Comparison of three corneal trephines for use in therapeutic penetrating keratoplasties for large corneal perforations.

Corneas with large perforations complicate penetrating keratoplasty due to the increased risk of anterior chamber collapse they pose. We hypothesize that suction trephines should produce more uniform corneal openings than non-suction trephines. Penetrating keratoplasties using Franceschetti-type freeblades, and Hanna and Hessberg-Barron suction trephines were performed on human eye bank eyes with large corneal perforations.

Accumulation of ornithine by chick retinal pigment epithelium cells in culture.

This study was designed to measure the accumulation of ornithine in retinal pigment epithelium cells grown in culture. Ornithine accumulated in retinal pigment epithelium cells in which the ornithine aminotransferase activity was inhibited with L-canaline. The effect of L-canaline was eliminated by the concomitant presence of methionine.

Structure-activity studies of antagonists of luteinizing hormone-releasing hormone with emphasis on the amino-terminal region.

The structure-activity relationship of the hydrophobic amino terminal region of the antagonist [N-Ac-D-Nal1,D-pClPhe2,D-Trp3,D-Arg6,Phe7,D-Ala10]-LH- RH has been investigated by the incorporation of a variety of amino acids with emphasis on positions 1, 2, and 3. The analogues were prepared by routine solid-phase peptide synthesis. All purifications were performed in two stages: gel permeation chromatography followed by preparative, reversed-phase, high-performance chromatography.

Effect of reductive alkylation of D-lysine in position 6 on the histamine-releasing activity of luteinizing hormone-releasing hormone antagonists.

The reductive alkylation of the D-Lys side chain in position 6 of the LH-RH antagonist [N-Ac-D-Nal1,D-Phe2,3,D-Arg6,Phe7,D-Ala10]LH-RH was investigated in an attempt to reduce the histamine-releasing activity inherent to most potent antagonists while retaining high antiovulatory activity. The protected parent analogue was prepared by conventional solid-phase peptide synthesis. After selective removal of the Lys Fmoc side-chain protection, the resin-bound peptide was readily and conveniently alkylated at the epsilon amino group with various aldehydes and ketones in the presence of NaCNBH3.

Use of the cutaneous anaphylactoid test to detect differences in mast cell mediator-releasing activity among LHRH peptides.

The cutaneous anaphylactoid test was evaluated as a screen for assessing mast cell mediator-releasing (MCMR) activity of luteinizing hormone-releasing hormone (LHRH) peptides. As expected, LHRH and D-Trp6-LHRH, an agonist of LHRH, were nonreactive at concentrations of antagonist which induced pronounced skin lesions. Differences between LHRH and antagonists were obvious, but it was difficult to detect differences among antagonists using arithmetic means.

Antagonistic analogs of luteinizing hormone-releasing hormone are mast cell secretagogues.

The histamine-releasing activity of luteinizing hormone-releasing hormone (LHRH) antagonistic analogs has been documented. Antagonists of LHRH elicited in vitro histamine release from mast cells obtained from previously unexposed rats. Intradermal injection of the antagonists caused increased local skin permeability.

Adenohypophyseal response to hypophysiotropic hormones in male obese Zucker rats.

Description of the recessive, homozygote obese Zucker rat (fafa) includes disorders of growth and reproduction. The aim of this study was to compare responsiveness of adenohypophyseal cells, obtained from male fafa rats and from their lean siblings, to growth hormone-releasing factor (GRF) and to luteinizing hormone-releasing hormone (LHRH). Pituitary cells were cultured for 4 days and were then challenged with either GRF-29 (the NH2-terminal 29 amino acid GRF peptide that expresses full biological activity of its parent 44 amino acid molecule) or [D-Trp6]LHRH (LHRH-A, an LHRH agonist).

Improved antagonists of luteinizing hormone-releasing hormone modified in position 7.

The structure-activity relationship of position 7 in the antagonist [N-Ac-D-Nal1,D-pClPhe2,D-Trp3,D-Arg6,D-Ala10]-LH-RH has been investigated by the incorporation of a series of amino acids at this position. The analogues were prepared by solid-phase peptide synthesis. All purifications were performed in two stages: gel permeation followed by preparative reversed-phase high-performance liquid chromatography.

Direct and indirect inhibition of ovulation in rats by an antagonist of luteinizing hormone-releasing hormone.

The importance of a central or peripheral site of action in the blockade of ovulation induced by LHRH antagonists was investigated. To determine if hCG could induce ovulation in cycling animals given antiovulatory doses of 10 or 100 micrograms [N-Ac-D-p-Cl-Phe1,D-p-Cl-Phe2,D-Trp3,D-Arg6,D-Ala10] LHRH, the antagonist was injected at 1200 h of proestrus followed by varying doses of hCG at 1600 h. hCG did induce ovulation in animals given the analog, which suggested that the antagonist acted primarily at the hypothalamus-pituitary level to inhibit ovulation; however, 20 IU hCG induced ovulation in all animals given 10 micrograms antagonist, but 40 IU were required for the same effect in animals injected with 100 micrograms.

An extremely sensitive in vitro model for elucidating structure-activity relationships of growth hormone-releasing factor analogs.

An improved rat anterior pituitary primary cell culture technique for studying GH-releasing activity of human pancreatic GH-releasing factor (hpGRF) and its analogs is described. Male pituitaries, dispersed by a combination of trypsin digestion and mechanical agitation, were plated at a density of 200,000 cells per well and cultured for 4 days. The attached cells were then stimulated with synthetic hpGRF which was comprised of the first 29 residues of the larger, originally isolated forms and which was amidated at the C-terminal (hpGRF-29).

Administration of human pancreatic growth hormone-releasing factor (GRF) analogs enhances responsiveness of cultured rat pituitary cells to GRF.

The aim of this study was to investigate whether anterior pituitary responsiveness to human pancreatic growth hormone-releasing factor containing 29 amino acids (GRF-29) can be modulated by GRF-29 itself. Male rats were injected (sc) daily for 3 days with 50 ug of GRF-29, or were treated twice daily for 14 days with 5 ug of [D-Ala-2]-GRF-29 (a potent GRF agonist). Control animals were injected with saline.

Synthesis and biological activity of LH-RH antagonists modified in position 1.

As part of our studies on the design of more potent antagonists of the LH-RH (luteinizing hormone-releasing hormone) decapeptide, twelve new highly soluble D-Arg6-analogs have been synthesized. These peptides contain modifications in position 1 and are typified by the general formula (N-acetyl-X1, D-p-Cl-Phe2, D-Trp3, D-Arg6, D-Ala10) LH-RH. We have found that a lyophilic, aromatic substituent is required in position 1 in order to elicit antiovulatory activity at a dose as low as 3 micrograms.

Suppression of ovulation in the rat by an orally active antagonist of luteinizing hormone-releasing hormone.

A synthetic antagonist of luteinizing hormone-releasing hormone blocked ovulation in rats in a dose-dependent manner when given by gavage on the afternoon of proestrus. Ovulation was delayed for at least 1 day in all animals given 2 milligrams of antogonist and in some of the animals treated with 1 or 0.5 milligram.

Peptide antagonists of LH-RH: large increases in antiovulatory activities produced by basic D-amino acids in the six position.

It has been assumed, usually with good reason, that D-amino acids with large aromatic side-chains must be present in position 6 of both LH-RH superagonists and antagonists for the highest levels of biological activity to be reached. However, using one of a recent generation of potent lH-RH inhibitory analogs as a model, we have found that the insertion of D-lysine or, better still, D-arginine in this position results in greater antiovulatory activity in the rat over corresponding D-phenylalanine6- and D-tryptophan6-analogs. For instance, [Ac-D-p-Cl-Phe1,2,D-Trp3,D-Arg6,D-Ala10]-LH-RH exhibits antiovulatory activity at a dose of 750 ng per animal and appears to be the first competitive antagonist with activity in the nanogram region in vivo.

LH-RH antagonists: further analogs with ring-substituted aromatic residues.

Although the systematic substitution of benzene and other aromatic ring systems with various atoms and groups has been a standard approach in conventional pharmaceutical research, it has only recently received the attention it deserves in peptide research. The observation that D-p-Cl-Phe inserted in position 2 of certain LH-RH (luteinizing hormone-releasing hormone) analogs results in large improvements in antagonist activity led us to examine the effect of this and other substituents on position 1 and 2 D-phenylalanyl analog side chains. Analogs containing two D-p-Cl-phenylalanines were found to be particularly powerful competitive inhibitors when assayed in cycling rat for blockade of ovulation.

Failure of gonadotropins to induce in vitro maturation of mouse oocytes treated with dibutyryl cyclic AMP.

In sufficient concentration, dibutyryl cAMP (DBC) prevents the spontaneous in vitro maturation of mouse oocytes. The effects of luteinizing hormone (LH) and follicle stimulating hormone (FSH) on this inhibition were tested in an oil-free chamber-slide culture system. Mouse oocytes devoid of cumulus cells were incubated in the presence of DBC and/or gonadotropins.