Hans-Jörg Ruoff

Intrinsic and Acquired Drug Resistance in Malignant Tumors

Drug resistance is the major reason for failure in cancer chemotherapy. Resistance may be either pre-existent (intrinsic resistance), or induced by drugs (acquired resistance), So far, no strategy has been found to overcome resistance, which is based on highly complex and individually variable biological mechanisms. In present clinical practice, drug resistance can only be recognized during treatment, after long lag times. Thus diagnostic tests are re quired, indicating resistance at an earlier stage, in order to avoid unnecessary medication, frequently associated with toxic side-effects. A number of new anti-cancer drugs are now available. In contrast to the unspecifically acting cytostatic chemotherapy, these compounds have targeted actions. However, as recent studies have shown, resistances and severe side-effects can also be found with targeted drugs. With the increasing number of new treatment regimens, the early diagnosis of resistance will optimize therapy, and indeed will be indispensable for individual cancer therapy. The resistance assays available for use in clinical practice should be integrated into cancer therapy. Research into this neglected area needs to be intensified.

Modulation of rat gastric mucosal prostaglandin E2 release by dietary linoleic acid: Effects on gastric acid secretion and stress-induced mucosal damage

We studied chronic intake of diets deficient in or supplemented with linoleic acid to determine whether it affects gastric acid secretion, release of prostaglandin E2, and stress-induced lesions. For 8-10 wk rats were fed three dietary regimens supplying 3.5% (control group), 0.3%, and 10% of total calories as linoleic acid. We found that diets deficient in linoleic acid (0.3%) reduced release of prostaglandin E2 into the gastric lumen (-77%) and increased basal (+133%) and pentagastrin-stimulated acid secretion (+93%) and the area of cold restraint-induced gastric mucosal lesions (+280%), when compared with the control group. Diets supplemented with linoleic acid (10%) increased prostaglandin E2 release into the gastric lumen (+106%) and reduced basal (-44%) and pentagastrin-stimulated acid secretion (-78%) and the area of cold restraint-induced mucosal.lesions (-80%). Prevention of these lesions by the 10% linoleic acid diet was confirmed by quantitative histology. Pretreatment with indomethacin (8 mg/kg intraperitoneally) abolished the effects of the 10% linoleic acid diet on prostaglandin formation, acid secretion, and mucosal injury. We conclude that in rats chronic intake of dietary linoleic acid reduces acid secretion and prevents cold restraint-induced mucosal lesions, possibly because of augmented synthesis of endogenous prostaglandins in the gastric mucosa.

Comparison of the effect of PGE2 and somatostatin on histamine stimulated14C-aminopyrine uptake and cyclic AMP formation in isolated rat gastric mucosal cells

In isolated rat gastric cells somatostatin and PGE2 were compared in respect to their effects on the cAMP system and on the histamine-stimulated H+-production, measured by14C-aminopyrine (14C-AP) uptake. Like PGE2 somatostatin activated adenylate cyclase (AC) for all in non-parietal cells. This effect on AC declined in cell fractions with increasing number of parietal cells. Activation of AC or elevation of cellular cAMP and uptake of14C-AP in response to histamine were inhibited by 10−9 to 10−5 mol/l PGE2 and somatostatin. The results indicate remarkable similarity between somatostatin and PGE2: both activate a non-parietal cell AC and both inhibit H+-production, likely by interfering at the histamine sensitive AC of the parietal cell.

Adenylate cyclase and H+ production of isolated rat parietal cells in response to glucagon and histamine.

The effect of glucagon and its interaction with histamine on adenylate cyclase (AC), cellular cAMP and [14C]aminopyrine ( [14C]AP) uptake, a reliable index of parietal cell H+ production, was studied in isolated rat gastric cells. AC activation in response to glucagon and histamine correlated with the number of parietal cells. Glucagon (10(-10)-10(-6) mol/l) increasingly stimulated AC (maximal effect: 92% by 10(-7) mol/l) and cellular cAMP (86% by 10(-9) mol/l) of fractions enriched with 80% parietal cells but did not cause a pronounced change of the histamine-stimulated enzyme. If there was any interaction, the effect of both hormones was additive. Glucagon neither changed basal [14C]AP uptake nor interfered with that in response to histamine. The data suggest that if glucagon activates a parietal cell AC this process is not followed by parietal cell H+ production. Furthermore, unlike other inhibitors such as somatostatin or PGE2, glucagon does not reduce acid secretion via the cAMP system of the parietal cell.

Cellular origin and release of intrinsic factor from isolated rat gastric mucosal cells

The cellular content and secretion of intrinsic factor was measured by [57Co]cyanocobalamin binding using isolated rat gastric mucosal cells. The intrinsic factor/R-protein ratio was above 9:1 as evaluated by specific anti-intrinsic factor antibodies. In unfractionized cells with 23 +/- 1.3% parietal cells the intrinsic factor content of 148 +/- 47 fmol/10(6) cells remained almost unchanged over 3 h, whereas basal secretion rose up to 57 +/- 10. In fractionized cells (Percoll) with 3-85% parietal cells most intrinsic factor was found in the parietal cell-depleted fraction (content: 441 +/- 30, secretion/3 h: 139 +/- 16, mean formation/h: 50 +/- 12 fmol/10(6) cells). The intrinsic factor content of the different cell fractions correlated with that of pepsin. [14C]Aminopyrine uptake, an indirect measure of parietal cell H+ production, was inversely related. Carbachol (1 X 10(-6)-10(-3) mol/l) stimulated intrinsic factor secretion, 1 X 10(-3) mol/l being maximally effective (90 +/- 8% above basal). This response was inhibited by atropine and pirenzepine, but not by prostaglandin E2 (PGE2) and somatostatin. Dibutyryl cyclic adenosine monophosphate (dibutyryl cAMP, 43 +/- 7%) and hexoprenaline (24 +/- 5%) enhanced intrinsic factor secretion less effectively and pentagastrin like histamine lacked any stimulatory effect. We conclude that in the rat intrinsic factor is produced and released from chief cells mainly under cholinergic control.

A calmodulin antagonist inhibits histamine-stimulated acid production by isolated rat parietal cells ☆

The role of calmodulin in the regulation of histamine-stimulated parietal cell function was studied in isolated rat parietal cells using [14C]aminopyrine uptake as a quantitative index of acid production. In enriched (77-87%) intact parietal cells the calmodulin antagonist naphthalene sulfonamide W 7 dose-dependently inhibited the response to 10(-4) M histamine (IC50: 2 X 10(-6) M). The mechanism of this inhibition was examined further with two other stimuli of H+-production: forskolin which directly activates the parietal cell adenylate cyclase without interacting at the histamine H2-receptor and dbcAMP which mimics the biological action of cAMP without preceding activation of adenylate cyclase. W 7 effectively inhibited the responses to 10(-4) M forskolin (IC50: 6 X 10(-7) M), 10(-3) M dbcAMP (IC50: 10(-6) M) and to 10(-2) M K+ (IC50: 3 X 10(-6) M). The action of W 7 followed non-competitive kinetics since the antagonist reduced the entire range of the concentration-response curves without shifting them rightwards towards higher concentrations of the respective stimulants. The effect of W 7 was reversed by washing the cells. ATP-induced [14C]aminopyrine uptake into digitonin-permeabilized oligomycin-inhibited parietal cells reflects H+-production independent of oxidative phosphorylation and was also inhibited by W 7 (IC50: 10(-5) M). Inhibition of K+-stimulated H+/K+-ATPase activity required even higher W 7-concentrations (IC50: 1.4 X 10(-4) M). Our data suggest that calmodulin might be involved in the intracellular mediation of the response to histamine. Between histamine-induced cAMP-generation and the H+-secreting tubulovesicular system W 7 seems to inhibit an intracellular step that finally activates the H+/K+-ATPase. Yet, direct inhibition of the ATPase requires W 7 concentrations of questionable specificity and is unlikely to be the mechanism behind the action of W 7 on the parietal cell response to histamine.

Effects of hormones (calcitonin, GIP) and pharmacological antagonists (ranitidine and famotidine) on isolated rat parietal cells

The rationale for the present study was to compare calcitonin and gastric inhibitory polypeptide (GIP) versus two histamine H2 receptor antagonists with respect to their potency of inhibiting parietal cell functions. Adenylate cyclase activity and acid production ([14C]aminopyrine uptake) of isolated rat parietal cells were stimulated by histamine. At 10(-7) and 10(-6) mol/l, calcitonin and GIP reduced the response to histamine by 10-20% following noncompetitive kinetics. Ranitidine and famotidine (MK 208) inhibited the response to histamine by about 50% at 10(-7)-10(-6) mol/l, and at 10(-5) mol/l abolished the histamine effect. On a molar basis famotidine turned out to be 6 times more potent than ranitidine. Both antagonists revealed competitive kinetics. Our data suggest direct inhibition of the parietal cells by the tested compounds which were shown to interfere at the adenylate cyclase cAMP system or at the histamine H2 receptor. However, compared to the histamine H2 receptor antagonists, hormonal inhibition is less pronounced and mediated by a different mechanism.

Intrinsic factor secretion from isolated human gastric mucosal cells.

Human gastric mucosal cells were isolated from the resected fundic mucosa of peptic ulcer patients. The intracellular content and secretion of intrinsic factor were estimated by binding to cyano[57Co]cobalamin. The content was maximal in the enriched parietal cell fraction which also displayed the highest H+ production as measured by amino[14C] pyrine uptake. Secretagogues evoked full response after 15 min of incubation: pentagastrin (181% of basal secretion), carbachol (208%), histamine (250%) and dibutyryl cyclic adenosine monophosphate (304%). The phosphodiesterase inhibitor isobutylmethylxanthine was slightly more effective even than dibutyryl cAMP. The response to histamine was abolished by ranitidine, indicating activation of adenylate cyclase via histamine H2 receptors, but remained unaffected by atropine, which in turn blocked the carbachol effect, whereas ranitidine was ineffective. The mean formation rate was 8.4 fmol intrinsic factor/10(6) cells per h under basal conditions and 14.3 fmol in response to histamine.

Histamine-sensitive adenylate cyclase in human gastric mucosa: Cellular localization and interaction by PGE2, somatostatin and secretin

Introduction According to studies with isolated and enriched parietal cells of dog gastric mucosa the intracellular accumulation of cAMP in response to histamine seems to be localized in parietal cells, whereas the accumulation in response to prostaglandins is in nonparietal cells [1, 2]. The present study describes the cellular localization of histamine and PGE2-sensitive adenylate cyclase (AC) in human gastric mucosa and the effect of the gastric acid secretion inhibitors, PGE2, somatostatin and secretin, on the histamine-activated enzyme.

Effect of pentagastrin on gastric mucosal nucleic acid synthesis of fed guinea pigs.

In normally fed guinea pigs the effect of a sustained release pentagastrin preparation was studied on gastric mucosal weight, protein, DNA and RNA concentration and thymidine and uridine incorporation into DNA and RNA, respectively. Treatment with pentagastrin for 3 days significantly increased antral fundic mucosal DNA and RNA concentration and thymidine and uridine incorporation. It is concluded that pentagastrin stimulates gastric mucosal nucleic acid synthesis beyond the driving force elicited by food.