity areactively modulated during the cell cycle. The volume-activatedCl current associated with RVD may therefore play animportant role during the cell cycle progress.

     

NMR Conformational and Dynamic Consequences of a Gly to Ser Substitution in an Osteogenesis Imperfecta Collagen Model Peptide

Close packing of three chains in a standard collagen triple helix requires Gly as every third residue. Missense mutations replacing one Gly by a larger residue in the tripeptide repeating sequence in type I collagen are common molecular causes of osteogenesis imperfecta. The structural and dynamic consequences of such mutations are addressed here by NMR studies on a peptide with a Gly-to-Ser substitution within an α1(I) sequence. Distances derived from nuclear Overhauser effects indicate that the three Ser residues are still packed in the center of the triple helix and that the standard 1-residue stagger is maintained. NMR dynamics using H-exchange and temperature-dependent amide chemical shifts indicate a greater disruption of hydrogen bonding and/or increased conformational flexibility C-terminal to the Ser site when compared with N terminal. This is consistent with recent suggestions relating clinical severity with an asymmetric effect of residues N- versus C-terminal to a mutation site. Dynamic studies also indicate that the relative position between a Gly in one chain and the mutation site in a neighboring staggered chain influences the disruption of the standard hydrogen-bonding pattern. The structural and dynamic alterations reported here may play a role in the etiology of osteogenesis imperfecta by affecting collagen secretion or interactions with other matrix molecules.Mutations in collagen result in a variety of connective tissue diseases (1, 2), with the clinical phenotype depending on the location and function of the collagen type. For instance, mutations in type I collagen, the major collagen in bone, lead to a bone disorder, osteogenesis imperfecta (OI),³ whereas mutations in type III collagen, which is present in high amounts in blood vessels, lead to aortic rupture in Ehlers-Danlos syndrome type IV (1, 2). All collagens have a triple helix motif composed of three polyproline II-like chains that are staggered by 1 residue and supercoiled about a common axis. The smallest residue Gly is typically present as every 3rd residue in each chain because of the tight packing of the chains, which generates the characteristic (Gly-Xaa-Yaa)_n repeating sequence. The Gly residues are all buried in the center, and the structure is stabilized by interchain N–H (Gly) … CO (Xaa) hydrogen bonds (3–5). The most common type of mutation leading to collagen disorders is a missense mutation that replaces 1 Gly in the repeating sequence by a larger residue.The best characterized collagen disease is OI, or brittle bone disease, which is distinguished by fragile bones due to mutations in type I collagen (2, 6). More than 400 Gly substitution missense mutations in the α1(I) and α2(I) chains of type I collagen have been reported to lead to OI (7). The severity of the disease varies widely from mild cases with multiple fractures to perinatal lethal cases (2, 6, 7). A single base change in a Gly codon can lead to one of 8 residues (Ser, Ala, Cys, Val, Arg, Asp, Glu, Trp) or a missense mutation. The smallest residue Ala is underrepresented in OI, suggesting that it may not always lead to pathology, whereas Ser mutations are overrepresented, corresponding to the most common substitutions observed. The 152 mutations leading to a Gly to Ser substitution account for ∼39% of all missense mutations in the α1(I) of type I collagen (7), with 115 associated with mild phenotypes and 37 associated with lethal phenotypes.The identity of the residue replacing Gly may be a determinant in the clinical severity of OI. Model peptide studies indicate that the degree of triple helix destabilization depends on the residue replacing Gly, with a ranking of the least destabilizing to the most destabilizing Ala,Ser8). There is some correlation between clinical severity of OI cases and this destabilization scale, with the strongly destabilizing residues Val, Arg, Asp, and Glu associated largely with lethal phenotypes (8). However, as cited above, a Gly to Ser mutation can lead to a mild, a severe, or a lethal OI case, with no obvious molecular explanation. Other factors suggested to contribute to clinical phenotype include the rigidity of its immediate sequence environment; its location with respect to the C terminus; its proximity to salt bridges; and its presence at an interaction site, such as the binding site for proteoglycans on collagen fibrils (7, 9). A recent study of the stability of OI collagens supported the importance of the domain location of the mutation (10), whereas a network analysis of the mutations suggested the importance of a destabilizing tripeptide sequence C-terminal to the mutation site (11).The standard triple helix conformation must undergo some structural perturbation as a result of a Gly replacement that is likely to relate to the development of the disorder. Thus it is important to define the structural consequences of a Gly substitution. It has not proved possible to obtain molecular information for the long collagen molecules themselves, but model collagen peptides have proved amenable to x-ray crystallography and NMR techniques (12, 13). The structure of a peptide containing a Gly to Ala substitution near the center of the peptide (Pro-Hyp-Gly)₁₀ has been solved by x-ray crystallography (5). This structure shows an overall straight molecule with standard triple helical structures at both ends and a localized conformational deformation at the Ala replacement site. The direct N–H (Gly) … CO (Xaa) hydrogen bond is replaced by a water-mediated hydrogen bond N–H (Ala) … H₂O … CO (Xaa).Here, NMR spectroscopy is used to define the structural and dynamic effect of a Gly to Ser replacement through the application of recently developed NMR methodology on selectively ¹³C/¹⁵N doubly labeled collagen peptides (14). This strategy includes chain assignments, measurement of NOEs, and scalar J-couplings to define the conformation of the peptide. These results combined with NMR hydrogen exchange experiments and temperature-dependent chemical shift data demonstrate the disturbed dynamic features and hydrogen bonding around the Ser substitution site. The NMR data of the Gly to Ser peptide are compared with the NMR and x-ray high resolution structure of the peptide containing a Gly to Ala substitution (5).