Biomedical subjects

N Kenny

Publications and source records attributed to N Kenny.

22 records · Page 2Linked to original sources

PubMed1989

Morphometric quantification of mitochondria in the two steroidogenic ovine luteal cell types.

Progesterone secretion is regulated by different mechanisms in large and small steroidogenic ovine luteal cells. Large cells secrete approximately 7-fold more progesterone in an unstimulated state than small cells. Since cholesterol side-chain cleavage, which is catalyzed by an inner mitochondrial membrane enzyme complex, is a major rate-limiting step in progesterone synthesis, mitochondrial components were quantified in the two steroidogenic cell types throughout the estrous cycle. Corpora lutea collected on Days 4 (n = 4), 8 (n = 4), 12 (n = 5), and 16 (n = 6) of the estrous cycle were prepared for electron microscopy. Volume densities of cell types within corpora lutea and mitochondrial densities within cell types were estimated by point-counting; nuclear and cytoplasmic volume densities were estimated by planimetric analysis. A total of 570 micrographs (magnification 5300 X) were analyzed. Large cell volume density was unchanged during the cycle (35 +/- 1%) while small cell volume density increased (p less than 0.05) from 13 +/- 1% on Day 4 to 20 +/- 3% on Day 12. Large cell mitochondrial volume density increased (p less than 0.05) from 13 +/- 1% on Day 4 to 23 +/- 1% on Day 16 accompanied by an increase in cytoplasmic volume density such that nuclear to cytoplasmic ratio increased (p less than 0.05) from 1:14 to 1:34 between Days 4 and 16. Small cell mitochondrial volume density increased from 11 +/- 1% on Day 4 to 14 +/- 1% (p less than 0.05) for the rest of the cycle while the nuclear to cytoplasmic ratio remained at 1:14.(ABSTRACT TRUNCATED AT 250 WORDS)

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PubMed1987

Progesterone production, LH receptors, and oxytocin secretion by ovine luteal cell types on days 6, 10 and 15 of the oestrous cycle and day 25 of pregnancy.

Corpora lutea were collected from sheep on Days 6, 10, and 15 of the oestrous cycle and Day 25 of pregnancy and dissociated into single cell suspensions. Purified preparations of large and small luteal cells were prepared by elutriation on all days except Day 6. Basal progesterone production by large cells was 6-8-fold higher than by small cells (36-65 vs 6-9 fg/cell/min). Oxytocin secretion was maximal on Day 6 (1.0 fg/cell/min) and declined thereafter. The number of receptors for LH increased between Day 6 and Day 10 and the two cell types had an equal number of receptors on Days 10 and 15 (19,000-23,000). Large cells on Day 25 of pregnancy had fewer receptors (12,000) than did small cells (26,000). Progesterone secretion by small luteal cells from all days examined was stimulated by LH (0.01-1000 ng/ml) in a dose-dependent manner; maximum sensitivity to LH occurred on Day 10. Despite the presence of receptors for LH on large cells, LH failed to stimulate progesterone production. Basal production of progesterone by large and small cells, and the response of small cells to LH, was not influenced by day examined. Re-combinations of large and small cells from Day 10 synergized to increase progesterone secretion. Prostaglandin E-2 (0.1-1000 ng/ml) did not stimulate progesterone secretion by large or small cells.

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PubMed1986

Prostaglandin F2 alpha-induced functional luteolysis: interactions of LH, prostaglandin F2 alpha and forskolin in cyclic AMP and progesterone synthesis in isolated rat luteal cells.

The acute antigonadotrophic action of prostaglandin F2 alpha (PGF2 alpha) was examined in dispersed luteal cell preparations from immature superluteinized rat ovaries. Cell suspensions prepared by collagenase digestion and purification over a Percoll density gradient were incubated for 1 h in Eagle's minimum essential medium in the presence and/or absence of LH, PGF2 alpha, N6,O2'-dibutyryladenosine 3',5'-cyclic monophosphate (dbcAMP) and forskolin. Medium was assayed for total progesterone and adenosine 3',5'-cyclic monophosphate (cAMP). Luteal cell preparations showed typical steroidogenic (progesterone) responses to LH, mimicked by both dbcAMP and forskolin. Whilst the threshold LH dose to increase cAMP synthesis was greater than that for progesterone (100 micrograms/l compared with 1 microgram/l), 24 mumol forskolin/l was the threshold dose for both cAMP and progesterone responses. Furthermore, combined doses of LH and forskolin synergistically raised cAMP yet produced less than additive increases in progesterone. Similarly, combinations of dbcAMP plus forskolin produced less than additive progesterone increases. These data suggest that forskolin may not act as a simple mimic of LH. Prostaglandin F2 alpha dose-dependently inhibited forskolin-induced cAMP and progesterone synthesis and also inhibited progesterone synthesis induced by dbcAMP. These data suggest that the antigonadotrophic effect of PGF2 alpha has more than one locus of action, i.e. it both inhibits an adenylate cyclase event associated with cAMP generation and blunts the cellular response to cAMP. The present uncertainty over the exact locus of forskolin's action within the adenylate cyclase complex limits further delineation of the inhibitory action of PGF2 alpha on LH-responsive adenylate cyclase.

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PubMedDate not supplied

Spontaneous apoptosis of cells prepared from the nonregressing corpus luteum.

At the end of a nonconception estrous cycle, the sheep corpus luteum undergoes involution (luteolysis), a process thought to involve apoptotic deletion of cells. It is not yet clear which of the heterogeneous luteal cell types is involved or what mechanisms drive the apoptotic progression. We examined intact paraffin-embedded corpora lutea (in situ terminal dUTP nick end-labeling method) and found direct evidence for apoptotic deletion of cells during luteolysis, but not in healthy, nonregressing corpora lutea. We then sought to implement in vitro models to dissect apoptotic mechanisms in the constituent cells of the corpus luteum. Cells prepared using standard collagenase dispersion of corpus luteum were evaluated for evidence of apoptosis (DNA laddering) by direct agarose gel electrophoresis, a method that obviates the need for DNA extraction, so allowing examination of relatively few cells (< or = 0.5 x 10(6)). When cells were prepared from nonregressing corpus luteum for in vitro manipulation, a population(s) of cells undergoing spontaneous apoptosis was detected. Apoptosis was inhibited by Zn2+ (5 mM), by the tyrosine phosphatase inhibitor sodium orthovanadate (100 microM), or by maintenance at 4 degrees C. It appears that simple collagenase digestion of intact corpus luteum removes a subset of constituent cells from their survival signal, leading to rapid initiation of endonuclease activity and apoptotic cell death. Identification of the required survival factors and their actions is being pursued to facilitate development of appropriate in vitro models for this endocrine system.

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