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Biomedical subjects

O Alabaster

Publications and source records attributed to O Alabaster.

At least 19 recordsLinked to original sources
PubMed1997

Absence of p53 gene mutations in rat colon carcinomas induced through the synergistic interaction between methylazoxymethanol and X-irradiation.

p53 is one the most frequently mutated genes found in human colonic tumors. Because colonic neoplasms induced in rats by certain chemical carcinogens are similar to human colonic tumors in their histological features and proliferation characteristics, the rat has been used as an experimental model to study the pathogenesis of colon cancer. However, p53 mutations were not detected in the chemically induced colonic tumors analyzed for p53 mutations. X-irradiation has also been shown to induce colonic neoplasms in rats that resemble human colonic tumors histopathologically. Because the incidence of colonic tumors induced by methylazoxymethanol (MAM) in rats was shown to be enhanced by X-irradiation, we immunohistochemically analyzed these colonic carcinomas for the presence of p53 gene mutations. The immunohistochemical analyses clearly showed the absence of nuclear immunoreactivity in all ten tumors examined. The results from the present study indicate that point mutations in p53, at least in the coding region, are not involved in the development of colon cancer induced by the combination of MAM and X-irradiation. Our observations, together with the data from previous studies, further suggest that rat colon carcinogenesis, unlike human colon cancer, may not involve p53 mutation as an obligatory event.

Adenocarcinoma↗
PubMed1997

Inhibition by wheat bran cereals of the development of aberrant crypt foci and colon tumours.

As variation in both type of fibre and its physical properties can influence physiological effects, the effects of different dietary levels (1, 4, 8%, w/w) of unprocessed wheat bran (WB) were compared with those of two of its processed commercial formulations used in breakfast cereals, on the formation of aberrant crypt foci (ACF) and colon tumours in Fischer 344 rats following azoxymethane (AOM) administration. All diets were high in fat (20 g/100 g) and low in calcium (0.2%, w/w). The rats were fed the experimental diets for 2 wk before receiving two sc injections of AOM (15 mg/kg body weight/wk). 8 wk following the first injection of AOM, five rats per group were killed and the formation of ACF was measured. 23 wk following the first injection of AOM, 12 rats per group were killed and the colon tumour incidence in different dietary groups was measured. The results showed that increasing the dietary concentration of fibre from 1 to 8% (w/w), using all the wheat bran formulations, significantly reduced the number of ACF per rat. None of the diets showed any significant effect on the normal growth of rats. No statistically significant differences were observed between the protective properties of WB and the two commercial formulations under investigation in terms of the reduction of the number of ACF, or in terms of the reduction of the colon adenocarcinoma incidence. The results suggest that wheat bran and its two commercial formulations can offer protection against colon cancer even when they are consumed with a high-fat/low-calcium diet. The addition of any of these formulations of wheat bran fibre is likely to be equally effective in the prevention of colon cancer in human populations that habitually consume high-fat/low-fibre Western-style diets.

Animals↗
PubMed1997

Suppression of intestinal polyp development by low-fat and high-fiber diet in Apc(delta716) knockout mice.

Most epidemiological and animal studies show a positive correlation of the dietary intake of fat with the incidence of colon cancer, whereas an inverse correlation of the dietary intake of fiber. In rats fed a diet low in fat and high in wheat bran fiber and calcium, a significant decrease was reported in the number of azoxymethane-induced aberrant crypt foci compared with those fed a high-fat, low-fiber and low-calcium diet. Mutations in the human APC gene play a key role, not only in familial adenomatous polyposis, but also in many sporadic cancers of the entire digestive tract. We previously constructed a mouse strain Apc(delta716), carrying a truncation mutation at codon 716 of the Apc gene, the homolog of human APC (10). The heterozygous mice developed numerous intestinal polyps, and all microadenomas dissected from the earliest polyps had already lost the wild-type allele, indicating the loss of heterozygosity. Using these Apc(delta716) knockout mice, we have investigated the effect of a low-fat and high-fiber diet (LRD for 'low-risk' diet) on intestinal polyposis, and compared it with that of a high-fat and low-fiber diet (HRD for 'high-risk' diet). The mice were fed either diet for 7 weeks, and the number and size of intestinal polyps were scored. The LRD-fed mice had fewer polyps than the HRD-fed mice, by 36% in the small intestine and by 64% in the colon. As for the polyp size distribution, there was no significant difference between the HRD- and LRD-fed mice. These results indicate that LRD can suppress intestinal polyposis compared with HRD which does not, and suggest that its suppression is at the initiation of polyp formation. This is likely to be due to a decreased frequency of loss of heterozygosity, rather than a retarded growth of the polyp adenomas.

Animals↗
PubMed1996

Microsatellite instability in aberrant crypt foci from human colons.

Aberrant crypt foci (ACF) are distinct microscopic lesions of the colon thought to be the earliest identifiable precursors of colon cancer. As precursors of colon cancer, ACF may contain mutations in genes that are altered early in colorectal tumorigenesis. Candidates for these genes include APC, K-Ras, and those of the DNA mismatch repair system. Some colon cancers with mutations in DNA mismatch repair genes are characterized by genomic instability at simple repeated sequences, also known as microsatellite instability. In this study, we analyzed 19 ACF (> or = 20 crypts/focus) and adjoining, microscopically normal colonic mucosa from 10 colon cancer patients for the presence of microsatellite instability. DNA from two ACF from two different patients displayed microsatellite instability. None of the DNA samples from normal mucosa displayed microsatellite instability. These observations support the role of ACF as a precursor to colon cancer and provide some evidence that mutations in DNA mismatch repair genes are early somatic events in colon cancer.

Colon↗
PubMed1996

A rapid dual organ rat carcinogenesis bioassay for evaluating the chemoprevention of breast and colon cancer.

In this study we evaluated the effect of dietary administration of a high fat, low fiber diet (HRD) with or without 2% phytic acid (PA) on the development of mammary cancer and/or colon cancer in rats exposed to methylnitrosourea (MNU), azoxymethane (AOM) or MNU + AOM. The rats were fed a HRD alone or a HRD + 2% PA. At the end of week 2, the rats were given either a s.c. injection of MNU (50 mg/kg body wt) or one of normal saline (vehicle). At the end of weeks 3 and 4, the rats were given either a s.c. injection of AOM (15 mg/kg body wt per week) or one of normal saline (vehicle). Nine weeks after the injection of MNU or saline, 10 rats from each group were sacrificed and the mammary tumor incidence and the number of colonic aberrant crypt foci (ACF) were compared between different groups. The administration of different diets was continued for an additional 21 weeks and the mammary tumor and colon tumor incidence between different groups were compared. Results showed that rats injected with MNU alone did not develop ACF or colon tumors while those injected with AOM alone did not develop mammary tumors. Linear regression analysis of the number of ACF at 11 weeks versus colonic tumor incidence at 32 weeks, and the linear regression analysis of mammary tumor incidence at 11 weeks versus mammary tumor incidence at 32 weeks, both showed good linear correlation. These results demonstrate the potential value of the short term dual organ carcinogenesis bioassay for screening chemopreventive agents for their relative ability to inhibit the development of mammary cancer and/or colon cancer while on high risk diet.

Animals↗
PubMed1996

Dietary fiber and the chemopreventive modelation of colon carcinogenesis.

Comparative international epidemiological data indicate that the difference between the highest and lowest colon cancer incidence is approximately 10-fold. This suggests that the dominant causes of colon cancer are environmental rather than genetic in origin, with the dominant environmental cause being the typical diet of Western industrialized countries. Many epidemiological and experimental studies have suggested an important role for dietary fiber in the prevention of colon cancer. Using the Fischer-344 rat as the experimental model, data clearly demonstrate a strong protective effect of a diet that is low in fat, high in fiber and high in calcium (low-risk diet). Such a diet prevents the development of both preneoplastic aberrant crypt foci (ACF) and colon tumors. Recent experiments have also demonstrated a direct relationship between a ras point mutation in ACF at different stages of rat colon carcinogenesis, and a ras point mutation that is subsequently present in colon tumors. Using wheat bran as the model dietary fiber source, its effects were compared to the effects of psyllium, phytic acid, vitamin E, beta-carotene, folic acid, alone or in combination, for their ability to prevent colon cancer in rats on high-risk Western-style diets. Our studies clearly demonstrated the ability of wheat bran to reduce ACF and colon tumors in rats that consumed high-fat, Western-style diets. Although phytic acid, which is a constituent of wheat bran, alone demonstrated strong cancer-preventive potential, our experiments provided evidence for the cancer-preventive effect of the crude fiber fraction that is independent of the effect of phytic acid. The synergistic combination of wheat bran with the soluble fiber psyllium led to enhanced protection; while the combination of wheat bran with beta-carotene showed only an additive effect. Beta-carotene appeared to show higher protection than wheat bran at an intake level that is nutritionally relevant to humans, suggesting the possibility of using beta-carotene to enhance the effects of dietary fiber in high-risk Western populations. Using ACF as an intermediate endpoint, it was also shown that vitamin E and beta-carotene appear to inhibit progression of ACF to colon cancer, while wheat bran and folic acid appeared to have weak cancer-preventive potential at this late stage of carcinogenesis. In conclusion, wheat bran alone, or in combination with psyllium, appears to have greater potential to inhibit earlier phases of carcinogenesis, while beta-carotene and vitamin E may also inhibit later stages of carcinogenesis. Despite considerable epidemiological and experimental evidence that increasing the fiber and lowering the fat content of the Western diet could substantially reduce the risk of cancer and heart disease, the real challenge is to find effective ways to educate and motivate people to overcome their intrinsic cultural resistance to such changes in their eating habits.

Animals↗
PubMed1996

The effect of dietary fat on the promotion of mammary and colon cancer in a dual-organ rat carcinogenesis model.

Female Sprague-Dawley rats (n = 270) were randomly divided into nine subgroups (30 rats/group) and fed diets with fat contents ranging from 5% to 45% of total energy, in which fat was isocalorically substituted for carbohydrates. At Week 3, the rats were given a single injection of methylnitrosourea (MNU, 50 mg/kg body wt). At Weeks 4 and 5, rats were given an injection of azoxymethane (AOM, 15 mg/kg body wt sc). The rats were maintained on their respective experimental diets until they were sacrificed. Nine weeks after the injection of MNU, 10 rats from each group were killed and their mammary tissue was examined for tumors and their colons for aberrant crypt foci (ACF) formation. The results showed no significant increase in the number of colonic ACF or mammary tumors at 11 weeks among individual groups consuming 5-25% energy from fat or among groups consuming 30-45% energy from fat. Data analysis indicated that the effects of the two diets (5-25% energy from fat and 30-45% energy from fat) on ACF formation as well mammary tumor development were statistically different. Continuation of the diets for 32 weeks led to the development of colonic and mammary tumors. Analysis of the combined group tumor incidence data clearly showed that the colonic and mammary tumor incidence in groups fed 30-45% of energy from fat was significantly higher than that observed among groups fed 5-25% of energy from fat. Linear regression analysis of the colonic ACF at 11 weeks, colonic tumor incidence at 32 weeks, and mammary tumor incidence at 11 and 32 weeks showed good linear correlation, demonstrating the potential value of ACF and mammary tumors at 11 weeks for evaluating the carcinogenic risk associated with different diets. The main conclusion of the experiment is that the incidence of colon and mammary cancers increased rapidly when the dietary levels of fat were increased from 15% to 30% of calories and that there was no significant influence on the colon and mammary cancer risk beyond 30% of calories from fat.

Adenocarcinoma↗
PubMed1995

Absence of p53 gene mutations in rat colon carcinomas induced by azoxymethane.

The K-ras and p53 genes are two of the most frequently mutated genes found in the human colonic tumors. Since azoxymethane (AOM) induced rat colonic neoplasms are similar to human colonic tumors in their histological features and proliferation characteristics, the rat has been used as an experimental model to study the pathogenesis of colon cancer in humans. Although the presence of K-ras point mutations has been reported in AOM induced rat colonic tumors, there are no reports describing the frequency for mutation of the p53 gene in these tumors. In this study, colon adenocarcinomas induced in rats by AOM were examined for the presence of point mutations in exons 5-8 of the p53 gene, using a combination of single strand conformation (SSCP) analysis, immunohistochemistry and direct DNA sequencing. SSCP analysis showed no differences in banding patterns between the normal mucosa and any of the 20 adenocarcinomas analyzed. Nuclear p53 immunoreactivity was absent in all tumors examined. Since p53 point mutations predominate in malignant colonic tumors, five adenocarcinomas with the greatest local invasiveness were analyzed by direct DNA sequencing of exons 5-8 of the p53 gene. Direct DNA sequencing did not reveal mutations in any of the adenocarcinomas analyzed, within the coding region of p53 gene that were sequenced. The results from the present study indicate that point mutations in the p53 gene, at least in the coding region (exons 5-8) are not involved in the development of colon cancer induced by AOM in the rat.

Adenocarcinoma↗
PubMed1995

Inhibition of progression of aberrant crypt foci and colon tumor development by vitamin E and beta-carotene in rats on a high-risk diet.

In this study we evaluated the effect of dietary administration of a high-fat, low-fiber diet (HRD) supplemented with Vitamin E, beta-carotene or folic acid and wheat bran on the growth of pre-existing aberrant crypt foci (ACF) that had been induced in Fischer-344 rats exposed to azoxymethane (AOM) and a HRD for 10 weeks. The rats (25 rats/dietary group) were fed a HRD for 2 weeks and were then given 2 subcutaneous injections of AOM (15 mg/kg body weight) while the rats continued on the HRD. After 6 weeks, rats were either maintained on the HRD (control) or crossed over to a HRD containing non-toxic levels of either Vitamin E, beta-carotene, folic acid or wheat bran. At 10, 14 and 18 weeks after the initiation of the experiment, 5 rats from each group were killed and the number of aberrant crypt foci (ACF) with different multiplicities were compared between groups. The dietary intervention was continued for 30 weeks to determine whether the inhibitory effect on the growth of ACF influenced the subsequent development of colonic tumors. The results revealed that vitamin E and beta-carotene caused a significant decrease in the number of ACF of different multiplicities when compared to the effect of the HRD alone. The decrease in the number of ACF due to folic acid and wheat bran appeared to be much smaller and in most cases was not significant. However, there was also a significant decrease in the incidence of colonic tumors and tumor multiplicity in both the vitamin E and beta-carotene groups that was not seen in the control group. The reports clearly demonstrates the ability of vitamin E and beta-carotene to inhibit the growth of colonic ACF, even in the presence of the strong promoting effect of high levels of dietary fat, using a post-initiation experimental design.

Animals↗
PubMed1995

Effect of beta-carotene and wheat bran fiber on colonic aberrant crypt and tumor formation in rats exposed to azoxymethane and high dietary fat.

The typical high-fat, low-fiber American diet promotes colon cancer. An alternative to radical changes in dietary habits is to reinforce the diet with cancer protectors. Experiments to evaluate the effects of beta-carotene in the presence of high fat and low and high dietary levels of wheat bran fiber were designed using the Fischer-344 rat colon cancer model. Rats (20/group), were given either high fat (20% w/w), low wheat bran, fiber (1% w/w) diets, or high fat (20% w/w) high wheat bran fiber (8% w/w) diets, with different levels of beta-carotene. After 2 weeks of adaptation, half were given two weekly s.c. injections of azoxymethane (AOM, 15 mg/kg body wt); and half two weekly s.c. injections of saline. Six weeks later, five rats from each dietary group were killed to evaluate the comparative effect of different dietary regimens on the induction of colon aberrant crypt foci (ACF). The remaining rats were maintained on their respective diets for an additional 20 weeks to examine the effect on colon tumor incidence. The total number of ACF/rat in the low-fiber groups declined from 44.0 +/- 4.18 to 12.8 +/- 1.95 in response to increasing amounts of beta-carotene from 1 to 20 mg/kg diet. A similar progressive reduction in total ACF/rat was also seen in the high-fiber groups (20.8 +/- 2.92 to 9.2 +/- 0.58). ACF did not develop in the saline-exposed groups. Similarly colon tumor incidence declined from 73% to 20% in high-fiber groups and from 27% to 13% in low-fiber groups in response to increasing amounts beta-carotene from 1 to 20 mg/kg diet. The results showed that beta-carotene and wheat bran, individually and when combined, protected the colon in rats consuming high-fat, western-style diets from ACF and benign or malignant tumor formation.

Animals↗
PubMed1994

Sequential analysis of K-ras mutations in aberrant crypt foci and colonic tumors induced by azoxymethane in Fischer-344 rats on high-risk diet.

Forty Fischer-344 male rats were given a high-risk diet (HRD) that was high in fat, low in fiber and low in calcium. After 4 weeks, the rats were given two weekly s.c. injections of azoxymethane (AOM, 15 mg/kg body wt), and remained on the same diet till death. Eight rats were killed at 12 weeks and again at 20 weeks in order to microdissect aberrant crypt foci (ACF) containing four or more crypts/focus from their colons. The remaining 24 rats were killed at 30 weeks to harvest colonic tumors. The polymerase chain reaction (PCR) was used to amplify specific DNA segments in the K-ras gene from ACF and colonic tumors. The PCR-amplified DNAs were sequenced to identify the point mutations in codons 12 and 13. All the mutations detected in the ACF and colonic tumors were G to A transitions in the second position of codon 12. These mutations were present in the ACF of 2/8 (25%) and 3/8 (37%) rats at 12 and 20 weeks respectively. The mutations were present in colonic tumors of 7/24 (29%) rats. These results provide important evidence for the significance of K-ras mutations in ACF (> 4 crypts/focus) as early markers of malignant potential in the colons of F344 rats exposed to AOM while receiving a high-risk western style diet.

Adenocarcinoma↗
PubMed1993

Potential synergism between wheat bran and psyllium: enhanced inhibition of colon cancer.

In this study we compared the influence of high fat (20% w/w) diets that combine low levels of calcium (0.18% w/w) and low (1% w/w), medium (4% w/w) and high (8% w/w) levels of dietary fiber from wheat bran (WB), with high (8% w/w) levels of dietary fiber from psyllium (PS) alone or in various combinations with WB, on the induction of colon tumors in Fischer-344 rats following exposure to azoxymethane (AOM). The rats were fed the experimental diets for 2 weeks, and then were given two s.c. injections of AOM (15 mg/kg body wt./week). Twenty-three weeks following the first injection of AOM, the incidence of colon tumors in the different dietary groups (12 rats/group) was compared. The results clearly showed that by increasing the dietary fiber concentration of WB from 1 to 8% significantly reduced the number of colon tumors/group. When the influence of 8% dietary fiber from WB on the development of colon tumors was compared with that of PS (WB:PS = 0:100), no significant difference was observed. However, combinations of WB and PS showed a greater protective effect than either WB or PS alone, at comparable levels of dietary fiber. The 50:50 combination of WB and PS showed maximum protection, while 25:75 and 75:25 combinations both produced intermediate effects. None of the diets showed any significant effect on the normal growth of rats. The results indicate that WB and PS fiber alone, and to a greater degree in combination, can offer protection against colon cancer promoted by high fat, low calcium diets. Diets that include wheat bran in combination with psyllium could be an effective means of reducing colon cancer risk in human populations addicted to high risk western diets.

Animals↗
PubMed1992

Fecal bile acid excretion and composition in response to changes in dietary wheat bran, fat and calcium in the rat.

The effect and possible interactive influence of different dietary amounts of wheat bran, fat and calcium on the fecal excretion, concentration and composition of bile acids was studied in Fischer-344 rats. The fecal bile acids were analyzed using gas-liquid chromatography. Dietary wheat bran increased both total bile acid excretion and fecal weight without changes in fecal bile acid concentration. The proportion of fecal hyodeoxycholic acid decreased with increasing dietary fiber, whereas that of lithocholic and deoxycholic acids increased significantly with fiber intake. The percent content of fecal chenodeoxycholic acid did not change. Increasing dietary fat led to an increase in bile acid excretion without changes in either fecal weight or bile acid concentration. In contrast, the level of dietary calcium did not affect the total excretion of bile acids. However, since calcium increased the fecal weight, it consequently diluted bile acids and decreased their fecal concentration. Dietary fat and calcium had no influence on fecal bile acid composition. There were no interactive effects of wheat bran, fat and calcium on fecal bile acids. The finding in this study that dietary fiber, fat and calcium induce significant changes in fecal bile acids may be of relevance to the potential of bile acids to promote carcinogenesis.

Animals↗
PubMed1992

The effect of high-risk and low-risk diets on aberrant crypt and colonic tumor formation in Fischer-344 rats.

Eighty Fischer-344 male rats were divided into eight groups. Half the groups were given a high-risk diet (HRD) that was high in fat, low in fiber and low in calcium, while the remaining groups were given a low-risk diet (LRD) that was low in fat, high in fiber and high in calcium. After 4 weeks, four groups were then given two weekly s.c. injections of azoxymethane (AOM) (15 mg/kg body wt), and four groups were given saline injections. At the end of 6 weeks the rats were either continued on HRD, LRD, or crossed over from HRD to LRD, or LRD to HRD for an additional 6 weeks. The rats were then killed and the influence of different dietary regimens on the induction of foci of aberrant crypts (AC) in their colons was compared. The number of foci of AC was highest in the AOM-injected HRD/HRD dietary group (8.83 +/- 0.98), followed by the AOM-injected LRD/HRD group (5.37 +/- 0.75), the HRD/LRD group (3.32 +/- 0.36) and the LRD/LRD group (2.93 +/- 0.32). Except for the HRD/HRD control group, which developed a small but significant number of foci of AC (1.24 +/- 0.19), the other saline-injected dietary groups showed no significant numbers of foci of AC. Two groups of rats on HRD/HRD (AOM and saline-injected), and two groups of rats on HRD/LRD (AOM and saline-injected) were continued on HRD and LRD respectively for an additional 14 weeks. A continuous HRD in AOM-injected rats induced colon tumors in 92% of rats, while the crossover from HRD to LRD suppressed the tumor incidence to 33%. Neither diet induced any tumors in saline-injected rats. The results clearly show that the influence of HRD and LRD on the induction of foci of AC and their distribution in the colon, correlated with the induction and distribution of tumors in the colon.

Animals↗
PubMed1992

Magnesium and calcium absorption in Fischer-344 rats influenced by changes in dietary fibre (wheat bran), fat and calcium.

Magnesium and calcium absorption were affected by changes in dietary wheat bran fibre and calcium, but not fat, in Fischer-344 rats when studied in a full factorial study which was a portion of a larger study of diet and colon carcinogenesis. For four weeks, nine-week-old rats were fed experimental purified diets to which had been added: wheat bran 0, 2.5 10, or 20%; fat 1, 5 or 10%; and calcium 0.18, 0.52, or 1.04% of diet weight. From day 26 to 29 all faeces were collected in metabolic cages, and food consumption noted. Dietary magnesium intake and net magnesium absorption increased in direct relation to the quantity of wheat bran in the diet. Calcium supplementation inhibited magnesium absorption on fibre-free diet, but had little effect on magnesium absorption when fibre was present. Fat had no measurable effect on magnesium absorption. A low dietary fibre content enhanced Ca absorption compared to that on a fibre-free diet. However, further increases in fibre content slightly inhibited calcium absorption. We conclude that the magnesium content of dietary wheat bran fibre is available for absorption to rats. Calcium supplementation inhibits magnesium absorption in a fibre-free diet, but presence of dietary fibre protects magnesium absorption from the calcium inhibition observed on a fibre-free diet. Absorption of calcium is increased by including some fibre in the diet. However, calcium absorption may be diminished slightly by increasing wheat bran content of the diet to a high level, probably through calcium binding and excretion with undigested fibre.

Animals↗
PubMed1991

Serum gastrin increases with increasing dietary calcium but not with increasing dietary fat or fiber in Fischer-344 rats.

We studied the effects of dietary calcium, fat and fiber on serum gastrin in Fischer-344 rats in a full factorial experiment as part of a larger study of diet and colon cancer risk factors. Nine- to 10-wk-old male rats were fed standard or experimental diets for 4 wk. Wheat bran was the sole source of fiber. Wheat bran levels were 0, 2.5, 10 and 20%; fat levels were 1, 5 and 10%; calcium levels were 0.18, 0.52 and 1.04% of diet weight. On d 29 serum was collected and stored at -80 degrees C until analyzed. There was a significant (P less than 0.0001) dose-dependent increase in serum gastrin from 102 to 173 ng/L, with increasing calcium. No other significant changes in serum gastrin were noted with the dietary changes. A long-term change in the level of serum gastrin, caused by dietary modification, will influence the trophic effect that gastrin has on colonic mucosa as well as on colon carcinomas. We speculate that calcium supplementation, although slowing colonic proliferation, might have an undesirable effect on the growth of early undetected colonic tumors.

Adenocarcinoma↗
PubMed1989

Influence of microenvironmental pH on adriamycin resistance.

Resistance to Adriamycin (ADR) is frequently dependent upon enhanced efflux associated with the expression of the MDR1-encoded P membrane glycoprotein. Since enhanced expression of the MDR1 gene in ADR-resistant cells may be the result of spontaneous genetic mutation or amplification, it is presumed to be relatively stable and unalterable. Yet, reducing ADR efflux could increase sensitivity, and has been attempted using calcium channel blockers and other drugs. However, since the tumor cell microenvironment varies with respect to pH because of differences in vascularization, oxygenation, and metabolite clearance, the possibility exists that these factors could influence drug transport and the critical biochemical pathways which determine cytotoxicity, even in resistant cells. Using flow cytometric analysis of ADR fluorescence, the influx and efflux of 10 microM ADR dissolved in MES buffer (pH 6.5) and 4-(2-hydroxyethylene)-1-piperazineethanesulfonic acid buffer (pH 7.5 and 8.5) was measured in sensitive P388 and resistant P388/R84 cells in vitro. Substantially enhanced uptake of ADR was detected at alkaline pH in both cell populations, while the proportion of ADR-positive cells and the level of ADR uptake was decreased at lower pH. Acidification reduced ADR efflux, whereas alkalinization increased efflux when the uptake pH was 6.5 or 7.5. At uptake pH 8.5, the pH of the external buffer had little effect, even in resistant cells. In resistant cells in an alkaline microenvironment, ADR transport and retention were superior to that observed in sensitive cells in an acidic microenvironment. No differences were observed in ADR transport when the transmembrane pH gradient was equilibrated. These observations are especially relevant to the effect of ADR on tumor cell subpopulations that are acidic, and in which drug diffusion is inefficient. Efforts to alkalinize tumor cells prior to ADR therapy might reduce ADR resistance, even of genetic origin.

Animals↗