Fructus Amomi extract attenuates nasal inflammation by restoring Th1/Th2 balance and down-regulation of NF-κB phosphorylation in OVA-induced allergic rhinitis

Fructus Amomi Cardamomi (FA) is the mature fruit of Amomum villosum Lour (family Zingiberaceae) and is commonly used in Chinese traditional medicine to treat various gastrointestinal disorders. FA’s possible benefits as an allergic rhinitis (AR) treatment, however, have not been examined. We used an ovalbumin (OVA)-induced AR mouse model to identify any anti-allergic effects associated with the administration of 200 mg/kg FA or dexamethasone (Dex) 2.5 mg/kg by oral administration. The results of our testing confirm that FA ameliorated nasal symptoms and alleviated nasal epithelium swelling, reduced the goblet cell hyperplasia and eosinophil cell infiltration in the nasal epithelium, and inhibited lung tissue inflammation and Dex as well. Significantly decreased Th2 cytokine (interleukin (IL)-1β, IL-4, and IL-5) expression, and a correspondingly significant increase in Th1 cytokine (IL-12, interferon (IFN)-γ) production, was observed in nasal lavage fluid (NALF) taken from mice that received FA or Dex treatment. FA also reduced the presence of OVA-specific immunoglobulin (Ig) E, OVA-specific IgG₁, and histamine levels in serum, and inhibited mast cell degranulation in vitro. In addition, these effects were involved with the reduction in NF-κB phosphorylation. These results suggest that FA restores Th1/Th2 balance and inhibits NF-κB phosphorylation and mast cell degranulation, thereby achieving a notable anti-inflammatory effect. Accordingly, it has the potential to be used as an efficacious therapeutic treatment for AR.     

The MukB-topoisomerase IV interaction mutually suppresses their catalytic activities

The bacterial condensin MukB and the cellular chromosomal decatenase, topoisomerase IV interact and this interaction is required for proper condensation and topological ordering of the chromosome. Here, we show that Topo IV stimulates MukB DNA condensation by stabilizing loops in DNA: MukB alone can condense nicked plasmid DNA into a protein–DNA complex that has greater electrophoretic mobility than that of the DNA alone, but both MukB and Topo IV are required for a similar condensation of a linear DNA representing long stretches of the chromosome. Remarkably, we show that rather than MukB stimulating the decatenase activity of Topo IV, as has been argued previously, in stoichiometric complexes of the two enzymes each inhibits the activity of the other: the ParC subunit of Topo IV inhibits the MukF-stimulated ATPase activity of MukB and MukB inhibits both DNA crossover trapping and DNA cleavage by Topo IV. These observations suggest that when in complex on the DNA, Topo IV inhibits the motor function of MukB and the two proteins provide a stable scaffold for chromosomal DNA condensation.     

Granal stacking of thylakoid membranes in higher plant chloroplasts: the physicochemical forces at work and the functional consequences that ensue.

The formation of grana in chloroplasts of higher plants is examined in terms of the subtle interplay of physicochemical forces of attraction and repulsion. The attractive forces between two adjacent membranes comprise (1) van der Waals attraction that depends on the abundance and type of atoms in each membrane, on the distance between the membranes and on the dielectric constant, (2) depletion attraction that generates local order by granal stacking at the expense of greater disorder (i.e. entropy) in the stroma, and (3) an electrostatic attraction of opposite charges located on adjacent membranes. The repulsive forces comprise (1) electrostatic repulsion due to the net negative charge on the outer surface of thylakoid membranes, (2) hydration repulsion that operates at small separations between thylakoid membranes due to layers of bound water molecules, and (3) steric hindrance due to bulky protrusions of Photosystem I (PSI) and ATP synthase into the stroma. In addition, specific interactions may occur, but they await experimental demonstration. Although grana are not essential for photosynthesis, they are ubiquitous in higher plants. Grana may have been selected during evolution for the functional advantages that they confer on higher plants. The functional consequences of grana stacking include (1) enhancement of light capture through a vastly increased area-to-volume ratio and connectivity of several PSIIs with large functional antenna size, (2) the ability to control the lateral separation of PSI from PSII and, therefore, the balanced distribution of excitation energy between two photosystems working in series, (3) the reversible fine-tuning of energy distribution between the photosystems by State 1-State 2 transitions, (4) the ability to regulate light-harvesting via controlled thermal dissipation of excess excitation energy, detected as non-photochemical quenching, (5) dynamic flexibility in the light reactions mediated by a granal structure in response to regulation by a trans-thylakoid pH gradient, (6) delaying the premature degradation of D1 and D2 reaction-centre protein(s) in PSII by harbouring photoinactived PSIIs in appressed granal domains, (7) enhancement of the rate of non-cyclic synthesis of adenosine triphosphate (ATP) as well as the regulation of non-cyclic vs. cyclic ATP synthesis, and (8) the potential increase of photosynthetic capacity for a given composition of chloroplast constituents in full sunlight, concomitantly with enhancement of photochemical efficiency in canopy shade. Hence chloroplast ultrastructure and function are intimately intertwined.     

Regulation of fragile X mental retardation 1 protein by C-terminus of Hsc70-interacting protein depends on its phosphorylation status

The fragile X mental retardation 1 (FMR1) protein binds mRNA and acts as a negative regulator of translation. Lack of FMR1 causes the most common neurological disorder, fragile X syndrome, while its overexpression is associated with metastasis of breast cancer. Its activity has been well-studied in nervous tissue, but recent evidence as well as its role in cancer indicates that it also acts in other tissues. We have investigated the expression of FMR1 in brain and other tissues of mouse and examined its regulation. We detected expression of FMR1 in liver and heart tissues of mice as well as in brain tissue, supporting other contentions that it acts in non-nervous tissue. Expression of FMR1 inversely correlated with expression of the C-terminus of Hsc70-interacting protein (CHIP) and, based on the known activity of CHIP in protein homeostasis, we suggest that CHIP regulates expression of FMR1. CHIP ubiquitinated FMR1 for proteasomal degradation in a molecular chaperone-independent manner. FMR1 expression was reduced following treatment with okadaic acid, a phosphatase inhibitor, but not in CHIP-depleted cells. Also, a non-phospho FMR1 mutant was much less efficiently ubiquitinated by CHIP and had a longer half-life compared to either wild-type FMR or a phospho-mimic mutant. Taken together, our results demonstrate that CHIP regulates the levels of FMR1 as an E3 ubiquitin ligase in phosphorylation-dependent manner, suggesting that CHIP regulates FMR1-mediated translational repression by regulating the levels of FMR1.     

The phenotype of Arabidopsis ovule mutants mimics the morphology of primitive seed plants

In seed plants, the ovule is the female reproductive structure, which surrounds and nourishes the gametophyte and embryo. This investigation describes the PRETTY FEW SEEDS2 (PFS2) locus, which regulates ovule patterning. The pfs2 mutant exhibited developmental defects in the maternal integuments and gametophyte. This mutation was inherited as a maternal trait, indicating that gametophyte defects resulted from ovule patterning aberrations. Specifically, the boundary between the chalaza and the nucellus, two regions of the ovule primordia, shifted towards the distal end of pfs2 ovule primordia. Results indicated that the PFS2 locus could: (i) be involved in the development of either the nucellus or the chalaza; or (ii) establish a boundary between these two regions. Examination of genetic interactions of the pfs2 mutation with other well-characterized ovule loci indicates that this locus affects integument morphogenesis. Interestingly, the pfs2 inner no outer and pfs2 strubbelig double mutants had inner integuments that appeared similar to their ancestral precursor. The fossil record indicates that the inner integument evolved by fusion of sterilized sporangia or branches around a central megasporangium. The question of whether the structures observed in these double mutants are homologous or merely analogous to the ancestral precursors of the inner integument is discussed.     

CD99 costimulation up-regulates T cell receptor-mediated activation of JNK and AP-1

Although CD28 is the principal T cell costimulatory molecule for the T cell receptor, a number of other cell surface proteins have costimulatory functions and perform specific roles in different contexts. Here we analyzed the mechanism of CD99 costimulation of the T cell receptor. Cooperation of CD99 engagement with suboptimal TCR/CD3 signals resulted in greatly enhanced CD4+ T cell proliferation. CD99 costimulation also led to elevated expression of CD25 and GM1 on the CD4+ T cell surface within 3 days. In Jurkat TAg cells, CD99 costimulation led to increased apoptosis compared to stimulation with CD3 or CD99 alone. CD99 costimulation also augmented activation of MAP kinases, especially of JNK, and increased AP-1 activation was also observed using a luciferase reporter assay. These results show that CD99 has a costimulatory function for T cells and acts by a mechanism distinct from CD28.     

Improved embryo development with decreased apoptosis in blastomeres after the treatment of cloned bovine embryos with beta-mercaptoethanol and hemoglobin

In preliminary experiments, the treatment of donor somatic cells with beta-mercaptoethanol (ME) or hemoglobin (Hb) improved in vitro-development of bovine cloned embryos. This study was subsequently evaluated whether the exposure to Hb and/or ME during in vitro-maturation or embryo culture could further promote the development of embryos cloned with ME-treated donor cells. A prospective, randomized study was conducted and, embryo development, cell number, and apoptosis in blastocysts were monitored. A significant (P < 0.05) effect was found after the combined treatment of cloned embryos with Hb (1 microg/ml) and ME (10 microM); the development of morulae (53 vs. 35%) was greatly improved, which resulted in enhanced blastocyst formation (38%). However, cell number and apoptosis in blastocysts were predominantly affected by ME rather than Hb; a significant increase in total cell number of blastomeres (142-154 vs. 123 cells/embryo), inner cell mass (ICM) (39-41 vs. 27), and trophectoderm (TE) (103-114 vs. 98), and the ratio of ICM to TE cell number (0.26-0.27 vs. 0.22) was found. Also, the apoptosis index indicating the ratio of apoptotic cell to normal blastomere number was greatly reduced after ME treatments (0.85 vs. 0.056-0.069). When embryos cloned with ME-treated cells were cultured in Hb + ME-containing medium, any of the treatments to recipient oocytes before enucleation did not further promote the development. In conclusion, combined treatment of cloned embryos with Hb + ME not only improved in vitro-development but also decreased blastomere apoptosis. The use of ME-treated donor cells and the culture of cloned embryos in Hb + ME-containing medium yielded the optimal results for promoting the production of blastocysts with improved quality.     

Polyglutamine expansion in ataxin-3 does not affect protein stability: implications for misfolding and disease

Polyglutamine proteins that cause neurodegenerative disease are known to form proteinaceous aggregates, such as nuclear inclusions, in the neurons of affected patients. Although polyglutamine proteins have been shown to form fibrillar aggregates in a variety of contexts, the mechanisms underlying the aberrant conformational changes and aggregation are still not well understood. In this study, we have investigated the hypothesis that polyglutamine expansion in the protein ataxin-3 destabilizes the native protein, leading to the accumulation of a partially unfolded, aggregation-prone intermediate. To examine the relationship between polyglutamine length and native state stability, we produced and analyzed three ataxin-3 variants containing 15, 28, and 50 residues in their respective glutamine tracts. At pH 7.4 and 37 degrees C, Atax3(Q50), which lies within the pathological range, formed fibrils significantly faster than the other proteins. Somewhat surprisingly, we observed no difference in the acid-induced equilibrium and kinetic un/folding transitions of all three proteins, which indicates that the stability of the native conformation was not affected by polyglutamine tract extension. This has led us to reconsider the mechanisms and factors involved in ataxin-3 misfolding, and we have developed a new model for the aggregation process in which the pathways of un/folding and misfolding are distinct and separate. Furthermore, given that native state stability is unaffected by polyglutamine length, we consider the possible role and influence of other factors in the fibrillization of ataxin-3.     

Kinetic analysis and modelling of enzymatic (R)-phenylacetylcarbinol batch biotransformation process

Initial rate and biotransformation studies were applied to refine and validate a mathematical model for enzymatic (R)-phenylacetylcarbinol (PAC) production from pyruvate and benzaldehyde using Candida utilis pyruvate decarboxylase (PDC). The rate of PAC formation was directly proportional to the enzyme activity level up to 5.0 U ml-1 carboligase. Michaelis-Menten kinetics were determined for the effect of pyruvate concentration on the reaction rate. The effect of benzaldehyde followed the sigmoidal shape of the Monod-Wyman-Changeux (MWC) model. The biotransformation model, which also included a term for PDC inactivation by benzaldehyde, was used to determine the overall rate constants for the formation of PAC, acetaldehyde, and acetoin. These values were determined from data for three batch biotransformations performed over a range of initial concentrations (viz. 50-150 mM benzaldehyde, 60-180 mM pyruvate, 1.1-3.4 U ml-1 enzyme activity). The finalized model was then used to predict a batch biotransformation profile at 120/100 mM initial pyruvate/benzaldehyde (initial enzyme activity 3.0 U ml-1). The simulated kinetics gave acceptable fitting (R2 = 0.9963) to the time courses of these latter experimental data for substrates pyruvate and benzaldehyde, product PAC, by-products acetaldehyde and acetoin, as well as enzyme activity level.