Involvement of histamine in growth of mouse and rat tumors: antitumoral properties of monofluoromethylhistidine, an enzyme-activated irreversible inhibitor of histidine decarboxylase.

The present study suggests that newly synthesized histamine is involved in the development of some animal tumors (e.g., Lewis lung carcinoma in mice and Morris hepatoma in rats). A marked induction of histidine decarboxylase (HDC) and an increase in the histamine concentration were observed in the tumors approximately 1 week after inoculation, and there were parallel increases in ornithine decarboxylase activity and the concentrations of polyamines. The H2 receptor antagonist, cimetidine, significantly reduced tumor growth in the animal models while the H1 receptor antagonist, dexchlorpheniramine, had no effect, suggesting that histamine could act via H2 receptor sites. Extensive depletion of tumor histamine induced by local injection of Compound 48/80 did not result in a significant cytostatic effect. Monofluoromethylhistidine (MFMH), an enzyme-activated irreversible inhibitor of HDC, retarded the growth of hepatoma tissue culture cells grown in culture, and when infused s.c. at 60 mg/kg/day it greatly inhibited the development of tumors induced i.m. by hepatoma tissue culture cells in Buffalo rats. MFMH also had pronounced antitumoral effects on EMT6 sarcomas and Lewis lung carcinomas in mice, which were associated with inhibition of HDC and depletion of the histamine content of the tumors. These cytostatic effects were clearly enhanced when MFMH was combined in therapy with the specific ornithine decarboxylase inhibitor, DL-alpha-difluoromethylornithine. The antitumoral effects of the combination were associated with marked decreases in the tumor histamine and putrescine contents. It is proposed that nascent histamine, like newly synthesized putrescine and spermidine, plays a role in the rapid proliferation of animal tumors. Inhibition of HDC by essentially nontoxic drugs such as MFMH could represent a novel approach to the control of neoplastic growth.

Plasma membrane transglutaminase and cornified envelope competence in cultured human keratinocytes.

When confluent cultures of the transformed human keratinocyte line SV-K14 are shifted to serum-free medium the cells achieve, within 4 days, the ability to synthesize a cornified envelope after challenge with the Ca2+ ionophore A23187. During these 4 days the enzyme transglutaminase (EC 2.3.2.13), which catalyses the cross-linking of different envelope precursor proteins, is partially transferred from the cytosolic pool into the plasma membrane. The association of the enzyme with the plasma membrane proves to be an essential step in the envelope formation since a direct correlation between plasma membrane-bound transglutaminase and envelope competence is observed. Retinoids block the insertion of the enzyme and therefore prevent envelope formation.

Properties of partially purified bovine brain dopamine beta-hydroxylase.

Dopamine beta-hydroxylase has been partially purified from bovine brain. A 140-fold purification factor was achieved using solubilization with Triton X-100, ammonium sulphate fractionation between 20-50 per cent saturation, affinity chromatography on concanavalin A-Sepharose 4 B and then filtration through Sephadex G200. The specific activity at the end was 51 nmoles/h/mg protein. The majority of endogenous inhibitors were lost. Immunological studies, kinetic studies, studies on the interaction with lectins and the effect of carboxylic acids on enzyme activity were carried out. Our data are in favour of the close similarity between the bovine brain and adrenal enzymes. No major differences could be found, at least with the characterization experiments using in the present study.

alpha-Fluoromethyl histidine. Inhibition of histidine decarboxylase in pylorus ligated rat.

alpha-Fluoromethyl histidine is an irreversible inhibitor of histidine decarboxylase. The injection of a single dose to pyloric-ligated rats inhibits gastric mucosal histidine decarboxylase in a dose-dependent manner but does not modify histamine content and gastric acid secretion even at the highest dose used. Administration of cimetidine increases histidine decarboxylase activity, decreases histamine level in gastric mucosa and inhibits gastric acid secretion. The co-administration of alpha-fluoromethyl histidine blocks the augmentation in enzyme activity, maintains lowered histamine level and prolongs the antisecretory action of cimetidine.

Histamine receptor blockade (H2) versus inhibition of histamine synthesis in stress ulceration in rats.

Cold and restraint stress in rats induced gastric mucosa lesions, increased gastric mucosa histidine decarboxylase activity and elevated hypothalamic content. Cimetidine did not modify the biochemical effects induced by stress but partially protected against gastric ulceration. alpha-Fluoromethyl histidine inhibited the increase of histidine decarboxylase activity in the gastric mucosa, inhibited the rise of hypothalamic histamine content and decreased the incidence of gastric lesions to the same extent as did cimetidine.

The rhino mouse model: the effects of topically applied all-trans retinoic acid and CD271 on the fine structure of the epidermis and utricle wall of pseudocomedones.

The histological and ultrastructural effects following 3 weeks' topical treatment with two agents (all-trans retinoic acid and a new synthetic retinoid-like substance, CD271) were evaluated on the epidermis and the epithelial wall of the pseudocomedones in rhino mouse skin. The comedolytic effects of these drugs were similar, and consisted of a reduction of the utricular diameter, with normalization of follicular units. Morphological examinations revealed a hyperplastic response with an increase in the number of cell layers of both epidermis and follicular epithelium, and modifications in keratinocyte differentiation. Ultrastructural changes in the epidermis and epithelial wall were observed mainly in the granular and horny layers, with increased desquamation, and a decrease in the cohesiveness of corneocytes. During the first week of treatment, some cutaneous toxic effects were noticed, but they normalized within two weeks. On the other hand, a fine granular material persisted in the intercellular spaces. It is confirmed that the skin of the rhino mouse is a good model for the evaluation of the comedolytic effects of drugs. Moreover, it reveals the specific effects of retinoids on epidermal differentiation. We have demonstrated that topically applied CD271 induces modifications similar to those obtained with all-trans retinoic acid. It is thus concluded that CD271 is a potentially effective anti-acne agent.

Effect of alpha-monofluoromethyl histidine, an irreversible inhibitor of histidine decarboxylase, on gestation in mice.

Chronic administration of alpha-monofluoromethyl histidine, a specific inhibitor of histidine decarboxylase, to pregnant mice blocked histidine decarboxylase activity and markedly depleted histamine levels in 18-day-old foetuses and in newborn mice. Such treatment had no effect on implantation or embryonic development or parturition suggesting that de novo synthesis of histamine is not a significant factor in these processes in the mouse.

Topical retinoids. Their uses in dermatology.

The retinoids provide an important new way of treating dermatologic disorders. They have also proved to have a role in the prevention of new lesion formation. New retinoids, of which adapalene is one, have recently been synthesized in order to obtain similar or better efficacy while reducing skin irritation potential. These new molecules are currently under clinical investigation. Preliminary results are encouraging. In the near future, an expanded range of topical retinoids should be available.

Induction of ornithine decarboxylase activity in hairless rat epidermis as a pharmacological model: validation of the animal model.

We use a mutant hairless Sprague Dawley rat to evaluate the capacity of retinoids to inhibit the epidermal ornithine decarboxylase activity induced by sellotape stripping. In order to minimize the variability introduced by the animals in our model we decided to validate the hairless rats used. A number of animal parameters were examined using a single lot of 50 males and 50 females aged from 4 to 11 weeks and acclimatized to laboratory conditions. The body weight growth curves were established. Nude animals present two periods of hair growth, the first at 6-7 weeks and the second at about 10-11 weeks. Hair development is more pronounced in males. No histological change was observed in the stratum corneum but an increase in epidermal thickness was noted in males aged 9 weeks. Removal of the stratum corneum by sellotape stripping was more effective and reproducible in the females, as determined histologically. Sellotape-stripping induction of ornithine decarboxylase in the epidermis was higher in rats aged 5-6 weeks and reached a plateau in animals aged 6-12 weeks. Individual variations obtained were lower in females (about 5%-10% in females and 10%-20% in males). The present research suggests that female rats aged about 8 weeks provide maximum reproducibility of response and ease of use.

Comparative study of plasma dopamine-beta-hydroxylase activities in noradrenaline-secreting and adrenaline-secreting phaeochromocytomae.

1. Circulating dopamine-beta-hydroxylase activities were measured in two hypertensive patients, with a phaeochromocytoma tumour. Tumours were found to differ by their secretory properties, one secreting noradrenaline, the other adrenaline. After removal of the tumour, the plasma dopamine-beta-hydroxylase activities in both patients gradually decreased to reach a stable value in correlation with urinary, excreted catecholamine levels. The only difference was the rate of the plasma enzyme activity decrease. 2. Thus, it appeared that some phaeochromocytomae are able to secrete dopamine-beta-hydroxylase in addition to catecholamines. Therefore, dopamine-beta-hydroxylase activity measurements may be of interest: (1) in determining secretory properties of phaeochromocytoma tumours, and (2) in following the evolution during the postoperative period.

Differential effects of calcium channel antagonists on histamine and pentagastrin-stimulated gastric acid secretion in the rat.

Different calcium channel antagonists have been assessed for their ability to inhibit gastric acid secretion in rats. Whereas verapamil, diltiazem and cinnarizine inhibited pentagastrin-induced gastric acid secretion but not histamine-induced secretion, nifedipine selectively inhibited the stimulant effect of histamine. In contrast, the vasodilator hydralazine had non-selective effects. These findings indicate that calcium-antagonists may have differential effects against different secretagogues and these effects are not simply related to hypotensive effects.

Inhibition of histamine biosynthesis and gastric function in the rat: effect on pentagastrin-induced gastric acid secretion.

A very simple and rapid method to measure gastric acid secretion in the conscious unoperated fasted rat is reported. Antisecretory activity of cimetidine and hypersecretory activity of pentagastrin are detected by this procedure. Pentagastrin induces gastric acid secretion and causes an increase by 2-fold in histidine decarboxylase activity. There is a delay between the increase of gastric acid output and the appearance of histidine decarboxylase activity. Co-administration of monofluoromethyl histidine, a specific irreversible inhibitor of histidine decarboxylase, does not modify the peak response to pentagastrin but shortens the duration of stimulation. We suggest from this data that histamine biosynthesis is needed for maintenance of elevated gastric acid secretion.

Stereochemistry of reactions catalysed by mammalian-brain L-glutamate 1-carboxy-lyase and 4-aminobutyrate: 2-oxoglutarate aminotransferase.

Deamination of 4-aminobutyrate by mammalian or bacterial 4-aminobutyrate aminotransferases involves the abstraction of the pro-S hydrogen on C-4 of 4-aminobutyrate. Decarboxylation of L-glutamate by rat brain glutamate decarboxylase occurs with retention of configuration. Inhibition of this enzyme by (S)-4-aminohex-5-ynoic acid involves the abstraction of the proton at C-4 of the inhibitor. On the basis of this finding, we postulate the existence of an abnormal reaction of glutamate decarboxylase in which the proton at C-4 of (S)-4-aminohex-5-ynoic acid is removed in a manner similar to the one which normally occurs in enzymatic transaminations of L-amino acids. This reaction is presumably facilitated by the acetylenic group adjacent to the eliminated proton.

Radiosensitization of human tumor cells by alpha-difluoromethylornithine.

The effect of polyamine depletion on the radiosensitivity of a human tumor cell line was investigated. CAL 18 A cells, derived from a breast carcinoma, were incubated with alpha-difluoromethylornithine (DFMO)--a specific and irreversible inhibitor of ornithine decarboxylase (ODC)--at a 1 mM or 10-mM concentration for either 1 hr or 24 hr and irradiated thereafter. Survival curves of exponentially growing cells revealed a moderate but significant enhancement of radiosensitivity as compared to untreated irradiated cells. Maximum radiosensitization was observed at a concentration of 10 mM after 1 hr incubation. Plateau-phase cells were used to study the effect of polyamine inhibition on repair of radiation-induced potentially lethal damage (PLD). DFMO enhanced the radiation response and significantly inhibited PLD repair in these cells. Measurement of ODC indicated that this enzyme was markedly inactivated upon brief incubation of CAL 18 A cells with DFMO, reflecting a depletion of polyamine synthesis. These results extend findings that have demonstrated enhancement of drug-induced cytotoxicity, and raise the possibility of clinical use of this substance for potentiation of radiation response.

A rapid and simple test system for the evaluation of the inhibitory activity of retinoids on induced ornithine decarboxylase activity in the hairless rat epidermis.

In mouse skin, antiproliferative agents including retinoids inhibit induction of ornithine decarboxylase activity by a variety of hyperproliferative stimuli. In the hairless rat skin, ornithine decarboxylase activity was induced by ten successive strippings with cellotape and by topical application of 12-O-tetradecanoyl-phorbol-13-acetate. Topical application of all trans-retinoic acid (25 nmol/cm2) immediately after the tape stripping of the skin significantly inhibited the induction of ornithine decarboxylase activity at all time points measured. The inhibition by all trans-retinoic acid of ornithine decarboxylase induced by cellotape stripping was dose dependent as was found to be the case for arotinoid, retinol, Ro-10-1670, motretinid, 13-cis-retinoic acid, etretinate, and vitamin A. Oral administration of all trans-retinoic acid also inhibited the ornithine decarboxylase activity induced by cellotape stripping. We propose the assay of ornithine decarboxylase activity in the hairless rat epidermis after tape stripping for a rapid evaluation of new retinoids.

Effect of prolonged inhibition of histidine decarboxylase on tissue histamine concentrations.

In rats, chronic infusion of alpha-fluoromethyl histidine, a selective irreversible inhibitor of mammalian histidine decarboxylase, caused a marked depletion of histamine in all tissues examined. There were no gross pharmacological effects associated with this depletion.

Hexadecane-induced skin hyperplasia in the hairless rat: time course of histological and biochemical events related to the synthesis of polyamines and DNA.

Several biochemical parameters including ornithine decarboxylase activity (ODC) and tissue polyamine levels were measured during the hexadecane-induced epidermal hyperplasia of hairless rat skin. Animals received three applications of 200 microliters pure n-hexadecane on day 1. ODC activity and polyamine levels (putrescine, spermidine and spermine) in the epidermis were significantly increased and reached maximum elevations at 12 h after the start of n-hexadecane treatment with DNA synthesis peaking at 24 h. Histological studies confirmed a significant cellular edema at 24 h after the beginning of the treatment followed at 48 h by an epidermal hyperplasia which was maximum at 72 h. These data support the view that ODC activation, increased biosynthesis of polyamines and DNA are early events in epidermal cell hyperproliferation.

Repeated application of dinitrochlorobenzene to the ears of sensitized guinea pigs: a preliminary characterization of a potential new animal model for contact eczema in humans.

We have evaluated a subchronic model of contact hypersensitivity in the guinea pig to mimic human chronic/recurrent eczema. Repeated challenges of the ears of previously sensitized guinea pigs with 0.1% dinitrochlorobenzene (once a week for 4 weeks) induced a typical oedema response, which increased during the first 48 h after each challenge. Crusts were detectable (48 h after challenge) and histological observations (72 h after challenge) revealed hyperplasia, papillomatosis, hyperkeratosis and some mononuclear cell infiltrates in the dermis. In agreement with clinical observations in humans, topical treatment of challenged animals with corticosteroid (1% hydrocortisone) reduced the oedema, hyperplasia, papillomatosis, and leucocyte infiltrates, while application of 5% bufexamac (a non-steroidal drug) was associated with a slight enhancement of the inflammatory response. Thus, this model presents clinical and histological similarities with human eczema. Its pharmacological relevance is also suggested, although further investigations are required to better define its selectivity.