![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
* Faculty of Nutrition,
Department of Veterinary Anatomy and Public Health and
** Department of Statistics, Texas A&M University, College Station, TX 77843
3To whom correspondence should be addressed. E-mail: jlupton{at}tamu.edu.
 |
ABSTRACT |
|---|
 TOP ABSTRACT MATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
KEY WORDS: • apoptosis • reactive oxygen species • antioxidant enzymes • oxidative damage
Colorectal cancer is anticipated to be the third most frequently diagnosed cancer in the United States this year, and it is predicted that almost half of individuals diagnosed will die from the disease within 5 y (1). Yet many of these cases could be prevented by appropriate diet and lifestyle modifications. Dietary fat and fiber are 2 of the most widely investigated dietary components with respect to colon cancer prevention (2,3). There is substantial evidence that diets rich in (n-3) PUFA, such as those found in fish oil (eicosapentaenoic acid and docosahexaenoic acid), protect against colon carcinogenesis (4,5), whereas diets rich in (n-6) PUFA, such as those found in corn oil, appear to promote cancer development in the colon. However, the chemopreventive abilities of fiber, fermentable and nonfermentable, have been the subject of much debate and have shown varied results in intervention trials and epidemiologic studies (6,7). Poorly fermented fibers, such as cellulose, have been considered protective in their ability to dilute putative carcinogens that may be present in the fecal stream. Yet, the products of highly fermentable fibers (i.e., butyrate) have been shown, at least in vitro, to possess chemopreventive qualities (8). We propose that the inconsistent effect seen with fiber may be attributable to the composition of the fat in the diet (9). Specifically, we have shown that the fermentable fiber, pectin, in combination with fish oil, has a protective effect in multiple stages of colon cancer (4,10).
The protective effect of this diet was shown to be primarily through enhancement of apoptosis, a form of programmed cell death (4,11). However, the mechanism by which the fish oil and pectin diet induces apoptosis has not been clearly elucidated. Recent evidence suggests that reactive oxygen species (ROS)4 comprise an important mediator of apoptosis [reviewed in (12)]. Considering the degree of unsaturation of (n-3) PUFA in combination with the rapid colonocyte oxidation of butyrate, this protective diet may alter cellular ROS in a manner sufficient to induce apoptosis in the colonocyte. Yet ROS can also damage and potentially mutate DNA (13); therefore, cells employ several defenses against ROS including antioxidant enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT). Although these enzymes are key players in preventing cellular damage caused by endogenous ROS (14,15), overexpression or addition of these enzymes to tissue systems has been shown to enhance tumorigenesis and block the action of several chemotherapeutic drugs by suppressing ROS-induced apoptosis (16,17). The ability of diet to modulate antioxidant enzyme expression and activity has been documented (18,19). However the ability of diet to simultaneously influence additional redox factors, such as ROS generation, in the colon has not been characterized. This investigation evaluates the ability of dietary lipid and fiber to alter the oxidative status of rat colonocytes, via ROS generation and modulation of antioxidant enzyme activity, thus creating an environment permissive for apoptosis.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
-tocopherol, 
-tocopherol, and tert-butyl hydroquinone equal to the levels found in corn oil. Food and water were freely available. To minimize fatty acid oxidation, diets were stored at –80°C and fresh food was provided every 24 h. Food intake and body weights were measured weekly. Tissue collection and cell isolation. After the rats were killed by CO2 asphyxiation and cervical dislocation, the colon was removed and flushed with warm Ca2+- and Mg2+-free PBS (GibcoBRL). For each rat, the last half of the colon was taken as the distal colon. The last centimeter of the distal colon was taken for histology and fixed in 4% paraformaldehyde for 4 h, followed by washing in 50 and 70% ethanol. The remaining colon segment was cut longitudinally to expose the lumen and placed in warm Ca2+- and Mg2+-free HBSS, 30 mmol/L EDTA, 5 mmol/L dithiothreitol (DTT), 0.1% fatty acid–free bovine serum albumin (BSA; wt/v), 1 mmol/L glutamine, and 1 mmol/L butyrate (pH 7.4). After a 15-min shaking incubation, the mucosal side was gently scraped with a rubber policeman. This procedure is designed to remove intact crypts and surface cells leaving behind the lamina propria (23). Removal of crypts and surface cells was confirmed by histological examination of the remaining intestinal tissue after the scraping procedure. The isolated crypts were then centrifuged at 100 x g and washed twice in warm HBSS containing Ca2+, Mg2+, 0.1% BSA (wt/v), 1 mmol/L glutamine, and 1 mmol/L butyrate. An aliquot of cells from the distal colon was taken for antioxidant enzyme analysis and fragment length analysis using repair enzymes (FLARE).
Measurement of apoptosis using the dUTP nick end-labeling (TUNEL) assay. Paraffin sections of the 4% paraformaldehyde fixed tissues were utilized for in situ measurement of apoptosis using ApopTag kits (Intergen) as previously described (24). This technology is based on the terminal deoxynucleotidyl transferase-mediated TUNEL technique. Intestinal crypts were scored according to positive staining by DAB and morphological criteria previously described by Kerr et al. (25). The apoptotic index for each crypt was determined by dividing the number of apoptotic cells in a crypt column by the crypt column height (number of cells). The mean apoptotic index of 25 crypts within a tissue was used as the apoptotic index for that tissue.
Measurement of antioxidant enzyme activity. Activities of CAT, GPx, and SOD in isolated colonocytes were measured spectrophotometrically using commercial assay kits (Calbiochem). Cell lysates were prepared by homogenization of cells in 50 mmol/L potassium phosphate buffer [250 mmol/L sucrose, 1 mmol/L EDTA, 1 mmol/L DTT, 0.1% Triton X-100 (v/v)] followed by centrifugation for 3 min at 10,000 x g. The supernatant was used for enzyme assays following protocols provided in each kit. Briefly, SOD activity was determined by measuring the rate of generation of a chromophore at 525 nm. CAT activity was determined by measuring the absorbance of quinoneimene dye at 520 nm. GPx was determined indirectly by oxidation of NADPH to NADP+ measured at 340 nm using H2O2 as the preferred substrate. Sodium azide (NaN3) was used to inhibit catalase competition for H2O2. Samples were analyzed in triplicate in 96-well microplates with standards provided in kits or purchased separately from Calbiochem. Microplates were read on a Spectra Max 250 microtiter plate reader using SoftMax Pro v.1.2 software (Molecular Devices). Activity was normalized to protein concentration as determined by Coomassie Blue assay (Pierce).
Detection of reactive oxygen species. Samples of isolated colonocytes (maintained at 37°C) were prepared in duplicate and incubated for 15 min with chloromethyl-2',7'-dichlorodihydrofluorescein diacetate (Molecular Probes), a fluorescence probe sensitive to such cellular oxidants as hydrogen peroxide (H2O2), hydroxyl radicals (OH•), and peroxyl radicals (OOH•). This probe passively diffuses into cells and upon oxidation by ROS forms a fluorescent adduct that remains trapped in the cell. Fluorescence was monitored on a Meridian Ultima confocal microscope (Meridian Instruments) with a 530-nm barrier filter and laser excitation at 488 nm, as previously described (26). Fluorescence intensity was used as an indirect measure of prevalence of ROS. Data for each sample were collected from 15 fields/treatment for each rat. Viability of the cells used for analysis was determined after each treatment by staining with ethidium homodimer-1 (Molecular Probes). Mean viability was 81 ± 4.5% (n = 60).
Measurement of oxidative DNA damage using the FLARE assay. This assay is a modification of single-cell gel electrophoresis, which uses Escherichia Coli formamidopyrimidine-DNA glycosylase (fpg) to introduce DNA strand breaks specifically at 8-hydroxydeoxyguanosine (8-OHdG) adducts (27,28), a prevalent and potentially mutagenic oxidative DNA adduct (29). This process measures the levels of 8-OHdG on a single-cell level in intact nuclei (30). 8-OHdG adducts were quantified using a comet assay kit (Trevigen). After isolation, crypts were kept at 4°C for the entire procedure to minimize DNA repair and were dispersed into single cells by repeated aspiration through a 27-gauge needle and plated with agarose in duplicate on comet slides. Slides were exposed to lysis buffer (1% sodium lauryl sarcosinate (v/v), 2.5 mol/L NaCl, 100 mmol/L EDTA, 1% Triton X-100 (v/v), 10 mmol/L Tris base, pH 10) to remove the outer cell membrane leaving only intact nuclei. Slides were then immersed in 1X FLARE buffer (10 mmol/L HEPES-KOH, 100 mmol/L KCl, pH 7.4) followed by the addition of the fpg enzyme (Trevigen) diluted 1:50 with reaction buffer (25X FLARE buffer, 100X BSA). Control slides received only reaction buffer without the fpg enzyme. After treatment with alkali solution (1 mmol/L EDTA, pH 12.5) to denature DNA strands, slides were exposed to electrophoresis (1 V/cm, 20 min) and immersed briefly in 70% ethanol (v/v). Nuclei were viewed by epifluorescence microscopy using SYBR green staining (Molecular Probes). Quantitation of the relative tail moment [tail moment/(tail moment + head moment)] (31) was measured using Metamorph software (Nikon, Garden City, NY). One hundred randomly selected cells were analyzed per treatment group for each rat.
Statistical analysis. Analyses of ROS, antioxidant enzyme activity, and oxidative DNA damage were performed by mixed model ANOVA using SAS 8.0 (SAS Institute). Apoptotic indices were analyzed with Poisson regression using Proc Genmod. The relation between antioxidant enzymes and ROS was determined by regression analysis in SPSS. The relation between ROS and oxidative DNA damage was examined by regression analysis using the generalized estimation equation approach with an identity link (32). The covariates considered in the model included the combinations of oil and fiber as well as oxidative DNA damage nested within these treatment combinations. To explain the relation of ROS and apoptotic index, the linear relation between the log-transformed apoptotic index and ROS levels was modeled. To reduce the influence from potential outlying observations, a robust regression using Huber’s weight function was performed (33). Differences were considered significant at P < 0.05.
|
RESULTS |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
Apoptosis. The combination of dietary fish oil and pectin yielded a greater apoptotic index compared with the other experimental diets (P < 0.008) (Fig. 1). These results are in agreement with previous findings from our laboratory in which a diet of fish oil and pectin enhanced apoptosis during experimentally induced colon cancer (4,11).
|
|
|
|
|
|
DISCUSSION |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
Important determinants of cellular antioxidant capacity are the enzymes SOD, CAT, and GPx, which are responsible for the elimination of ROS. Because these enzymes act sequentially to remove ROS, the balance of the activity of these enzymes may be as critical in the defense against ROS as the activity of the enzymes alone (37). Data from this experiment showed that the activity of these enzymes, especially CAT, is influenced by dietary fiber. Specifically, antioxidant enzyme activity in colonocytes from rats fed a pectin diet was less than that observed in those fed the cellulose diets. Furthermore, the combination of fish oil with pectin in the diet resulted in even lower activity for CAT and SOD. Interestingly, the difference in CAT activity was greater than the change seen with SOD, suggesting a possible enzyme activity imbalance. SOD converts superoxide (O2•–) to H2O2, which is then converted to water and/or O2 by CAT and GPx. Thus, dramatically diminished CAT activity coupled to only a subtle reduction in SOD (an increase in SOD/CAT) may yield a system that can no longer eliminate H2O2 at the rate it is formed. Indeed, dietary pectin and to a lesser extent dietary fish oil elevate the SOD/CAT ratio compared with cellulose and corn oil, respectively. Furthermore, the greatest enhancement in SOD/CAT was seen when fish oil and pectin were combined in the diet. Similar experiments in rat colonocytes found that dietary fish oil reduces antioxidant enzyme activity (18), and a recent dietary intervention trial showed that diets high in fiber and (n-3) PUFA are capable of reducing antioxidant enzyme activity in humans (19). Although the mechanism by which these dietary constituents achieve a reduction in enzyme activity is unclear, these results suggest that dietary lipid and dietary fiber, specifically fish oil and pectin, work coordinately to alter antioxidant enzyme activity and balance in a manner that may create a prooxidant environment in the colonocytes.
ROS measurements further suggest that dietary fish oil may create a more oxidative environment in the colonocytes compared with a corn oil diet. Diets with fish oil as the lipid source enhanced ROS generation in the colonocytes. This is not unexpected considering the high degree of unsaturation found in the long-chain (n-3) PUFA in fish oil. The primary (n-3) fatty acids in fish oil, eicosapentaenoic acid [20:5(n-3)] and docosahexaenoic acid [22:6(n-3)] have up to 3 times as many double bonds per molecule than the (n-6) fatty acids found in corn oil, such as linoleic acid [18:2(n-6)]. This increases the opportunity for oxidant attack and can contribute to the propagation of ROS. Furthermore, we have shown dietary (n-3) PUFA to be readily incorporated into the mitochondrial membrane (38), predisposing the mitochondria to enhanced lipid peroxidation and membrane damage and contributing to the propagation of ROS generated by the mitochondrial electron transport system (36). It is likely that many of the fatty acids consumed in these experimental diets maintained a high degree of unsaturation because the diets were kept at –80°C to prevent fatty acid oxidation. However, these conditions may not be practical outside of the laboratory setting, and less than optimal storage may enhance the degree of fatty acid oxidation before consumption. This may have a substantial impact on the physiologic effects of dietary lipid and definitely warrants further attention.
Although dietary fish oil and pectin alter the antioxidant capacity and ROS generation of colonocytes to favor a prooxidant environment, the outcome of these diet-induced cellular modifications is critical. An oxidative environment may favor apoptosis, or alternatively, may increase the potentially mutagenic event of damaging DNA. Therefore, it was important to determine apoptosis and oxidative DNA damage in the same rats in which the diet-induced changes in ROS and antioxidant enzyme activity were measured. In the fish oil/pectin diet, apoptosis was enhanced and the oxidative environment created by this diet was associated with a decline in oxidative DNA damage. Alternatively, in the corn oil/cellulose diet, apoptosis was less than in the fish oil/pectin diet and as ROS increased, oxidative damage did not decrease as in the fish oil/pectin diet. Thus, the suppression of oxidant protection systems and enhancement of ROS generation by dietary fish oil and pectin appears to protect the colon against oxidative DNA damage by promoting ROS-mediated apoptosis. However, continued investigation is required to further elucidate the multifaceted relation between cellular redox status and apoptosis. Additional investigation is also warranted to determine whether these dietary modifications of the redox environment and the resulting increase in apoptosis continue during the initiation and progression stages of colon carcinogenesis. Although we showed previously that apoptosis was enhanced by dietary fish oil and pectin during the initiation (11) and progression (4) of colon carcinogenesis, whether the mechanism is associated with alteration of the redox environment remains to be determined.
In summary, dietary fish oil and pectin work coordinately to enhance colonocyte apoptosis by modulation of the cellular redox environment. In this ex vivo model, we showed that dietary fish oil enhanced ROS in colonocytes, whereas the effects of dietary pectin were more clearly seen in the attenuation of antioxidant enzyme activity. Therefore, it is not surprising that the combination of these dietary constituents worked in concert to create an environment permissive for apoptosis, thereby protecting cells from severe and possibly mutagenic DNA damage. This study reinforces the importance of diet for the prevention of cancer and strengthens the growing realization that the effects of individual diet components may not be as important as the combination of foods consumed in the diet. Further investigations should evaluate the influence of other components of the diet matrix as well as the effect of alterations to diet components (e.g., fatty acid oxidation before consumption).
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
2 Supported by National Institutes of Health grants CA61750, CA82907, CA59034, CA57030, DK53055, NSBRI NASA NCC 9-58, NIEHS-P30-ES09106, and a Grant-In-Aid of Research from Sigma Xi.
4 Abbreviations used: BSA, bovine serum albumin; CAT, catalase; DTT, dithiothreitol; FLARE, fragment length analysis using repair enzymes; fpg, formamidopyrimidine-DNA glycosylase; GPx, glutathione peroxidase; 8-OHdG, 8-hydroxydeoxyguanosine; ROS, reactive oxygen species; SOD, superoxide dismutase; TUNEL, dUTP nick end-labeling.
Manuscript received 22 July 2004. Initial review completed 12 August 2004. Revision accepted 8 September 2004.
|
LITERATURE CITED |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
1. Jemal, A., Tiwari, R. C., Murray, T., Ghafoor, A., Samuels, A., Ward, E., Feuer, E. J. & Thun, M. J. (2004) Cancer statistics, 2004. CA Cancer J. Clin. 54:8-29.
2. Reddy, B. (2000) Novel approaches to the prevention of colon cancer by nutritional manipulation and chemoprevention. Cancer Epidemiol. Biomark. Prev. 9:239-247.
3. Lipkin, M., Reddy, B., Newmark, H. & Lamprecht, S. A. (1999) Dietary factors in human colorectal cancer. Annu. Rev. Nutr. 19:545-586.[Medline]
4. Chang, W.-C.L., Chapkin, R. S. & Lupton, J. R. (1998) Fish oil blocks azoxymethane-induced rat colon tumorigenesis by increasing cell differentiation and apoptosis rather than decreasing cell proliferation. J. Nutr. 128:491-497.
5. Latham, P., Lund, E. K. & Johnson, I. T. (1999) Dietary n-3 PUFA increases the apoptotic response to 1,2-dimethylhydrazine, reduces mitosis and suppresses the induction of carcinogenesis in the rat colon. Carcinogenesis 20:645-650.
6. Alberts, D. S., Martinez, M. E., Roe, D. J., Guillen-Rodriguez, J. M., Marshall, J. R., Van Leeuwen, B., Reid, M. E., Ritenbaugh, C., Vargas, P. A., Bhattacharyya, A., Earnest, D. L. & Sampliner, R. E. (2000) Lack of effect of a high-fiber cereal supplement on the recurrence of colorectal adenomas. N. Engl. J. Med. 342:1156-1162.
7. Peters, U., Sinha, R., Chatterjee, N., Subar, A. F., Ziegler, R. G., Kulldorff, M., Bresalier, R., Weissfeld, J. L., Flood, A. & Schatzkin, A. (2003) Dietary fibre and colorectal adenoma in a colorectal cancer early detection programme. Lancet 361:1491-1495.[Medline]
8. Lupton, J. R. (2004) Microbial degradation products influence colon cancer risk: the butyrate controversy. J. Nutr. 134:479-482.
9. Lupton, J. R. (2000) Is fiber protective against colon cancer? Where the research is leading us?. Nutrition 16:558-561.[Medline]
10. Hong, M. Y., Chang, W.-C.L., Chapkin, R. S. & Lupton, J. R. (1997) Relationship among colonocyte proliferation, differentiation and apoptosis as a function of diet and carcinogen. Nutr. Cancer 28:20-29.[Medline]
11. Hong, M. Y., Lupton, J. R., Morris, J. S., Wang, N., Carroll, R., Davidson, L., Elder, R. & Chapkin, R. (2000) Dietary fish oil reduces O6-methylguanine DNA adduct levels in rat colon in part by increasing apoptosis during tumor initiation. Cancer Epidemiol. Biomark. Prev. 9:819-826.
12. Mates, J. M. & Sanchez-Jimenez, F. M. (1999) Role of reactive oxygen species in apoptosis: implications for cancer therapy. Int. J. Biochem. Cell Biol. 32:157-170.
13. Beckman, K. B. & Ames, B. N. (1997) Oxidative decay of DNA. J. Biol. Chem. 272:19633-19636.
14. Beckman, K. B. & Ames, B. N. (1998) The free radical theory of aging matures. Physiol. Rev. 78:547-581.
15. Mates, J. M., Perez-Gomez, C. & Castro, I.N.D. (1999) Antioxidant enzymes and human diseases. Clin. Biochem. 32:595-603.[Medline]
16. Hirose, K., Longo, D., Oppenheim, J. & Matsushima, K. (1993) Overexpression of mitochondrial manganese superoxide dismutase promotes the survival of tumor cells exposed to interleukin-1, tumor necrosis factor, selected anticancer drugs, and ionizing radiation. FASEB J. 7:361-368.[Abstract]
17. Gorman, A., McGowan, A. & Cotter, T. G. (1997) Role of peroxide and superoxide anion during tumour cell apoptosis. FEBS Lett. 404:27-33.[Medline]
18. Nieto, N., Fernandez, M., Torres, M., Rios, A., Suarez, M. & Gil, A. (1998) Dietary monounsaturated n-3 and n-6 long chain polyunsaturated fatty acids affect cellular antioxidant defense system in rats with experimental ulcerative colitis induced by trinitrobenzene sulfonic acid. Dig. Dis. Sci. 43:2676-2687.[Medline]
19. Bruce, B., Spiller, G., Klevay, L. & Gallagher, S. (2000) A diet high in whole and unrefined foods favorably alters lipids, antioxidant defenses and colon function. J. Am. Coll. Nutr. 19:61-67.
20. Maciorowski, K. G., Turner, N. D., Lupton, J. R., Chapkin, R. S., Shermer, C. L., Ha, S. D. & Ricke, S. C. (1997) Diet and carcinogen alter the fecal microbial populations of rats. J. Nutr. 127:449-457.
21. Pickering, J. S., Lupton, J. R. & Chapkin, R. S. (1995) Dietary fat, fiber, and carcinogen alter fecal diacylglycerol composition and mass. Cancer Res. 55:2293-2298.
22. Zoran, D. L., Turner, N. D., Taddeo, S. S., Chapkin, R. S. & Lupton, J. R. (1997) Wheat bran reduces tumor incidence in a rat model of colon cancer independent of effects on distal luminal butyrate concentrations. J. Nutr. 127:2217-2225.
23. Zhang, J., Wu, G., Chapkin, R. S. & Lupton, J. R. (1998) Energy metabolism of rat colonocytes changes during the tumorigenic process and is dependent on diet and carcinogen. J. Nutr. 128:1262-1269.
24. Chang, W. C., Chapkin, R. S. & Lupton, J. R. (1997) Predictive value of proliferation, differentiation, and apoptosis as intermediate markers for colon tumorigenesis. Carcinogenesis 18:721-730.
25. Kerr, J., Gobe, G., Winterford, C. & Harmon, B. (1995) Anatomical methods in cell death. Methods Cell Biol. 46:1-27.[Medline]
26. Barhoumi, R. & Burghardt, R. C. (1996) Kinetic analysis of the chronology of patulin- and gossypol-induced cytotoxicity in vitro. Fundam. Appl. Toxic. 30:290-297.
27. Collins, A. R., Ma, A. G. & Duthie, S. J. (1995) The kinetics of repair of oxidative DNA damage (strand breaks and oxidized pyrimidine dimers) in human cells. Mutat. Res. 336:69-77.[Medline]
28. Singh, N. P., McCoy, M. T., Tice, R. R. & Schneider, E. L. (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp. Cell Res. 175:184-191.[Medline]
29. Feig, D. I., Reid, T. M. & Loeb, L. A. (1994) Reactive oxygen species in tumorigenesis. Cancer Res. 54:1890S-1894S.
30. Bancroft, L. K., Lupton, J. R., Davidson, L. A., Taddeo, S. S., Murphy, M. E., Carroll, R. J. & Chapkin, R. S. (2003) Dietary fish oil reduces oxidative DNA damage in rat colonocytes. Free Radic. Biol. Med. 35:149-159.[Medline]
31. Riso, P., Santangelo, A. & Porrini, M. (1999) The comet assay for the evaluation of cell resistance to oxidative stress. Nutr. Res. 19:325-333.
32. Diggle, P., Liang, K. & Zeger, S. (1994) Analysis of Longitudinal Data 1994 Oxford University Press Oxford, UK.
33. Heiberger, R. & Becker, R. (1992) Design of an S function for robust regression using iteratively reweighted least squares. J. Comput. Graph. Stat. 1:181-196.
34. Renehan, A., Booth, C. & Potten, C. (2001) What is apoptosis, and why is it important?. Br. Med. J. 322:1536-1538.
35. Wiseman, H. & Halliwell, B. (1996) Damage to DNA by reactive oxygen and nitrogen species: role in inflammatory disease and progression to cancer. Biochem. J. 313:17-29.
36. Hong, M. Y., Chapkin, R. S., Barhoumi, R., Burghardt, R. C., Turner, N. D., Henderson, C. E., Sanders, L. M., Fan, Y.-Y., Davidson, L. A., Murphy, M. E., Spinka, C. M., Carroll, R. J. & Lupton, J. R. (2002) Fish oil increases mitochondrial phospholipid unsaturation, upregulating reactive oxygen species and apoptosis in rat colonocytes. Carcinogenesis 23:1919-1926.
37. Li, S., Yan, T., Yang, J.-Q., Oberley, T. D. & Oberley, L. W. (2000) The role of cellular glutathione peroxidase redox regulation in the suppression of tumor cell growth by manganese superoxide dismutase. Cancer Res. 60:3927-3939.
38. Chapkin, R., Hong, M., Fan, Y., Davidson, L., Sanders, L., Henderson, C., Barhoumi, R., Burghardt, R., Turner, N. & Lupton, J. (2002) Dietary n-3 PUFA alter colonocyte mitochondrial membrane composition and function. Lipids 37:193-199.[Medline]
This article has been cited by other articles:
|
|
 |
|
  J. Vanamala, A. Glagolenko, P. Yang, R.J. Carroll, M.E. Murphy, R.A. Newman, J.R. Ford, L.A. Braby, R.S. Chapkin, N.D. Turner, et al. Dietary fish oil and pectin enhance colonocyte apoptosis in part through suppression of PPAR{delta}/PGE2 and elevation of PGE3 Carcinogenesis, April 1, 2008; 29(4): 790 - 796. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. L. Wakefield, M. Lane, S. J. Schulz, M. L. Hebart, J. G. Thompson, and M. Mitchell Maternal supply of omega-3 polyunsaturated fatty acids alter mechanisms involved in oocyte and early embryo development in the mouse Am J Physiol Endocrinol Metab, February 1, 2008; 294(2): E425 - E434. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. S. N. Kolar, R. Barhoumi, E. S. Callaway, Y.-Y. Fan, N. Wang, J. R. Lupton, and R. S. Chapkin Synergy between docosahexaenoic acid and butyrate elicits p53-independent apoptosis via mitochondrial Ca2+ accumulation in colonocytes Am J Physiol Gastrointest Liver Physiol, November 1, 2007; 293(5): G935 - G943. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. S. N. Kolar, R. Barhoumi, J. R. Lupton, and R. S. Chapkin Docosahexaenoic Acid and Butyrate Synergistically Induce Colonocyte Apoptosis by Enhancing Mitochondrial Ca2+ Accumulation Cancer Res., June 1, 2007; 67(11): 5561 - 5568. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Ng, R. Barhoumi, R. B. Tjalkens, Y.-Y. Fan, S. Kolar, N. Wang, J. R. Lupton, and R. S. Chapkin The role of docosahexaenoic acid in mediating mitochondrial membrane lipid oxidation and apoptosis in colonocytes Carcinogenesis, November 1, 2005; 26(11): 1914 - 1921. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
