Biomedical subjects

F R DeRubertis

Publications and source records attributed to F R DeRubertis.

At least 55 records · Page 3Linked to original sources

PubMed1988

Role of activation of protein kinase C in the stimulation of colonic epithelial proliferation by unsaturated fatty acids.

Some, but not all, studies have suggested that high-fat diets promote colon carcinogenesis, possibly by stimulating the proliferative activity of colonic epithelium. Both the increase in colonic excretion of bile salts and of fatty acids that occur with an increase in fat ingestion have been implicated as stimuli of epithelial proliferative activity. In this study, we examined the role of activation of protein kinase C in fatty acid-induced stimulation of colonic epithelial proliferation in the rat. Intracolonic instillation of arachidonate, linoleate, or oleate at concentrations that did not induce surface cell injury or loss increased colonic mucosal ornithine decarboxylase activity and stimulated incorporation of [3H]thymidine into mucosal deoxyribonucleic acid. The saturated fatty acid palmitate was without effect. Arachidonate, linoleate, and oleate each induced the translocation of protein kinase C activity from the soluble fraction to the membrane fraction of colonic mucosa, an index of enzyme activation. The translocation of protein kinase C induced by unsaturated fatty acids occurred both in vivo after intracolonic instillation of these agents and in vitro upon incubation of isolated colonic crypt epithelium with fatty acids. The effects of the unsaturated fatty acids on both enzyme translocation and colonic epithelial proliferative activity were suppressed by 1-(5-isoquinolinyl)-2-methylpiperazine, an inhibitor of protein kinase C activity. Unsaturated fatty acids directly stimulated soluble colonic mucosal protein kinase C activity when added to the enzyme assay mixture. This action was blocked by 1-(5-isoquinolinyl)-2-methylpiperazine. However, unsaturated fatty acids also increased the breakdown of polyphosphoinositides when added to isolated colonic epithelium. The increase in polyphosphoinositide breakdown resulted in release of diacylglycerol, an endogenous activator of protein kinase C. Thus, unsaturated fatty acids may activate protein kinase C of colonic epithelium through either a direct intracellular effect or through an action on the cell membrane. The results support a role for protein kinase C in the stimulation of colonic epithelial proliferation by unsaturated fatty acids.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1988

Role of enhanced arachidonate availability through phospholipase A2 pathway in mediation of increased prostaglandin synthesis by glomeruli from diabetic rats.

Prostaglandin E2 (PGE2) production by superfused glomeruli from rats made diabetic for 2 wk by streptozocin injection is twofold higher than that by glomeruli from normal rats. The higher rates of PGE2 production by glomeruli from diabetic rats are associated with higher levels of labeled free arachidonate in glomeruli prelabeled with [3H]arachidonate, both basally and after stimulation with Ca2+ ionophore A23187. The difference between release of labeled arachidonate from phospholipids of diabetic versus normal glomeruli is likely underestimated by measurements of arachidonate alone due to more rapid incorporation of released arachidonate into triacylglycerol of diabetic glomeruli. A23187 induced a fall in labeled phosphatidylcholine, phosphatidylethanolamine, and phosphatidylinositol in glomeruli that had been prelabeled with [3H]arachidonate and also induced a reduction in the mass of these phospholipids. Consistent with the higher levels of labeled arachidonate, the reduction in both labeled phospholipids and phospholipid mass with A23187 was greater in glomeruli from diabetic than normal rats. Furthermore, the reduction in labeled phospholipids and phospholipid mass with A23187 was largely (62-80%) accounted for by a fall in phosphatidylcholine plus phosphatidylethanolamine in glomeruli from both normal and diabetic rats. These results suggest a primary role for phospholipase A2 in A23187 actions on glomerular arachidonate release in normal rats and for the higher levels of arachidonate found in glomeruli from diabetic rats. Nevertheless, A23187 also stimulated the production of inositol phosphates--a measure of cellular phospholipase C activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1987

Role for protein kinase C in A23187 induced glomerular arachidonate release and PGE2 production.

A high concentration of the calcium ionophore A23187 (10 uM) stimulated release of labeled arachidonate and immunoreactive PGE2 from isolated superfused glomeruli. A lower concentration of A23187 (1 uM) or 12-0-tetradecanoyl phorbol-13-acetate (TPA, 0.1 uM), a direct activator of protein kinase C, when added alone was without effect on these parameters. Combined addition of A23187 (1 uM) and TPA (0.1 uM) synergistically stimulated arachidonate release and PGE2 production. 1-(5-isoquinolinyl)-2-methylpiperazine (H-7) a known inhibitor of protein kinase C in other tissues, suppressed increases in arachidonate release and PGE2 production mediated by A23187 (10 uM) or TPA plus A23187 (1 uM). H-7 inhibited while TPA stimulated protein kinase C activity that had been partially purified from soluble fractions of glomerular homogenates. These results support a role for protein kinase C in A23187 mediated arachidonate release.

Animals↗

PubMed1987

Subcellular distribution of protein kinase C in rat colonic epithelial cells with different proliferative activities.

Activation of Ca2+ and phospholipid-dependent protein kinase C (PKC) is associated with increased proliferation in several cell types. When activated, PKC is tightly bound to the particulate cell fraction. Accordingly, we examined the subcellular distribution of PKC in superficial (nonproliferative) and proliferative colonic epithelial cells from rat colonic mucosa. PKC was determined in soluble and particulate fractions of these cells following partial purification of enzyme activity of cellular homogenates by DEAE-cellulose chromatography. In the superficial cells, 90% of the PKC was associated with the soluble fraction. By contrast only 42% of the enzyme activity was found in the soluble fraction of proliferative cells. The specific activity of protein kinase C was higher in the particulate fraction of proliferative compared to superficial cells when expressed as a function of either particulate protein or cellular DNA content. Addition of deoxycholate or 12-O-tetradecanoylphorbol-13-acetate induced a translocation of protein kinase C from the soluble to the particulate fraction. [3H]Thymidine incorporation into DNA was higher in colonic epithelial cells isolated from the colons of rats which had been exposed to deoxycholate or 12-O-tetradecanoylphorbol-13-acetate in vivo. Treatment of rats with 1-(5-isoquinolinyl)-2-methylpiperazine (H-7) suppressed basal [3H]thymidine incorporation into DNA and increases in this parameter induced by 12-O-tetradecanoylphorbol-13-acetate and deoxycholate. The results are consistent with a positive role for activation of protein kinase C in the control of colonic epithelial proliferation.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1987

Actions of sulfasalazine and 5-aminosalicylic acid as reactive oxygen scavengers in the suppression of bile acid-induced increases in colonic epithelial cell loss and proliferative activity.

Sulfasalazine suppresses mucosal injury in patients with ulcerative colitis, but the mechanism of its therapeutic action is uncertain. In the present study, we examined the mechanism of the protective action of sulfasalazine in a rat model in which colonic epithelial cell loss and subsequent increases in epithelial proliferative activity were induced by intracolonic instillation of sodium deoxycholate. Sulfasalazine or its therapeutically active metabolite 5-aminosalicylic acid suppressed the loss of deoxyribonucleic acid into the colonic lumen and the subsequent increases in mucosal ornithine decarboxylase activity and tritiated thymidine incorporation into deoxyribonucleic acid induced by sodium deoxycholate. Sulfasalazine and 5-aminosalicylic acid also blocked xanthine-xanthine oxidase-induced loss of deoxyribonucleic acid and the subsequent proliferative response. In vitro sodium deoxycholate increased reactive oxygen formation by colonic mucosal scrapings or isolated crypt epithelium. These actions of sodium deoxycholate on reactive oxygen formation were blocked by sulfasalazine or 5-aminosalicylic acid. Sulfapyridine, a therapeutically inactive metabolite of sulfasalazine, had no effect on sodium deoxycholate-induced increases in surface cell sloughing, ornithine decarboxylase, tritiated thymidine incorporation into deoxyribonucleic acid, chemiluminescence, or superoxide production. The ability of sulfasalazine and 5-aminosalicylic acid to scavenge reactive oxygen may play a role in their therapeutic effects of inflammatory bowel disease.

Aminosalicylic Acids↗

PubMed1987

Sequential alterations in glomerular prostaglandin and thromboxane synthesis in diabetic rats: relationship to the hyperfiltration of early diabetes.

The present study examined the role of enhanced production of prostaglandin (PG) E2 and 6-keto-PGF1a, the stable metabolite of PGI2, by glomeruli from streptozotocin diabetic rats in the mediation of hyperfiltration. Correlative measurements of insulin clearance (CIn) and glomerular production of PGE2, 6-keto PGF1a and thromboxane (TX) B2 (the stable metabolite of TXA2) were made at two time points, nine to 15 days and 25 to 28 days after streptozotocin. CIn was elevated by 40% to 50% in diabetic rats studied at nine to 15 or 25 to 28 days compared to values in age-matched controls. Basal production of PGE2, 6-keto PGF1a and TXA2 (as reflected by TXB2) and increases in response to A23187 were elevated in glomeruli from nine to 15-day diabetic rats compared to values in control glomeruli. Exogenous arachidonate abolished these differences. Treatment of nine-day diabetic rats with indomethacin (3 mg/kg/d) rapidly (within 24 hours) and reversibly suppressed CIn without altering CIn in control rats. Indomethacin had no effect on plasma glucose in control or diabetic rats. Treatment of nine to 15-day diabetic rats with insulin (10 U/kg/d by osmotic minipump) beginning 24 hours after streptozotocin lowered plasma glucose to values that were not significantly different from control and prevented the rise in CIn. Treatment of diabetic rats with insulin or incubation of glomeruli from untreated diabetic rats with insulin (0.3 mU/mL) for two hours in vitro reduced basal and A23187 induced increases in PGE2, 6-keto PGF1a, and TXB2 to values that were not different from those in control glomeruli.(ABSTRACT TRUNCATED AT 250 WORDS)

6-Ketoprostaglandin F1 alpha↗

PubMed1987

Relationship of bile salt stimulation of colonic epithelial phospholipid turnover and proliferative activity: role of activation of protein kinase C1.

The mechanism by which bile salts stimulate the proliferative activity of colonic epithelium is uncertain. One of the striking cellular actions of certain bile salts that enhance the proliferative activity of colonic epithelium, such as deoxycholate (DOC) and chenodeoxycholate, is the rapid stimulation of membrane phospholipid turnover. Increased membrane phosphoinositol turnover may lead to release of diacylglycerol (DAG). The latter is an endogenous activator of the calcium phospholipid-dependent enzyme protein kinase C (PKC) whose stimulation has been correlated with enhanced proliferation in several cell systems. In the present study, we examined the effects of DOC on PKC of colonic epithelium in vitro and in vivo. When added directly in vitro to partially purified soluble preparations of phospholipid, calcium-dependent PKC from crypts isolated from rat colon, DOC suppressed activity by 20%, presumably due to calcium complex formation. By contrast, the tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA), and the DAG derivative, 1-oleoyl-2-acetylglycerol (OAG), increased soluble PKC in vitro twofold. The nontumor promoters phorbol and 4 alpha-phorbol-12,13-didecanoate (4 alpha PDD) were without effect. However, in intact colonic epithelial crypt cells prelabeled with arachidonate, DOC caused rapid release of DAG and markedly increased the fraction of PKC associated with the particulate cell fraction, an index of PKC activation. TPA and OAG caused similar shifts in the subcellular distribution of PKC but did not stimulate DAG release, whereas phorbol and 4 alpha PDD were without effect on any parameter. In vivo intracolonic instillation of DOC, OAG, or TPA each induced a shift of soluble PKC to the particulate fraction of colonic mucosal scrapings.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1987

Role of activation of protein kinase C in the stimulation of colonic epithelial proliferation and reactive oxygen formation by bile acids.

Deoxycholate (DOC), chenodeoxycholate, 12-O-tetradecanoyl phorbol-13-acetate (TPA), or 1-oleoyl-2-acetyl-glycerol (OAG) activated colonic epithelial protein kinase C as reflected by translocation from the soluble to the particulate cell fraction. Activation of protein kinase C was correlated with stimulation of enhanced proliferative activity of colonic mucosa and reactive oxygen production. TPA and OAG, but not DOC, directly activated soluble protein kinase C in vitro. However, DOC rapidly increased labeled inositol phosphate and diacylglycerol accumulation in colonic epithelial cells. Retinoic acid inhibited protein kinase C activity and suppressed DOC-, TPA-, and OAG-induced increases in reactive oxygen production. The results support a role for protein kinase C in the stimulation of colonic epithelial proliferative activity and reactive oxygen production induced by bile acids, TPA and OAG. In contrast to TPA and OAG, which activate protein kinase C directly, bile acids appear to activate protein kinase C indirectly by increasing the diacylglycerol content of colonic epithelium.

Animals↗

PubMed1986

Profiles of eicosanoid production by superficial and proliferative colonic epithelial cells and sub-epithelial colonic tissue.

The profile of cyclooxygenase and lipoxygenase products in normal rat colonic epithelium and subepithelium was examined. Colons were thoroughly perfused to eliminate contamination with blood. Two preparations of colonic epithelium were employed. The first consisted of intact colonic crypts and epithelial sheets. The second yielded single cell suspensions of superficial versus proliferative epithelial cells. Lipoxygenase product formation by colonic epithelium as measured by hydroxyeicosatetraenoic acid (HETE) and leukotriene B4 (LTB4) production (5-HETE greater than 12-HETE greater than 15-HETE greater than LTB4) accounted for 58% of the total colonic production of these moieties, whereas epithelium accounted for only 20% of total colonic protein. By contrast, prostaglandin (PG) E2 and PGF2 alpha production occurred predominantly (greater than 97%) in the subepithelial layers. The present studies also demonstrate markedly higher levels of accumulation of lipoxygenase products in proliferative versus superficial epithelial cells, whereas prostaglandin accumulation was greater in superficial cells. Previous studies have supported a role for lipoxygenase and cyclooxygenase products in the control of colonic secretion, inflammatory cell infiltration and proliferative activity. The present results raise the possibility that the striking differences in the sites of production of these products within the colon has functional implications.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗

PubMed1986

Increased urinary excretion of PGE2 during inappropriate antidiuresis induced by DDAVP.

Increased urinary prostaglandin E excretion (UPGE2V) and renal production of PGE2 have been implicated in the mediation of escape from inappropriate antidiuresis induced by vasopressin (AVP). AVP-induced increases in PGE2 have been linked to its pressor activity. We examined the effects of the non-pressor AVP analogue, 1,desamino-8-D-arginine vasopressin (DDAVP), on UPGE2V in a rat model of inappropriate antidiuresis in which the escape response is not apparent. A dose of DDAVP known to cause antidiuresis (10 ng/hr), was given by osmotic minipump to rats during a water diuresis established by ad libitum ingestion of 5% dextrose in water (D/W). This dose of DDAVP reduced plasma sodium (P[Na+] ) to 127 +/- 2 mEq/liter within 24 hr. P[Na+] in controls given 5% D/W alone was 151 +/- 2. In the first eight hr after DDAVP, UPGE2V was fivefold higher than in control animals, an increase which was sustained for 48 hrs. Plasma renin (PRA) was threefold higher in control rats compared to rats that had received DDAVP, consistent with volume expansion in the latter. Despite declines in P[Na+] and PRA and increased UPGE2V in DDAVP treated rats, evidence of escape from inappropriate antidiuresis (secondary decline in Uosm or rise in urine volume (V) or P[Na+] ) was lacking. Moreover, DDAVP induced similar fivefold increases in UPGE2V when given to fluid deprived rats, demonstrating that increases of UPGE2V during inappropriate antidiuresis were not mediated by volume expansion. Administration of indomethacin to volume repleted DDAVP treated rats did not significantly increase Uosmol or decrease V compared to values observed in rats receiving DDAVP alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1986

Unilateral multinodular toxic goiter: scintiscan mimicking solitary toxic nodule.

A 58-year-old man presented with hyperthyroidism. By palpation, the left lobe of the thyroid was diffusely enlarged, and the right lobe was normal. Radioactive iodide scintiscan demonstrated homogeneous uptake that was localized to the enlarged left lobe, with near total suppression of uptake on the right. TSH stimulation led to clear visualization of a normal appearing right lobe, findings most consistent with an autonomously functioning solitary nodule on the left. Left hemithyroidectomy cured the hyperthyroidism. However, no single dominant nodule was found in the left lobe. Rather, there was diffuse thyroidal hyperplasia of the micronodular variety, consistent with multinodular toxic goiter. Thus, in this patient with a usually diffuse form of thyroid disease, the autonomously functioning hyperactive follicles were localized predominantly to one thyroid lobe. This variant expands the clinical spectrum of multinodular toxic goiter, and further emphasizes the extent of asymmetry in the distribution of hyperactive follicles than can occur in this disorder.

Adult↗

PubMed1986

Role of reactive oxygen in bile salt stimulation of colonic epithelial proliferation.

Our previous studies had suggested a link between bile salt stimulation of colonic epithelial proliferation and the release and oxygenation of arachidonate via the lipoxygenase pathway. In the present study, we examined the role of reactive oxygen versus end products of arachidonate metabolism via the cyclooxygenase and lipoxygenase pathways in bile salt stimulation of rat colonic epithelial proliferation. Intracolonic instillation of 5 mM deoxycholate increased mucosal ornithine decarboxylase activity and [3H]thymidine incorporation into DNA. Responses to deoxycholate were abolished by the superoxide dismutase mimetic CuII (3,5 diisopropylsalicylic acid)2 (CuDIPS), and by phenidone or esculetin, which inhibit both lipoxygenase and cyclooxygenase activities. By contrast, indomethacin potentiated the response. Phenidone and esculetin suppressed deoxycholate-induced increases in prostaglandin E2 (PGE2), leukotriene B4 (LTB4), and 5, 12, and 15-hydroxyeicosatetraenoic acid (HETE), whereas CuDIPS had no effect. Indomethacin suppressed only PGE2. Deoxycholate (0.5-5 mM) increased superoxide dismutase sensitive chemiluminescence 2-10-fold and stimulated superoxide production as measured by cytochrome c reduction in colonic mucosal scrapings or crypt epithelium. Bile salt-induced increases in chemiluminescence were abolished by CuDIPS, phenidone, and esculetin, but not by indomethacin. Intracolonic generation of reactive oxygen by xanthine-xanthine oxidase increased colonic mucosal ornithine decarboxylase activity and [3H]thymidine incorporation into DNA approximately twofold. These effects were abolished by superoxide dismutase. The findings support a key role for reactive oxygen, rather than more distal products of either the lipoxygenase or cyclooxygenase pathways, in the stimulation of colonic mucosal proliferation by bile salts.

Animals↗

PubMed1986

Relationship between loss of rat colonic surface epithelium induced by deoxycholate and initiation of the subsequent proliferative response.

Bile acids increase the proliferative activity of rat colonic epithelium. However, the mechanisms responsible are unknown. The present study examined the relationships between deoxycholate (DOC) induced surface cell sloughing, as measured by loss of DNA into the lumen and by light microscopy, and the subsequent increases in mucosal ornithine decarboxylase activity and [3H]thymidine (dThd) incorporation into mucosal DNA induced by deoxycholate. Intracolonic instillation of DOC (10 mumol; 5 mM) resulted in a progressive increase in luminal DNA content which was significant by 1 min and maximal by 1 h. No further increase in luminal DNA occurred between 1 and 4 h after DOC. Similarly, light microscopy demonstrated a progressive loss of surface epithelium between 10 min and 1 h after DOC instillation. By 4 h after DOC, the colonic mucosal surface was normal histologically. The rapid repair of the epithelial surface occurred without a detectable increase in [3H]dThd incorporation into DNA within 4 h. The latter finding thus suggested that upward migration of nondividing crypt epithelial cells rather than the rapid initiation of new DNA synthesis and new mitotic activity was responsible for surface repair. Enhanced proliferative activity of colonic mucosa, as measured by increased [3H]dThd incorporation into DNA, did occur subsequently (12 to 24 h) after instillation of DOC. The dose response of early surface cell loss and the subsequent proliferative response to DOC were identical, consistent with a link between these two DOC mediated events. However, two observations suggested that surface epithelial loss alone was not sufficient to trigger the proliferative response to DOC: intracolonic instillation of DOC followed by removal of the DOC solution at 1 h, at which time surface epithelial loss was maximal, did not result in an increase in ornithine decarboxylase activity or [3H]dThd incorporation into DNA when these parameters were assessed at 4 h or 12 to 48 h, respectively; phenidone, an antioxidant and radical scavenger, and bis[(3,5-diisopropyl-salicylato) (O,O) copper(II), a lipophilic agent with superoxide dismutase activity, abolished the DOC mediated proliferative response but did not prevent the early loss of surface cells. The results imply that events other than or in addition to surface cell loss are necessary for the expression of the action of DOC to stimulate the proliferative activity of colonic epithelium.

Animals↗

PubMed1985

Fatty acid stimulated N-demethylation of 1,2-dimethylhydrazine and tetramethylhydrazine by rat colonic mucosa.

A fatty acid stimulated, NADPH-independent pathway for the N-demethylation of 1,1-dimethylhydrazine (1,1-DMH) with the generation of HCHO was demonstrated in 10,000 g soluble fractions of colonic mucosal homogenates. Tetramethylhydrazine and, to a lesser extent, aminopyrine, but not 1,2-DMH or methylhydrazine, were also substrates for this reaction. Isolated superficial colonic epithelial cells metabolized 1,1-DMH at a faster rate than proliferative epithelial cells. Indomethacin, an inhibitor of cyclooxygenase activity, and 5,8,11,14-eicosatetraynoic acid (ETYA), an inhibitor of both cyclooxygenase and lipoxygenase activities, suppressed HCHO production from 1,1-DMH by 50 and 80%. However, in the presence of indomethacin or ETYA, arachidonate hydroperoxide stimulated HCHO formation. This suggested a peroxidative mechanism for 1,1-DMH metabolism, related in part to prostaglandin synthesis. A possible role for lipoxygenase activity in mediating 1,1-DMH metabolism was suggested by the ability of linoleate, which did not increase prostaglandin synthesis, to stimulate 1,1-DMH metabolism and by the fact that ETYA was more effective than indomethacin as an inhibitor of 1,1-DMH metabolism. The fatty acid stimulated pathway for N-demethylation was clearly distinct from the mixed function oxidase activities. NADPH did not stimulate 1,1-DMH metabolism to HCHO. 7,8-Benzoflavone or SKF-525A, inhibitors of cytochrome P-450, and methimazole, an inhibitor of N-demethylation catalyzed by the hepatic microsomal FAD-containing monooxygenase, did not suppress HCHO formation. To the extent that 1,1-DMH and tetramethylhydrazine reach the colon unchanged, the results suggest that fatty acid stimulated cooxidation pathways in colonic mucosa may contribute to the metabolism of these agents. Metabolism by superficial cells which are destined to slough may be an important defense mechanism against the toxic and carcinogenic actions of these hydrazines in colon.

1,2-Dimethylhydrazine↗

PubMed1985

16,16-Dimethyl prostaglandin E2 suppresses the increases in the proliferative activity of rat colonic epithelium induced by indomethacin and aspirin.

Treatment of rats with indomethacin rapidly increased ornithine decarboxylase (4 h) of colonic mucosa and [3H]thymidine incorporation into colonic mucosal deoxyribonucleic acid (DNA) (1 or 5 days) when this parameter was examined in vivo and ex vivo. The changes in colonic mucosal ornithine decarboxylase and DNA synthesis induced by indomethacin were correlated temporally with suppression of colonic prostaglandin synthesis, as assessed from ex vivo colonic production of prostaglandin E, the dominant prostaglandin product of colon. Autoradiographic studies indicated that the enhancement of proliferative activity of colonic epithelium after treatment with indomethacin for 1 day was confined to the lower third of the colonic crypt (normal proliferative zone). After 5 days of indomethacin treatment, however, there was an extension of the proliferative zone to the upper third of the colonic crypts. Concurrent treatment of rats with the stable prostaglandin E2 analogue, 16,16-dimethyl prostaglandin E2, suppressed indomethacin-induced increases in colonic mucosal ornithine decarboxylase and DNA synthesis. Concurrent administration of 16,16-dimethyl prostaglandin E2 also prevented the extension of the proliferative zone of colonic epithelium induced by 5 days of indomethacin administration. 16,16-Dimethyl prostaglandin E2 alone for 1-5 days had no detectable effects on colonic mucosal ornithine decarboxylase and DNA synthesis compared with corresponding control values. Increases in colonic mucosal DNA synthesis were also induced by treatment of rats for 5 days with aspirin (ASA). The stimulation of colonic mucosal DNA synthesis induced by ASA was significantly suppressed by concurrent administration of 16,16-dimethyl prostaglandin E2 and was also correlated with the inhibition of colonic prostaglandin synthesis by ASA. The colons of rats treated with indomethacin for 1 day or ASA for 5 days appeared normal by light microscopy. However, treatment of rats for 5 days with indomethacin resulted in mild to moderate inflammation of the lamina propria and some goblet cell depletion at the mucosal surface, but no loss of surface epithelium. The ultrastructure of the surface epithelium of the colons of rats treated with indomethacin or ASA was normal as assessed by electron microscopy. The results thus demonstrate that inhibition of local colonic prostaglandin synthesis is associated with increases in the proliferative activity of colonic epithelium, and that these increases are suppressed by administration of 16,16-dimethyl prostaglandin E2.(ABSTRACT TRUNCATED AT 400 WORDS)

16,16-Dimethylprostaglandin E2↗

PubMed1985

Ca2+-dependent modulation of renin release from isolated glomeruli: apparent independence from alterations in cGMP.

This study examined the role of cGMP in the control of renin release from isolated rat glomeruli. An inverse correlation between renin release and cGMP content of isolated glomeruli was found under several conditions of incubation. Thus, incubation of isolated glomeruli in Ca2+-free media containing EGTA, or the addition of the Ca2+ and calmodulin (CaM) antagonists trifluoperazine (TFP), N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamide hydrochloride (W-7), or (8-N, N-diethylamino)-octyl 3,4,5-trimethoxybenzoate-HCI (TMB-8) to glomeruli incubated in Ca2+ replete buffer lowers cGMP and stimulates renin release. These same incubation conditions enhance the release of renin induced by isoproterenol (DBcAMP) in isolated glomeruli. By contrast, raising media K+ to 60 mmol/L, or the incubation of glomeruli with angiotensin II (A-II) or ouabain--all of which are thought to increase intracellular Ca2+--increased glomerular cGMP and suppressed basal glomerular renin release and the increases in renin release induced by isoproterenol (DBcAMP). However, neither exogenous DBcAMP nor nitroprusside, an agent that increased the endogenous cGMP, content of glomeruli mimicked the suppressive effects of high K+, A-II, or ouabain on renin release. Moreover, DBcGMP and nitroprusside also failed to reverse the stimulatory effects of Ca2+ deprivation, TFP, W-7, or TMB-8 on glomerular renin release, even though nitroprusside clearly enhanced cGMP under these conditions of incubation. The results suggest that changes in glomerular cGMP and renin release occur concomitantly in response to alterations in glomerular Ca2+ homeostasis, but that cGMP does not mediate the changes in glomerular renin release.

1-Methyl-3-isobutylxanthine↗

PubMed1985

Fatty acid-stimulated oxidation of methylazoxymethanol by rat colonic mucosa.

The present study examined fatty acid-initiated metabolism of methylazoxymethanol (MAM) to formaldehyde (HCHO) by the 10,000 X g soluble fraction of rat colonic mucosa, and the role of prostaglandin synthase and lipoxygenase activities in mediating this process. Incubation of MAM with soluble fractions of rat colonic mucosa, in the absence of arachidonate, resulted in significant HCHO production compared to that observed in buffer alone or with the heated tissue fractions. Addition of arachidonate (100 microM), linoleate (100 microM), or arachidonate hydroperoxide, but not palmitate, increased HCHO formation by 50%. Indomethacin (25 to 100 microM) suppressed basal and arachidonate-stimulated HCHO production by 25 to 50%. However, indomethacin did not influence linoleate or arachidonate hydroperoxide-induced increases in HCHO. These data suggested a peroxidative mechanism for MAM oxidation, that was mediated in part by arachidonate metabolism via the prostaglandin synthase system. 5,8,11,14-Eicosatetraynoic acid (25 to 500 microM) suppressed HCHO production by 30 to 80% in the absence of fatty acids, and abolished stimulation by arachidonate or linoleate, but not by arachidonate hydroperoxide. MAM was also oxidized by an NAD+-dependent dehydrogenase, as evidenced by MAM-mediated NAD reduction in 10,000 X g soluble fractions of rat colonic mucosa. On a molar basis, the ability of the soluble fraction of rat colonic mucosa to oxidize MAM by the NAD+-dependent dehydrogenase pathway and the fatty acid-stimulated pathway were similar. However, NADPH did not stimulate HCHO formation by MAM. Moreover, 7,8-naphthoflavone; 2-diethylaminoethyl-2,2-diphenylvalerate; and methimazole, inhibitors of mixed-function oxidase activity, did not suppress HCHO formation, implying that MAM was not metabolized by the colonic mixed-function oxidase activity. MAM metabolism to HCHO was 3 to 4 times greater by soluble fractions of superficial epithelial cells isolated from rat colon compared to those of the isolated proliferative epithelial cell pool. The results are consistent with a role for both the prostaglandin synthase and lipoxygenase systems of colonic mucosa in the oxidative metabolism of MAM. Enhanced oxidation of MAM by superficial cells of colonic epithelium which are preparing to slough may serve to protect the colon against the carcinogenic effect of this drug.

5,8,11,14-Eicosatetraynoic Acid↗

PubMed1984

Calcium dependence of the stimulatory action of hypertonicity on renal medullary prostaglandin synthesis.

Previous studies have shown that hypertonic mannitol or NaCl increases the release of [3H]arachidonate and immunoreactive prostaglandin E in inner medullary slices incubated in Ca2+-free media containing EGTA. By contrast, the stimulation of these parameters by ionophore A23187 and by arginine-vasopressin are abolished in Ca2+-free media plus EGTA. In the present study, the effects of Ca2+ deprivation and the intracellular Ca2+ antagonist TMB-8 [8-N,N-diethylamino)octyl-3,4,5 -trimethoxybenzoate-HCl) were further examined to assess the Ca2+ dependence of the actions of different stimuli of prostaglandin E synthesis in rat renal inner medulla. Ca2+-free media without EGTA abolished increases in [3H]arachidonate and immunoreactive prostaglandin E release induced by ionophore A23187, but not those induced by arginine-vasopressin, suggesting that different pools of Ca2+ subserve expression of the actions of these two stimuli. At low concentrations, TMB-8 (10-25 microM) inhibited increases in [3H]arachidonate and immunoreactive prostaglandin E release induced by arginine-vasopressin, but did not influence effects of Ca2+ plus ionophore A23187 or hypertonicity on these parameters. At higher concentrations (100-500 microM), TMB-8 suppressed effects of ionophore A23187, hyperosmolar NaCl and mannitol on immunoreactive prostaglandin E and [3H]arachidonate release from slices. The effects of a sub-optimal inhibitory concentration of TMB-8 on ionophore A23187 actions were overcome by increasing Ca2+ in the media from 1.5 to 5 mM. Ca2+ deprivation, or concentrations of EGTA or TMB-8, that were effective in suppressing increases in immunoreactive prostaglandin E induced by ionophore A23187, arginine-vasopressin or hypertonicity, did not modify increases in immunoreactive prostaglandin E induced by exogenous arachidonate. Moreover, in microsomal fractions of inner medulla, TMB-8 suppressed Ca2+-dependent increases in phospholipase A2 and C activities, an effect which was competitive with Ca2+. Thus, Ca2+ deprivation and TMB-8 act at a step in the immunoreactive prostaglandin E synthetic pathway proximal to cyclooxygenase activity, and probably at the level of Ca2+-dependent acyl hydrolase activity. The results with TMB-8 indicate that an intracellular pool of Ca2+ is involved in expression of the actions of hypertonicity to increase [3H]arachidonate release and immunoreactive prostaglandin E in inner medulla.

Animals↗