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Biomedical subjects

C H Li

Publications and source records attributed to C H Li.

At least 19 recordsLinked to original sources
PubMed1992

Purification and functional reconstitution of the cystic fibrosis transmembrane conductance regulator (CFTR).

Circumstantial evidence has accumulated suggesting that CFTR is a regulated low-conductance Cl- channel. To test this postulate directly, we have purified to homogeneity a recombinant CFTR protein from a high-level baculovirus-infected insect cell line. Evidence of purity included one- and two-dimensional gel electrophoresis, N-terminal peptide sequence, and quantitative amino acid analysis. Reconstitution into proteoliposomes at less than one molecule per vesicle was accomplished by established procedures. Nystatin and ergosterol were included in these vesicles, so that nystatin conductance could serve as a quantitative marker of vesicle fusion with a planar lipid bilayer. Upon incorporation, purified CFTR exhibited regulated chloride channel activity, providing evidence that the protein itself is the channel. This activity exhibited the basic biophysical and regulatory properties of the type of Cl- channel found exclusively in CFTR-expressing cell types and believed to underlie cAMP-evoked secretion in epithelial cells.

Animals↗
PubMed1991

Insulin receptors prepared with iodoacetamide show enhanced autophosphorylation and receptor kinase activity.

In this study, we found that adding iodoacetamide to the homogenization buffer used in the preparation of mouse or rat liver plasma membranes resulted in an increase of insulin receptor autophosphorylation by 4-5-fold and receptor kinase activity by about 2-fold. Similar effects were obtained with iodoacetate and p-chloromercuriphenyl sulfonate. The effect of iodoacetamide was minimal when it was added to membranes prepared without the thiol reagent. The enhancing effect of iodoacetamide on insulin receptor autophosphorylation was the result of a more than 2-fold decrease in the Km and a more than 3-fold increase in Vmax for ATP. The presence of iodoacetamide in the preparation of plasma membranes also greatly increased the solubilization of the insulin receptor from the plasma membrane by Triton X-100. We propose that iodoacetamide acts to alkylate some unknown thiols released during tissue homogenization and that in its absence these thiols formed mixed disulfides with the insulin receptor, thus adversely affecting the process of receptor activation by insulin.

Animals↗
PubMed1991

Pharmacological characterization of the inhibitory activity of beta h-endorphin (beta h-EP), [Arg9,19,24,28,29]-beta h-EP, [Gln8,Gly31]-beta h-EP-Gly-Gly-NH2, in the neuroeffector junction of the mouse vas deferens.

The inhibitory opioid activities of beta h-endorphin (beta h-EP), its structurally related peptide analogues [Gln8,Gly31]-beta h-EP-Gly-Gly-NH2 (Gly-Gly-beta h-EP), [Arg9,19,24,28,29]-beta h-EP (Arg-beta h-EP) and methionine enkephalin have been examined in the electrically stimulated mouse vas deferens bioassay. All four peptides behaved as full agonists; methionine enkephalin was the most potent followed by Arg-beta h-EP, beta h-EP and Gly-Gly-beta h-EP. Neither Gly-Gly-beta h-EP nor Arg-beta h-EP antagonized the inhibitory action of beta h-EP or methionine enkephalin. An hour of tissue exposure to 30 nM beta-funaltrexamine followed by thorough washing, displaced to the right, in a parallel fashion, the concentration-response curves of beta h-EP and analogues. Whereas the displacement of the concentration response curves was 8 to 10-fold for beta h-EP and Arg-beta h-EP, it was only about 3-fold for Gly-Gly-beta h-EP and methionine enkephalin. Naltrindole was the most potent antagonist of methionine enkephalin with an apparent pA2 of 9.4; its potency as an antagonist of beta h-EP and related analogues was approximately one-tenth of this with pA2 values approximately 8.5. Norbinaltorphimine also antagonized the action of the opioid peptides with pA2 values close to 7.8.

Animals↗
PubMed1991

Calcium-permeable channels in rat hepatoma cells are activated by extracellular nucleotides.

Extracellular ATP is known to cause uptake of Ca2+ by rat liver cells. The specific pathway permitting influx of Ca2+ has not yet been identified. In the present investigations, we studied the properties of ATP-evoked 45Ca2+ uptake in rat hepatoma cell monolayers and then used patch-clamp electrophysiology to identify the channel that may account for this uptake. The results suggest that ATP-stimulated 45Ca2+ uptake occurs as a result of P2-purinergic receptor interaction because uptake was inhibited by Reactive Blue (100 microM), a blocker of this type of receptor. Furthermore, the ability of other adenine nucleotides to stimulate 45Ca2+ uptake was related to the selectivity sequence for binding to the P2-purinergic receptor. ATP-stimulated 45Ca2+ uptake occurs primarily through a conductance pore since it was inhibited by 70% upon dissipation of the membrane potential using the K+ ionophore valinomycin. The calcium channel blockers nifedipine and verapamil failed to inhibit 45Ca2+ uptake, but gadolinium (GdCl3) was an effective blocker. In cell-attached patch-clamp experiments, a single type of channel was activated with ATP (100 microM) addition to the bath in 18 of 32 trials. The current-voltage relationship of the ATP-activated channel is identical to that of the stretch-activated channel previously characterized in this laboratory as a calcium-permeable cation-nonselective channel [Am. J. Physiol. 258 (Cell Physiol. 27): C421-C428, 1990]. There are several lines of evidence which suggest that this cation-nonselective channel may account for ATP-stimulated 45Ca2+ uptake.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1979

Studies on prolactin. Selective reduction of the disulfide bonds of the ovine hormone.

Methods for selective reduction of the disulfide bonds in ovine prolactin are reported. Cleavage of all three disulfide bonds abolishes biological activity and denatures the hormone. Reduction-carbamidomethylation of one or two of the disulfide bridges does not diminish the biological activities in the pigeon crop-sac and mouse mammary gland bioassays. When compared to the native hormone, monomers of these two partially reduced-carbamidomethylated derivatives also show only modest changes in properties measured by exclusion chromatography, circular dichroism, and immunological cross-reactivities. However, cleavage of cystine-4--11 and cystine-191--199, followed by carbamidomethylation, destroys the biological activity of this derivative in a teleost fish bioassay (Gillichthys urinary bladder). In contrast, reduction of cystine-4--11 actually increased the teleost potency of this derivative compared to the intact hormone. Since teleost prolactin appears to lack a homologue to the cystine-4--11 disulfide bond in the amino-terminal loop of the ovine hormone, selective reduction of this bond in ovine prolactin may produce a derivative whose properties more closely resemble the fish hormone.

Amino Acid Sequence↗
PubMed1979

Effect of beta-endorphin on calcium uptake in the brain.

The uptake of 45Ca2+ by nerve-ending fractions from brains of mice was inhibited in vitro by 10(-9)M concentrations of beta-endorphin and in mice injected intraventricularly with 7 picomoles of beta-endorphin. That the effect was a specific opiate agonist response of beta-endorphin was demonstrated by use of the opiate antagonist, naloxone, which reversed the action. A role for beta-endorphin in the regulation of calcium flux and neurotransmitter release should be considered.

Animals↗
PubMed1979

Behavioral activities of opioid peptides and morphine sulfate in golden hamsters and rats.

The behavioral effects of beta-endorphin, [D-Ala2, D-Leu5]-enkephalin and morphine were investigated in golden hamsters and in rats. In golden hamsters, beta-endorphin and [D-Ala2, D-Leu5]-enkephalin induced loss of righting reflex, whereas morphine caused no such effect. Both opiate peptides and morphine caused the inhibition of tail-flick response and catalepsy in rats. beta-Endorphin was the most potent, followed by [D-Ala2, D-Leu5]-enkephalin and then by morphine. The catalepsy induced in rats by [D-Ala2, D-Leu5]-enkephalin was different from that of beta-endorphin and morphine in that it produced catalepsy without muscular rigidity. beta-Endorphin and [D-Ala2, D-Leu5]-enkephalin caused hypothermia in golden hamsters; morphine was less active in altering the body temperature. beta-Endorphin caused hypothermia at high doses and hyperthermia at low doses in rats. These heterogenous behavioral responses indicate that multiple types of receptors mediate the effects of opiates in the central nervous system.

Animals↗
PubMed1979

Human EEG response to beta-endorphin.

Beta-endorphin, morphine, and saline were given intravenously to a single schizophrenic subject on separate occasions in a double-blind design. EEG spectral analyses performed on data collected before and after drug injection demonstrated that beta-endorphin and morphine produced similar increases in alpha power within 5 to 15 minutes after injection. This effect could be distinguished from two placebo (saline) injections. These data suggest that intravenous beta-endorphin can produce changes in the central nervous system in humans.

Adult↗
PubMed1979

beta-Endorphin: analgesic and hormonal effects in humans.

The pharmacokinetics and the hormonal, analgesic, and behavioral effects of several doses of human beta-endorphin were evaluated after intravenous administration to three patients and intracerebroventricular administration to one patient with pain caused by cancer. These effects were compared to the hormonal effects of intravenously administered morphine sulfate in two patients and an enkephalin analog in two baboons. The mean terminal half-life after intravenous administration of 5 or 10 mg of human beta-endorphin to three patients was 37 min; the mean volume of distribution was 178 ml/kg, and the metabolic clearance rate was 3.2 (ml/min)/kg. The half-life of beta-endorphin in cerebrospinal fluid after intracerebroventricular administration was 93 min, and the volume of distribution was 0.74 ml/kg. A rapid rise in plasma prolactin followed both intravenous and intracerebroventricular beta-endorphin. Intravenous administration did not affect plasma growth hormone, but intracerebroventricular administration suppressed plasma growth hormone. No significant change in plasma growth hormone was noted after intravenous administration of morphine to humans, but plasma growth hormone decreased in one baboon after administration of the enkephalin analog. beta-Endorphin-stimulated release of prolactin occurred at doses lower than those required to produce analgesic and other behavioral effects. When both hormonal and analgesic effects were observed (after 7.5 mg were given intracerebroventricularly), the onset of the hormonal response slightly preceded the analgesic and behavioral responses. These studies suggest that the hormonal effects of beta-endorphin are species dependent and are similar to those of morphine. Hormonal and analgesic effects of beta-endorphin appear to result from the activation of opiate receptors that differ in their locations and characteristics.

Adult↗
PubMed1979

Properties and localization of beta-endorphin receptor in rat brain.

Stereospecific high-affinity binding sites for beta h-[3H]endorphin could be demonstrated in the P2 pellet of rat brain homogenate. Scatchard analysis of the binding data revealed binding sites with Kd values of 0.81 and 6.8 nM and density of 120 and 240 fmol/mg of protein. Distribution of beta h-[3H]endorphin binding in various brain regions parallels that of opiate receptor:striatum greater than thalamus greater than amygdala greater than hypothalamus, septum greater than cortex greater than midbrain, brainstem. Similar to their effect on 3H-labeled agonist binding, Na+ and other monovalent cations, GTP, trypsin, chymotrypsin, phospholipase A2, and N-ethylmaleimide all inhibited the specific binding of beta h-[3H]endorphin. In contrast to their action on alkaloid and enkephalin binding, Ca2+, Mg2+, and Mn2+ also inhibited beta h-[3H]endorphin binding. These data suggest a difference between beta h-endorphin and alkaloid/enkephalin binding sites.

Animals↗
PubMed1979

Semisynthesis of human somatotropin analogs.

Complementation of the natural NH2-terminal 134-amino acid fragment of the reduced, carbamoylmethylated human somatotropin with synthetic analogs of COOH-terminal fragments of 57, 42, or 38 amino acids of reduced, carbamoyl-methylated human somatotropin has been investigated. It was found that synthetic fragments of 57 and 42 amino acids gave recombinants with full growth-promoting activity, whereas attempts to obtain a recombinant with the synthetic 38-amino acid fragment were unsuccessful. The synthesis of two analogs of the COOH-terminal fragment of human somatotropin, human [N]e170, Ala165,182,189] somatotropin-(150-191) and human [Nle170, Ala165,182,189] somatotropin-(154-191), is herein described.

Amino Acid Sequence↗
PubMed1979

beta-Endorphin: synthesis of analogs modified at the carboxyl terminus with increased activites.

Three analogs of human beta-endorphin (beta h-EP) have been synthesized: [Gly31]beta h-EP, [Gly31]beta h-endorphinamide, and [Gly31]beta h-endorphinylglycine. All are more active than beta h-EP in both the guinea pig ileum bioassay and the opiate receptor binding assay. The last two analogs are about twice as active as beta h-EP in an assay for analgesia. Modification at position 31 and extension at the COOH terminus may afford a route toward analogs with even greater biological activity.

Analgesics↗
PubMed1979

beta-Endorphin: formation of alpha-helix in lipid solutions.

Human beta-endorphin adopts a partial helical conformation in aqueous solutions of cerebroside sulfate, ganglioside GM1, phosphatidylserine, and phosphatidic acid, but not of cerebroside and phosphatidylcholine, as evidenced by circular dichroic spectra. Addition of Ca2+ to the peptide in cerebroside sulfate solution can break up the helix; at 10 mM Ca2+ the peptide (12 microM) essentially exists in an unordered form. For comparison, sheep beta-lipotropin in acidic cerebroside sulfate solution (pH less than 4) also has a partial helical conformation of the complex between human beta-endorphin and lipids may be related to the opiatelike function of this peptide hormone.

Adrenocorticotropic Hormone↗