Predicting cognitive decline in patients with hypoxaemic COPD

AbstractThe objective was to identify predictors of cognitive decline in patients with hypoxaemic COPD on continuous oxygen therapy.Eighty-four consecutive ambulatory hypoxaemic COPD patients in stable clinical conditions were prospectively studied over the course of 2 yr. Baseline multidimensional assessment included respiratory function tests, blood gas analysis, Mini Mental Status (MMS) test, Geriatric Depression Scale (GDS), Activities of Daily Living (ADLs) and Charlson's index of comorbidity. Reassessments were made 1 yr and 2 yr thereafter. Sequential changes in MMS, GDS and ADLs were assessed by Friedman's ANOVA by rank test.Forty patients completed the study (group A), while 44 died or were lost to follow-up (group B). Group B was characterized by more severe respiratory function impairment and worse performances on ADLs and GDS. In group A, MMS deteriorated from baseline to the 1 yr and 2 yr reassessments (27 ± 2·9 vs. 25·8 ± 4·1 and 25·4 ± 4, P<0·005), whereas GDS and ADLs did not change significantly; the 23 patients experiencing a decline of MMS had baseline lower percentage predicted FVC (52·3 ± 17·1 vs. 66·9 ± 13·4, P<0·03) and FEV1 (27·2 ± 8·6 vs. 44 ± 26·8, P<0·02) values and better affective status (GDS score: 11·9 ± 7·7 vs. 16·5 ± 5·6, P<0·04). Two-year changes in MMS and in GDS scores were inversely correlated (Spearman's ρ = −0·32, P = 0·04).Cognitive decline is faster in the presence of severe bronchial obstruction and parallels the worsening of the affective status in COPD patients on oxygen therapy. The onset of depression rather than baseline depressive symptoms seems to be a risk factor for cognitive decline

Chemical-Scale Studies on the Role of a Conserved Aspartate in Preorganizing the Agonist Binding Site of the Nicotinic Acetylcholine Receptor

The nicotinic acetylcholine receptor and related Cys-loop receptors are ligand-gated ion channels that mediate fast synaptic transmission throughout the central and peripheral nervous system. A highly conserved aspartate residue (D89) that is near the agonist binding site but does not directly contact the ligand plays a critical part in receptor function. Here we probe the role of D89 using unnatural amino acid mutagenesis coupled with electrophysiology. Homology modeling implicates several hydrogen bonds involving D89. We find that no single hydrogen bond is essential to proper receptor function. Apparently, the side chain of D89 establishes a redundant network of hydrogen bonds; these bonds preorganize the agonist binding site by positioning a critical tryptophan residue that directly contacts the ligand. Earlier studies of the D89N mutant led to the proposal that a negative charge at this position is essential for receptor function. However, we find that receptors with neutral side chains at position 89 can function well, if the side chain is less perturbing than the amide of asparagine (nitro or keto groups allow function) or if a compensating backbone mutation is introduced to relieve unfavorable electrostatics

Probing the role of the cation–π interaction in the binding sites of GPCRs using unnatural amino acids

We describe a general application of the nonsense suppression methodology for unnatural amino acid incorporation to probe drug–receptor interactions in functional G protein-coupled receptors (GPCRs), evaluating the binding sites of both the M2 muscarinic acetylcholine receptor and the D2 dopamine receptor. Receptors were expressed in Xenopus oocytes, and activation of a G protein-coupled, inward-rectifying K^+ channel (GIRK) provided, after optimization of conditions, a quantitative readout of receptor function. A number of aromatic amino acids thought to be near the agonist-binding site were evaluated. Incorporation of a series of fluorinated tryptophan derivatives at W6.48 of the D2 receptor establishes a cation–π interaction between the agonist dopamine and W6.48, suggesting a reorientation of W6.48 on agonist binding, consistent with proposed “rotamer switch” models. Interestingly, no comparable cation–π interaction was found at the aligning residue in the M2 receptor

Reduced ratio of protective versus proinflammatory cytokine responses to commensal bacteria in HLA-B27 transgenic rats

Germ-free HLA-B27 transgenic (TG) rats do not develop colitis, but colonization with specific pathogen-free (SPF) bacteria induces colitis accompanied by immune activation. To study host-dependent immune responses to commensal caecal bacteria we investigated cytokine profiles in mesenteric lymph node (MLN) cells from HLA-B27 TG versus nontransgenic (non-TG) littermates after in vitro stimulation with caecal bacterial lysates (CBL). Supernatants from CBL-stimulated unseparated T- or B- cell-depleted MLN cells from HLA-B27 TG and non-TG littermates were analysed for IFN-γ, IL-12, TNF, IL-10 and TGF-β production. Our results show that unfractionated TG MLN cells stimulated with CBL produced more IFN-γ, IL-12 and TNF than did non-TG MLN cells. In contrast, CBL-stimulated non-TG MLN cells produced more IL-10 and TGF-β. T cell depletion abolished IFN-γ and decreased IL-12 production, but did not affect IL-10 and TGF-β production. Conversely, neither IL-10 nor TGF-β was produced in cultures of B cell-depleted MLN. In addition, CD4+ T cells enriched from MLN of HLA-B27 TG but not from non-TG rats produced IFN-γ when cocultured with CBL-pulsed antigen presenting cells from non-TG rats. Interestingly, IL-10 and TGF-β, but not IFN-γ, IL-12 and TNF were produced by MLN cells from germ-free TG rats. These results indicate that the colitis that develops in SPF HLA-B27 TG rats is accompanied by activation of IFN-γ-producing CD4+ T cells that respond to commensal bacteria. However, B cell cytokine production in response to components of commensal intestinal microorganisms occurs in the absence of intestinal inflammation

Chemotherapy and Stem Cell Transplantation Increase p16

AbstractThe expression of markers of cellular senescence increases exponentially in multiple tissues with aging. Age-related physiological changes may contribute to adverse outcomes in cancer survivors. To investigate the impact of high dose chemotherapy and stem cell transplantation on senescence markers in vivo, we collected blood and clinical data from a cohort of 63 patients undergoing hematopoietic cell transplantation. The expression of p16INK4a, a well-established senescence marker, was determined in T-cells before and 6months after transplant. RNA sequencing was performed on paired samples from 8 patients pre- and post-cancer therapy. In patients undergoing allogeneic transplant, higher pre-transplant p16INK4a expression was associated with a greater number of prior cycles of chemotherapy received (p=0.003), prior autologous transplantation (p=0.01) and prior exposure to alkylating agents (p=0.01). Transplantation was associated with a marked increase in p16INK4a expression 6months following transplantation. Patients receiving autologous transplant experienced a larger increase in p16INK4a expression (3.1-fold increase, p=0.002) than allogeneic transplant recipients (1.9-fold increase, p=0.0004). RNA sequencing of T-cells pre- and post- autologous transplant or cytotoxic chemotherapy demonstrated increased expression of transcripts associated with cellular senescence and physiological aging. Cytotoxic chemotherapy, especially alkylating agents, and stem cell transplantation strongly accelerate expression of a biomarker of molecular aging in T-cells

Lifestyle impacts on the aging-associated expression of biomarkers of DNA damage and telomere dysfunction in human blood: Measuring the influence of lifestyle on aging

Cellular aging is characterised by telomere shortening, which can lead to uncapping of chromosome ends (telomere dysfunction) and that activation of DNA damage responses. There is some evidence the DNA damage accumulates during human aging and that lifestyle factors contribute to the accumulation of DNA damage. Recent studies have identified a set of serum markers that are induced by telomere dysfunction and DNA damage and these markers showed an increased expression in blood during human aging. Here, we investigated the influence of lifestyle factors (such as exercise, smoking, body mass) on the aging associated expression of serum markers of DNA damage (CRAMP, EF-1α, Stathmin, n-acetyl-glucosaminidase, and chitinase) in comparison to other described markers of cellular aging (p16INK4a upregulation and telomere shortening) in human peripheral blood. The study shows that lifestyle factors have an age-independent impact on the expression level of biomarkers of DNA damage. Smoking and increased body mass indices were associated with elevated levels of biomarkers of DNA damage independent of the age of the individuals. In contrast, exercise was associated with an age-independent reduction in the expression of biomarkers of DNA damage in human blood. The expression of biomarkers of DNA damage correlated positively with p16INK4a expression and negatively with telomere length in peripheral blood T-lymphocytes. Together, these data provide experimental evidence that both aging and lifestyle impact on the accumulation of DNA damage during human aging

Two Neuronal Nicotinic Acetylcholine Receptors, α4β4 and α7, Show Differential Agonist Binding Modes

Nicotinic acetylcholine receptors (nAChRs) are pentameric, neurotransmitter-gated ion channels responsible for rapid excitatory neurotransmission in the central and peripheral nervous systems, resulting in skeletal muscle tone and various cognitive effects in the brain. These complex proteins are activated by the endogenous neurotransmitter ACh as well as by nicotine and structurally related agonists. Activation and modulation of nAChRs has been implicated in the pathology of multiple neurological disorders, and as such, these proteins are established therapeutic targets. Here we use unnatural amino acid mutagenesis to examine the ligand binding mechanisms of two homologous neuronal nAChRs: the α4β4 and α7 receptors. Despite sequence identity among the residues that form the core of the agonist-binding site, we find that the α4β4 and α7 nAChRs employ different agonist-receptor binding interactions in this region. The α4β4 receptor utilizes a strong cation-π interaction to a conserved tryptophan (TrpB) of the receptor for both ACh and nicotine, and nicotine participates in a strong hydrogen bond with a backbone carbonyl contributed by TrpB. Interestingly, we find that the α7 receptor also employs a cation-π interaction for ligand recognition, but the site has moved to a different aromatic amino acid of the agonist-binding site depending on the agonist. ACh participates in a cation-π interaction with TyrA, whereas epibatidine participates in a cation-π interaction with TyrC2

Effect of Cytotoxic Chemotherapy on Markers of Molecular Age in Patients With Breast Cancer

Senescent cells, which express p16 INK4a, accumulate with aging and contribute to age-related pathology. To understand whether cytotoxic agents promote molecular aging, we measured expression of p16 INK4a and other senescence markers in breast cancer patients treated with adjuvant chemotherapy

Effect of metal Ions (Ni2+, Cu2+ and Zn2+) and water coordination on the structure of L-phenylalanine, L-tyrosine, L-tryptophan and their zwitterionic forms

Methods of quantum chemistry have been applied to double-charged complexes involving the transition metals Ni2+, Cu2+ and Zn2+ with the aromatic amino acids (AAA) phenylalanine, tyrosine and tryptophan. The effect of hydration on the relative stability and geometry of the individual species studied has been evaluated within the supermolecule approach. The interaction enthalpies, entropies and Gibbs energies of nine complexes Phe•M, Tyr•M, Trp•M, (M = Ni2+, Cu2+ and Zn2+) were determined at the Becke3LYP density functional level of theory. Of the transition metals studied the bivalent copper cation forms the strongest complexes with AAAs. For Ni2+and Cu2+ the most stable species are the NO coordinated cations in the AAA metal complexes, Zn2+cation prefers a binding to the aromatic part of the AAA (complex II). Some complexes energetically unfavored in the gas-phase are stabilized upon microsolvation