Biomedical subjects

R Havouis

Publications and source records attributed to R Havouis.

At least 19 recordsLinked to original sources

PubMed2000

Polyamines are unevenly distributed within the rat and rabbit kidney.

Aliphatic polyamines have generally been measured on the whole kidney. Since the kidney is composed of a variety of cells, whole organ data are of limited value for the interpretation of the functions of the polyamines. The aim of this study was to establish the distribution pattern of putrescine, spermidine and spermine within the kidneys of male and female rats and rabbits. It is shown that the polyamines are unevenly distributed along the cortico-papillary axis. Each amine exhibited its own distinct distribution pattern. The polyamines are predominantly located in the cortex. Putrescine levels increased gradually from the cortex to the papillary tip in rabbits, whereas, in rats, fluctuations in putrescine level were marked. In the six zones of the rabbit kidney studied, spermidine and spermine concentrations were markedly higher in females than in males. This difference was less marked in rats.

Animals↗

PubMed2000

Polyamine profiles in tumor, normal tissue of the homologous breast, blood, and urine of breast cancer sufferers.

Polyamines are involved in the development of breast cancer. We assayed polyamines in erythrocytes, urines, and breast tissues (tumor tissue and histologically normal breast tissue close to the tumor) of patients with invasive breast cancer (n = 174) and benign breast disease (n = 71, used as controls). Polyamine levels in red blood cells and urine were similar to the polyamine concentrations found in healthy subjects, and thus cannot be used as diagnostic markers of breast cancer. In cancer tissue, polyamines were significantly increased in comparison with the polyamine concentrations in controls, and were correlated to the tumor aggressiveness as evaluated by histological grade and Ki-67 proliferative index. On the other hand, correlation was found between polyamine levels in the tumor and the status of the hormone receptors. In the mammary tissue close to the cancer, polyamines dramatically decreased in comparison with the polyamine levels of tissue samples removed around the histologically proven benign tumors. The changes of the polyamine concentrations in the histologically normal breast tissue in the vicinity of the cancer could play a role in the cancer development and need further studies, especially if polyamines are considered as a potential therapeutic target in breast cancer.

Adenocarcinoma↗

PubMed1998

Polyamine sulfonamides with NMDA antagonist properties are potent calmodulin antagonists and cytotoxic agents.

N1-Dansylspermine and related sulfonamides of the natural polyamines are very potent blockers of NMDA-type glutamate receptors. They exhibit pharmacological properties which were not predicted from the constituents of the conjugates. Cytotoxicity and calmodulin antagonism of N1-dansylspermine were especially impressive. Calmodulin antagonism implies that N1-dansylspermine prevents induction of ornithine decarboxylase and inhibits its own active uptake via the polyamine transport system. Structure-activity considerations demonstrated that an aromatic character of the substituent is not required; amide bond formation with an aliphatic sulfonic acid is sufficient to transform spermine into a highly toxic calmodulin antagonist. Cytotoxicity and calmodulin antagonism are properties which are intrinsic to spermine, but they are observed only at very high concentrations. Amide bond formation at N1 with a lipophilic residue appears to 'amplify' these normally latent properties. The use of polyamine conjugates structurally related to the amides described in this work for targeting tumours may be marred by their calmodulin antagonism.

3',5'-Cyclic-AMP Phosphodiesterases↗

PubMed1997

Tumour growth modifies intravascular polyamine transport by plasma lipoproteins in the mouse.

Polyamines are polycationic compounds which are implicated in cell division and tumor growth. We have evaluated the potential role of plasma lipoproteins in the transport of major polyamines, spermine, spermidine and putrescine, and the effect of tumor growth on such transport. Plasmas of healthy male BL6/DBA2 mice and of mice bearing Lewis lung carcinoma (3LL) were fractionated by isopycnic density gradient ultracentrifugation, and polyamine content determined in lipoprotein fractions. Spermidine was the most abundant polyamine in the lipoproteins of both control and tumor-bearing mice and was principally associated with HDL (d: 1.046-1.136 g/ml); approx. 40% of total plasma polyamines was lipoprotein-associated in control mice and 60% in cancerous mice. Only minor amounts were transported by LDL (< 10% of total lipoprotein-associated polyamines), while VLDL were devoid of these substances. Marked elevations of circulating levels of LDL were found in 3LL grafted mice: in these particles however, the contents of spermidine and spermine were significantly reduced. A preferential uptake of polyamines by red blood cells could in part explain this marked reduction of LDL polyamine content, but the consequence of this reduction on the net electrical charge and biochemical function of LDL remains unknown. Elevations of plasma LDL and HDL levels in 3LL-grafted mice underlie the finding that only minor modification was detected in the putrescine content of these particles. However, it is evident that elevated total amounts of putrescine were present in the plasma of such animals. Finally, the density profile of polyamines was modified in cancerous mice in which a shift to transport in lighter apo.AI-containing HDL particles was observed for spermidine; an even more marked shift was found for spermine. In conclusion, our data demonstrate that HDL particles constitute the major plasma vehicle for polyamine transport in both control and in tumor-bearing mice.

Animals↗

PubMed1997

Polyamine deprivation prevents the development of tumour-induced immune suppression.

Mice grafted with the 3LL (Lewis lung) carcinoma exhibit immune suppression: spleen cells showed decreased spontaneous interleukin 2 (IL-2) production and T-CD4+ and T-CD8+ lymphocyte populations; in addition the polyamine content in the spleen was increased. By treating the mice with a polyamine-deficient diet containing neomycin, metronidazole and inhibitors of ornithine decarboxylase and polyamine oxydase, tumour growth was reduced and the immune abnormalities were reversed. The spleen cells overproduced IL-2 by reducing exogenous sources of polyamines, but total blockade of all major polyamine sources was necessary to obtain an optimal effect both on IL-2 production and on spleen polyamine content. Irrespective of whether polyamine deprivation was started at an early or at an advanced stage of tumour growth, T-lymphocyte populations were restored to normal values, demonstrating that polyamine deprivation not only prevents tumour-induced immune suppression, but reverses established immunological disorders. In contrast to what was observed regarding IL-2 production by spleen cells and natural killer (NK) cell activity, the polyamine oxidase (PAO) inhibitor did not enhance the number of T lymphocytes. These findings are consistent with a direct effect of the polyamines on immune effector cell metabolism. They suggest an important role of the gastrointestinal polyamines and of PAO activity in the regulation of IL-2 production.

Animals↗

PubMed1997

Bis(7-amino-4-azaheptyl)dimethylsilane: a new tetramine with polyamine-like features. Effects on cell growth.

Bis(7-amino-4-azaheptyl)dimethylsilane is a new type of aliphatic polycation with structural features resembling those of spermine. The elongation to a seven-membered carbon chain in which the central CH2-group was substituted by (CH3)2Si renders the molecule more lipophilic than spermine. Cells accumulate the compound via the polyamine transport system. Due to suppression of ornithine decarboxylase, de novo putrescine biosynthesis is impaired, and intracellular putrescine and spermidine concentrations are depleted. Depletion of intracellular spermidine together with binding to functionally important polyamine binding sites are considered the main features of the compound which cause cytostatic, and at high concentrations, cytotoxic effects.

Animals↗

PubMed1997

Dimethylsilane polyamines: cytostatic compounds with potentials as anticancer drugs.

Several dimethylsilane tetramines [homologs of spermine with an Si(CH3)2 group in the central carbon chain], a carbon analog of the dimethylsilane tetramines [containing C(CH3)2 instead of Si(CH3)2] and a dimethylsilane hexamine were studied with regard to their cytotoxic activity and their ability to interact with double-stranded DNA. All polyamine analogs exerted cytostatic effects to several cell lines at micromolar concentrations. Their ability to condense DNA was comparable to and their ability to displace ethidium bromide from binding to DNA was superior to that of spermine. Their cytostatic effect was not correlated with the depletion of cellular spermidine concentrations. It is suggested that the new polyamine analogs act mainly by displacing spermidine from binding sites which are essential for the promotion of cell growth.

Animals↗

PubMed1996

Bis(7-amino-4-azaheptyl)dimethlysilane and bis(7-ethylamino-4-azaheptyl)dimethylsilane: inhibition of tumor cell growth in vitro and in vivo.

Bis(7-amino-4-azaheptyl)dimethylsilane (AzhepSi) and its bis(ethyl) derivative [bis(7-ethylamino-4-azaheptyl)dimethylsilane] (EtAzhepSi) are the first examples of a new type of aliphatic tetramine with a dimethylsilane group incorporated into the central carbon chain. AzhepSi shares certain properties with the natural polyamines, but in contrast with spermidine and spermine it inhibits the growth of L1210 leukemia cells in culture at micromolar concentrations. The bis(ethyl) derivative of AzhepSi was made, in analogy to bis(ethyl) spermine, a polyamine derivative, which gained much attention during the last decade as a potential anticancer drug. Chinese hamster ovary (CHO) cells accumulate the dimethylsilyl tetramines considerably more and are more sensitive to these drugs than are CHO-MG cells, a polyamine uptake-deficient mutant. This and related observations demonstrate that AzhepSi and EtAzhepSi are preferentially taken up by a polyamine transport system. Both tetramines inhibit the growth of a variety of tumor cells at micromolar concentrations. AzhepSi turned out to be either equipotent or more potent, but in no case less potent than EtAzhepSi. When given alone at daily doses of 25 micromol/kg, the compounds did not prolong the survival time of L1210 leukemia mice. However, in combination with 2-(difluoromethyl)ornithine and neomycin, AzhepSi had a significant effect on the life span of the animals. The growth rate of 3LL Lewis lung carcinoma was reduced by both compounds at daily doses of 25 micromol/kg. The observations presented in this work suggest that the dimethylsilyl tetramines are antiproliferative agents in vitro and in vivo. Due to enhanced general toxicity, the introduction of N-ethyl substituents was of no advantage in the case of these polyamine analogues.

Animals↗

PubMed1996

In vivo, synergestic inhibition of MAT-LyLu rat prostatic adenocarcinoma growth by polyamine deprivation and low-dose cyclophosphamide.

Polyamine deprivation in vivo produces significant tumor growth inhibition of the hormone-resistant, metastatic Dunning Mat-LyLu murine prostatic carcinoma. In order to produce a cytotoxic effect in addition to the cytostatic effect of polyamine deprivation, various chemotherapy regimens, combined with drug-containing polyamine-deficient chow (DC-PDC), were assessed. Triple chemotherapy combining methotrexate, cyclophosphamide and vindesine; and monochemotherapy with high-dose cyclophosphamide (90 mg. kg-1) and low-dose cyclophosphamide (20 mg.kg-1) were studied alone and in combination with DC-PDC. A variant of DC-PDC excluding the polyamine oxidase inhibitor MDL 72527 was also studied in combination with low-dose cyclophosphamide. The triple-chemotherapy regimen alone or in combination with polyamine deprivation was effective on tumor growth inhibition but was also toxic. High-dose cyclophosphamide alone produced significant tumor growth inhibition and an increase in life span. High-dose cyclophosphamide in combination with DC-PDC was also effective on tumor growth but was also toxic. Low-dose cyclophosphamide alone was moderately effective on tumor growth inhibition with a marginal increase in life span. When combined with polyamine deprivation, results with low-dose cyclophosphamide compared favourably with those of high-dose cyclophosphamide alone and prevented the formation of lung metastases. The polyamine oxidase inhibitor does not appear to be mandatory to achieve this effect if DC-PDC is combined with low-dose cyclophosphamide. Polyamine deprivation appears to be an important tool in anticancer therapy, allowing the use of reduced doses of cytotoxic agents with the same antitumoral efficacy.

Adenocarcinoma↗

PubMed1996

Polyamine deprivation provokes an antalgic effect.

It is well established that inhibition of putrescine formation using D,L-2-(difluoromethyl)ornithine and feeding a polyamine-deficient diet together with non-absorbable antibiotics (neomycin and metronidazole), prevent almost completely the growth of tumors in rats. A similar regimen given to patients with prostate cancer not only reduced the titer of prostate specific antigen in serum, but surprisingly provoked at the same time an antalgic effect. This observation led us to study the potentiation effect of polyamine deprivation on pain threshold in healthy rats. Animals were fed for 2 weeks with an artificial diet of known polyamine content, in combination with antibiotics and 2-(difluoromethyl)ornithine, and were then submitted to pain stimuli using two models, the Randall-Selitto test and the Tail-Flick test. Polyamine deprivation produced in these models an increase in the latency of the response, even under conditions which did not produce significant changes of the polyamine concentrations in blood and brain. From these observations, we may conclude that the polyamines play a role in the perception of nociceptive stimuli under physiological conditions.

Analgesia↗

PubMed1996

Red blood cell polyamines, anaemia and tumour growth in the rat.

In rats with Mat Lylu prostatic carcinoma, significant changes in blood composition and red blood cell (RBC) characteristics were observed. Anaemia, characterised by a decrease in the number of RBC and the reduction of haemoglobin and the iron content in plasma, was correlated with tumour size and the accumulation of spermidine and spermine in the RBC. In large tumours, spermidine levels were increased by 8-fold over normal value. Spleen weight and splenic spermidine concentrations were enhanced in animals with tumours. After splenectomy, the rate of tumour growth decreased by 30%. It is proposed that anaemia in tumour-bearing animals is caused by enhanced RBC lysis, owing to the alteration of the rheological properties of RBC. These may be caused by the alterated surface characteristics due to polyamine accumulation. RBC lysis and high concentrations of polyamines in RBC and spleen appear, not only to favour tumour growth, but also to compromise the immunological defence mechanisms against neoplastic invasion.

Anemia↗

PubMed1996

Observations with an S-adenosylmethionine decarboxylase inhibitor in rats with prostatic adenocarcinoma.

CGP 48664A (2-(4-aminoiminomethyl)-2,3-dihydro-1H-inden-1-ylidene dihydrochloride, CAS 149400-88-4) is a new potent inhibitor of S-adenosylmethionine decarboxylase with antitumor properties. In view of the eminent clinical problems in the treatment of non hormone dependent prostatic cancer, the antiproliferative potency of this compound was tested in Dunning MAT-LyLu rat prostatic adenocarcinoma. The compound proved inefficient in preventing the growth of this tumor, even at a near toxic dose. A reason for the lacking effect is presumably the inadequate accumulation of the drug in the tumor cells due to the excessive growth rate of the MAT-LyLu xenografts. Tumor growth seems not to be accompanied by a proportionally rapid vascularization of the tumor mass. CGP 48664A was found to be a potent inhibitor of polyamine oxidase. This property of the drug may have contributed to the activation of the phagocytic capacity of peritoneal macrophages from tumor-bearing rats.

Adenocarcinoma↗

PubMed1995

Transforming growth factor-beta stimulates arginase activity in macrophages. Implications for the regulation of macrophage cytotoxicity.

Macrophage arginine metabolism via nitric oxide (NO) synthase and arginase pathways reduces and enhances tumor cell proliferation, respectively. Transforming growth factor-beta (TGF-beta) has been shown to down-regulate the NO synthase pathway. The present study describes the effect of TGF-beta on the arginase pathway. TGF-beta up-regulated arginase activity in rat peritoneal macrophages as assessed by measuring the generation of [14C]urea from [14C]-L-arginine in the presence of NG-monomethyl-L-arginine (L-NMMA). The stimulation, which reached fivefold after a 48-h exposure of macrophages to 10 ng/ml TGF-beta, was due to reduction in Km value of arginase. TGF-beta-induced up-regulation of arginase activity led to the release of more polyamines, mainly putrescine. The role of this up-regulation on macrophage cytotoxicity toward L-929 tumor cells was analyzed in coculture experiments. Macrophages blunted DNA synthesis by L-929 cells as assessed by measuring the incorporation of [3H]TdR into the cells and the proportion of cells in the G2 phase. Addition of TGF-beta in the presence of L-NMMA permitted L-929 cells cocultured with macrophages to resume DNA synthesis. The mechanism responsible for this restoration was the up-regulation of arginase activity rather than the down-regulation of NO synthase activity since TGF-beta in the presence of L-NMMA failed to further reduce NO synthase activity whereas it still enhanced arginase activity; synthetic putrescine (1-10 microM) also blunted macrophage cytotoxicity toward L-929 cells. This is the first evidence that TGF-beta up-regulates arginase activity in macrophages and, hence, limits macrophage-dependent cytostasis.

Animals↗

PubMed1995

Three dimensional culture of pineal cell aggregates: a model of cell-cell co-operation.

Three dimensional (3-D) cultures of pineal cell aggregates were obtained by constant gyratory shaking the heterogenous cell populations, obtained from the rat pineals, in the DMEM (Dulbecco's modified Eagle's medium). Within 4 days, the pineal cells became organized into a tissue like configuration appearing as a compact ball, evidenced by the scanning electron microscopy. The 3-D aggregates seemed to be mainly composed of pinealocytes (round-oval cells), glial (elongated cells) and other unknown cells. The heterogenous cells were separated by intercellular spaces. The ultrastructural characteristics revealed by transmission electron microscopy exhibited the presence of granular lysosomes, typical of pinealocytes actively involved in the secretion. These pineal cell aggregates secreted melatonin and other indole amines i.e. 5-methoxytryptamine (5-MT), indole acetic acid (IAA), 5-methoxy-3-indole acetic acid (5-MIAA), tryptophol (TOL) and 5-methoxytryptophol (5-MTL) in the culture medium, indicating the functional aspect of pinealocytes. The 3-D aggregates cultures had advantages over the pineal monolayer cultures as, after 4 days of culture, the amounts of indole amines secreted by 3-D aggregates were higher than those secreted by monolayer cultures. Besides, the 3-D aggregates remained functional till 24 days in the gyratory culture conditions. In the continuous perifusion system, the 3-D aggregates secreted melatonin while challanged with isoproterenol. This 3-D model of pineal cell aggregates might be useful, in future, to perform other kinetic studies of the release of indole amines in perifusion experiments as this system allows the maintenance of pineal cells for a long period of time.

5-Methoxytryptamine↗

PubMed1994

Alterations in intestinal uptake of putrescine and tissue polyamine concentrations in tumor-bearing rats.

Intestinal absorption of putrescine and tissue metabolism of polyamines were investigated in rats grafted with the rapidly growing Mat-Lylu prostatic tumor. These animals exhibited a dramatic 21% decrease in weight and protein, but not DNA, content of their intestinal mucosa, relative to healthy rats reared under similarly controlled nutritional conditions. No significant variation in the specific activities of intestinal brush-border membrane enzymes was observed, however, suggesting a comparable differentiation state of intestinal cells exists in both groups. Putrescine uptake by brush-border membrane vesicles prepared from cancerous or healthy rat intestine was a time dependent process at 25 degrees C. Equilibrium uptake was much greater than could be explained by equilibration of the vesicle space with putrescine, indicating that the diamine was bound to membrane sites. Kinetics of putrescine uptake at 2 min revealed that the process involves two components, a saturable Michaelis-Menten carrier and passive diffusion. With respect to the kinetic parameters of putrescine transport, no significant changes were observed between the tumor-bearing and the control rats. After correction for nonspecific binding to the membranes, putrescine accumulation at equilibrium (75 min) was concentration-dependent and fit a single-site saturable model. Maximum accumulation of the diamine at equilibrium (Bmax) was increased by more than 46% in the cancerous rats relative to the controls, but the dissociation constant (Kd) was unchanged. Efflux of putrescine from the vesicles was slightly slower in the tumor-bearing group, but the differences were generally not significant. No change was observed with respect to the specific activity of ornithine decarboxylase and the concentration of polyamines in the intestinal mucosa. In Mat-Lylu grafted rats fed a standard diet supplemented with [14C]putrescine, about 19% of body radioactivity was recovered in the tumor within 24 h. This was concomitant with a decrease in the percentage of radioactivity retained in the intestinal, renal and hepatic tissues, relative to that retained in the same tissues of healthy rats. Our findings indicate that the presence of the tumor evolves an adaptive response in the small intestine of the rat, involving an increased capacity of the brush-border membrane to accumulate putrescine.

Adenocarcinoma↗

PubMed1994

Transport and metabolism of polyamines in wild and multidrug resistant human leukemia (K 562) cells.

Multidrug resistance (MDR) can be defined as the resistance of cancer cells not just to chemotherapeutic agents to which they have been exposed but also to other apparently unrelated compounds. This MDR phenotype is commonly associated with the high expression of levels of 170 kDa P-glycoprotein, encoded by MDR genes. In the present study, the uptake kinetics of polyamines and their biosynthesis were studied in wild and multidrug resistant (MDR) K 562 cells in culture. The rate (Vmax) of polyamine uptake was significantly lower in MDR cells than that in wild type cells, whereas the Km for the uptake was not significantly different in these cells, suggesting that polyamine transporter is not modified in MDR cells, though their different physiological state influences the uptake process. In a 32 h chase, the transported radioactive polyamines were gradually interconverted. [14C]putrescine was converted into [14C]spermidine following between 15 min and 32 h of culture, and into [14C]-spermine after 16 h of culture, in both the cell types; however, the levels of interconverted radioactive polyamines were always lower in MDR cells as compared with wild type cells. Similarly, internalized [14C]spermidine was converted into [14C]spermine, but not into [14C]putrescine in both the cells types. [14C]spermidine is metabolized into [14C]spermine after 4 h of culture in wild type cells, whereas in MDR cells the interconversion of [14C]spermidine into [14C]spermine is seen only after 16 h of culture. Blocking of the transmembrane drug efflux pump, expressed in the MDR cells, by preincubation in the presence of verapamil, did not influence the uptake of either of the two polyamines (putrescine and spermidine) by MDR cells. On the contrary, this kind of preincubation of wild type cells in the presence of verapamil significantly increased the uptake of these two polyamines. The levels of intracellular polyamine contents in MDR cells were always lower than those in the parental cell line. These results demonstrate that MDR cells are defective in both the uptake of polyamines and their biosynthesis as compared with wild type cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1994

[Involvement of polyamines in malignant proliferative processes: antineoplastic effects of a polyamine deficiency].

The fact that tumors require polyamines for growth has been repeatedly demonstrated. In vivo polyamines are available both from endogenous (intracellular biosynthesis) and exogenous sources (food and intestinal microflora). We investigated in rats grafted with Mat-Lylu prostatic adenocarcinoma the distribution between tumor and tissues of orally administered [14C]putrescine (Pt). The amount of radioactivity retained by tumors was directly proportional to the tumor volume. In a tumor of 25 cm3 19% of the totally retained radioactivity was found. The accumulation of Pt by intestinal brush-border membrane vesicles prepared from tumor-bearing animals was significantly higher than by vesicles from healthy rats. Our results indicate that the presence of a tumor induces an adaptive response in the small intestine which stimulates the uptake of exogenous polyamines. Our therapeutic strategy was to realise a total blockade of all endogenous and exogenous sources of polyamines by feeding animals with a drug (DFMO, MDL 72527, antibiotics) containing polyamine deficient chow. We observed that polyamine deprivation largely reduced both primary tumor and metastatic development. Natural Killer cell cytotoxic activity and blood formula were restored to normal values after treatment. Furthermore polyamine deprivation enhanced antitumoral efficacy of chemotherapy.

Animals↗

PubMed1992

Characterization of metformin transport system in NIH 3T3 cells.

The biochemical properties of the metformin transport system were studied in NIH 3T3 cells. 14C-metformin uptake appeared to be a sodium dependent process. Iso-osmotical replacement of Na+ by choline chloride in the assay medium resulted in a decrease of metformin uptake. Amiloride (200 microM) inhibited the metformin transport by 35% in these cells. Gramicidin, a channel ionophore, was the most effective in inhibiting the metformin transport as compared to valinomycin, a mobile ion carrier, and Ca2+ ionophore (A 23187). Loading of cells with asparagine, ornithine, or polylysine did not influence the uptake process. However, the addition of lysine or arginine significantly stimulated the metformin uptake by NIH 3T3 cells. Similarly, the addition of metformin stimulated the arginine uptake by these cells, suggesting that metformin shares the y+ transport system. Metformin inhibited competitively the uptake of 14C-spermidine, a molecule of the polyamine family, by NIH 3T3 cells, whereas the latter failed to influence the uptake of the former significantly by these cells. Incubation of NIH 3T3 cells in the presence of difluoromethyl-ornithine (a suicidal inhibitor of polyamine biosynthesis) stimulated the spermidine, but not the metformin, uptake by these cells. Interestingly, a prolonged incubation of these cells in the presence of metformin failed to down-regulate the spermidine transport process. The spermidine- and methylglyoxal-bis(guanylhydrazone), MGBG-transport deficient (3T3MG) cells which do not accumulate exogeneous spermidine or MGBG, took up 14C-metformin. However, 14C-metformin uptake by 3T3MG cells was lower than that by normal NIH 3T3 cells.

3T3 Cells↗