24 research outputs found
Distribution of common peptide YY-neuropeptide Y receptor along rat intestinal villus-crypt axis
Formation of dense erythrocyte in SAD mice exposed to chronic hypoxia: evaluation of different therapeutic regimens and additive benfit of comnbining oral clotrimazole and magnesium therapies.
Functional VIP receptors in the human mucus-secreting colonic epithelial cell line CL.16E
Combination therapy of erythropoietin, hydroxyurea, and clotrimazole in a beta thalassemic mouse: a model for human therapy
beta thalassemia (beta thal) in DBA/2J mice is a consequence of the spontaneous and complete deletion of the beta major globin gene. Homozygous beta thal mice have clinical and biological features similar to those observed in human beta thal intermedia. Erythrocytes in human beta thal are characterized by a relative cell dehydration and reduced K+ content. The role of this erythrocyte dehydration in the reduced erythrocyte survival, which typifies the disease, has not previously been evaluated. We examined for 1 month the effects on the anemia and the erythrocyte characteristics of beta thal mice of daily treatment with either clotrimazole (CLT), an inhibitor of red blood cell (RBC) dehydration via the Gardos channel, or human recombinant erythropoietin (r-HuEPO), or hydroxyurea (HU). The use of either r-HuEPO or HU induced a significant increase in hemoglobin (Hb), hematocrit (Hct), erythrocyte K+ and a decrease in percent reticulocytes, suggesting improved erythrocyte survival. CLT alone decreased only mean corpuscular hemoglobin concentration (MCHC) and cell density and increased cell K+. Thus, though the Gardos channel plays a major role in cell dehydration of murine beta thal erythrocyte survival. Combination therapy with r-HuEPO plus HU produced no incremental benefit beyond those of single drug therapy. However, addition of CLT to r-HuEPO, to HU, or to combined r-HuEPO plus HU led to statistically significant increase in Hb, Hct, and erythrocyte K+ compared with any of the regimens without CLT. These results suggest that CLT not only inhibits erythrocyte dehydration, but also potentiates the erythropoietic and cellular survival responses to r-HuEPO and HU
The Human Vpac(1) Receptor - Three-Dimensional Model And Mutagenesis Of The N-Terminal Domain
peer reviewedThe human VPAC(1) receptor for vasoactive intestinal peptide (VIP) and pituitary
adenylate cyclase activating peptide belongs to the class II family of
G-protein-coupled receptors with seven transmembrane segments. Like for all class
II receptors, the extracellular N-terminal domain of the human VPAC(1) receptor
plays a predominant role in peptide ligand recognition. To determine the
three-dimensional structure of this N-terminal domain (residues 1-144), the
Protein Data Bank (PDB) was screened for a homologous protein. A subdomain of
yeast lipase B was found to have 27% sequence identity and 50% sequence homology
with the N-terminal domain (8) of the VPAC(1) receptor together with a good
alignment of the hydrophobic clusters. A model of the N-terminal domain of
VPAC(1) receptor was thus constructed by homology. It indicated the presence of a
putative signal sequence in the N-terminal extremity. Moreover, residues
(Glu(36), Trp(67), Asp(68), Trp(73), and Gly(109)) which were shown to be crucial
for VIP binding are gathered around a groove that is essentially negatively
charged. New putatively important residues for VIP binding were suggested from
the model analysis. Site-directed mutagenesis and stable transfection of mutants
in CHO cells indicated that Pro(74), Pro(87), Phe(90), and Trp(110) are indeed
important for VIP binding and activation of adenylyl cyclase activation.
Combination of molecular modeling and directed mutagenesis provided the first
partial three-dimensional structure of a VIP-binding domain, constituted of an
electronegative groove with an outspanning tryptophan shell at one end, in the
N-terminal extracellular region of the human VPAC(1) receptor
Identification Of Key Residues For Interaction Of Vasoactive Intestinal Peptide With Human Vpac(1) And Vpac(2) Receptors And Development Of A Highly Selective Vpac(1) Receptor Agonist - Alanine Scanning And Molecular Modeling Of The Peptide
The widespread neuropeptide vasoactive intestinal peptide (VIP) has two receptors
VPAC(1) and VPAC(2). Solid-phase syntheses of VIP analogs in which each amino
acid has been changed to alanine (Ala scan) or glycine was achieved and each
analog was tested for: (i) three-dimensional structure by ab initio molecular
modeling; (ii) ability to inhibit (125)I-VIP binding (K(i)) and to stimulate
adenylyl cyclase activity (EC(50)) in membranes from cell clones stably
expressing human recombinant VPAC(1) or VPAC(2) receptor. The data show that
substituting residues at 14 positions out of 28 in VIP resulted in a >10-fold
increase of K(i) or EC(50) at the VPAC(1) receptor. Modeling of the
three-dimensional structure of native VIP (central alpha-helice from Val(5) to
Asn(24) with random coiled N and C terminus) and analogs shows that substitutions
of His(1), Val(5), Arg(14), Lys(15), Lys(21), Leu(23), and Ile(26) decreased
biological activity without altering the predicted structure, supporting that
those residues directly interact with VPAC(1) receptor. The interaction of the
analogs with human VPAC(2) receptor is similar to that observed with VPAC(1)
receptor, with three remarkable exceptions: substitution of Thr(11) and Asn(28)
by alanine increased K(i) for binding to VPAC(2) receptor; substitution of
Tyr(22) by alanine increased EC(50) for stimulating adenylyl cyclase activity
through interaction with the VPAC(2) receptor. By combining 3 mutations at
positions 11, 22, and 28, we developed the [Ala(11,22,28)]VIP analog which
constitutes the first highly selective (>1,000-fold) human VPAC(1) receptor
agonist derived from VIP ever described