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1. Rate sedimentation and isopycnic centrifugation were used to analyse the subcellular sites of enzymes in homogenates of goldfish intestinal mucosa. 2. The results allowed the following allocations to be made: carnitine acetyl transferase-mitochondrial and peroxisomal, xanthine dehydrogenase and NAD: alpha-glycerophosphate dehydrogenase soluble phase, NADP: isocitrate dehydrogenase soluble phase and mitochondrial, and 2-naphthyl laurate hydrolase microsomal and/or brush border. 3. Histochemistry confirmed the use of alkaline phosphatase and 1-naphthyl acetate esterase as brush border and microsome markers respectively. 4. Urate oxidase, allantoinase, allantoicase, xanthine oxidase and glycollate/lactate oxidase, activities were undetectable, and 1-naphthyl palmitate hydrolase was present only as a contaminant from pancreas.

We investigated the effect of high fat and nutrient depleted diets on the formation of colon tumors in female Swiss mice treated with 1,2-dimethylhydrazine (DMH). Mice received the following diets: control diet (laboratory chow, 5.5% fat) or chow with added starch and/or corn oil so as to supply a fat level of 5.5% (diet F6) or 23% (diet F23), while reducing the level of nutrients and dietary fiber per 100 calories to 61% of the level of the control diet. DMH was given as 5 weekly s.c. injections. Diets were given variously from 6 weeks before the first injection until 7 days after the last one (initiation period) or else for the subsequent 22 weeks until sacrifice of the mice (promotion period). A high fat diet (diet F23 vs. F6) tended to increase adenoma formation when fed during the initiation period but this requires further study. Feeding it in the promotion period increased the incidence of adenocarcinomas. A nutrient depleted diet (diet F6 vs. the control diet) caused a decreased incidence of adenocarcinomas.

The effect of diet on the lethality of 1, 2-dimethylhydrazine (DMH) injec tions was investigated using Swiss mice. Mortality was highest when mice were fed a nutrient-dilutealfhigh-fat diet, was intermediate with a nutrientdilutealflow-fat diet and was lowest with laboratory chow. Wheat bran was not protective. Mice developed tolerance to DMH after receiving several injections of progressively increasing dose.

During the refining of carbohydrate foods there is a sharp drop in the concentration of dietary fiber and of various vitamins and minerals. Estimates were made of the effect of refining on the total diet intake of fiber, selenium, folic acid, vitamins E and B6, choline, chromium, magnesium, zinc, manganese, copper, sodium and potassium. The health implications of this are discussed and it is concluded that the losses are likely to be detrimental.

Female Swiss mice were fed on a control diet (laboratory chow, 6% fat) or high fat diets (laboratory chow with added fat, principally corn oil, to produce a fat content of 16% or 23%). Homogenate of colon mucosa was fractionated by isopycnic banding on a linear sucrose gradient and the distribution of 5′-nucleotidase determined. Two main peaks were detected which we interpret as belonging to basolateral plasma membrane (BLPM) and brush borders. A high fat diet did not affect the relative sizes of the peaks or the median density of the BLPM peak but did significantly reduce the density of the brush border peak from 1.208 g/ml to 1.197 g/ml.

Female, Swiss mice were fed semi-synthetic diets for 33 days. The diets were fibre-free (FF) or supplemented with corn bran (CB) 12%, wheat bran (WB) 12%, alfalfa (AL) 12%, pectin (P) 6%, cellulose (CL) 6%, or lignin (LG) 6%. Fibre caused little hyperplasia of the colon mucosa. The number of cells per crypt was increased 9–13% and the crypt column length by 14–19% in the CL, AL and LG groups. CB caused rather less hyperplasia, WB less again and P caused none. The colon mucosal DNA content was approximately 5–10% lower in mice given supplemental fibre. The pH of the contents of the distal colon was apparently unaffected by fibre. Measurement of a non-specific, non-enzymic reducing activity indicated that activity was doubled by AL and LG, lowered 41% by CB but little changed by WB, CL and P. The deoxycholate binding capacity of the colon contents was increased 3–4-fold by LG, whereas the other fibre sources were without appreciable effect. This high binding capacity by LG was also seen in the material used for diet preparation. Analyses of the contents of the caecum and of the remaining colon indicated that as food residue passes from the caecum to the remaining colon little change in binding capacity occurs.

Medical research is here divided into two broad categories. The great majority of research is "complex" and includes all studies of intricate body mechanisms. A minority of research is "simple" and deals mainly with direct relationships between human environmental or life-style factors and disease incidence (or with analogous studies on animals). In addition, it includes studies of simple body mechanisms. Examples are provided to demonstrate that most of our medical knowledge of practical benefit derives from simple research. This is particularly the case in the area of prevention.

1. 1. Rate dependent and isopycnic banding in a zonal rotor were used to analyse the subcellular sites of enzymes in homogenates of guinea-pig small intestinal mucosa. 2. 2. The results demonstrate the following localizations: monoglyceride acyltransferase—microsomal; xanthine oxidase and dehydrogenase—soluble phase, and NADP: isocitrate dehydrogenase—soluble phase and mitochondrial. 3. 3. Alkaline phosphatase is confined to brush borders and is absent from the basolateral plasma membrane. A variable proportion of the activity, up to 40% is on brush borders which during homogenization break up into particles of reduced density and slow sedimentation rate.

Summary The effects of dietary supplements of .a-carotene (20-500 mg per kg diet) on hepatic microsomal drug-metabolizing enzyme activities were studied in mice. Supplementation for 14 days resulted in marked reductions in the concentrations of cytochrome P-4S0 and biphenyl 4-hydroxylase. The antipyrine N-demethylase and p-nitroanisol O-demethylase activities, however, were unchanged. Also apparently unchanged were the hepatic concentrations of microsomal protein, lipid peroxides, and superoxide dismutase. Supplemental .a-carotene was weakly protective against the acute toxic effects of an injection of 1,2-dimethylhydrazine (DMH), as indicated by a lowered mortality. This anti-carcinogenic action of p-carotene, including a protection against Di'\1H-induced colon carcinogenesis, suggests an alteration by this carotenoid in the metabolism of carcinogens by the liver.

The effects were investigated of cyclical rhythm and hormonal contraception on serum fat-mobilizing activity, glycerol, cholesterol and whole blood glucose during 2 menstrual cycles in a group of normally menstruating young women and a second group of young women using hormonal contraception. A control group of normal young men was also investigated. There was no evidence of any change in mean level of any of the parameters measured, among the follicular, ovulatory and luteal phases. No cyclical pattern was discernable in the male subjects. The mean value for serum cholesterol concentration in women using hormonal contraception was higher than the value for the untreated human female group. The overall mean value for serum glycerol concentration in the women was significantly (0.01 > P > 0.001) higher than the mean value obtaining in the men.

The effects of cabbage and vitamin E on colon carcinogenesis were investigated in Swiss mice treated with 1,2-dimethylhydrazine. Throughout the experiment the mice were fed a laboratory chow diet (46 mg vitamin E per kg) or chow containing 13 g cabbage per 100 g or 180 mg vitamin E per kg. Starting after 31 days of diet treatment the mice received 7 weekly s.c. injections of DMH. They were sacrificed 17 weeks after the first dose of DMH. While diet did not significantly alter colon tumor response, some trends were observed. Female mice given cabbage had a higher incidence (percent of mice with a tumor) and multiplicity (tumors per tumor bearing mouse) of colon tumors. Males were little affected by cabbage apart from a lower incidence of adenocarcinomas. Compared with mice fed the control diet those given vitamin E had a higher colon tumor incidence. This effect, which was stronger in females, was due to an increased incidence of adenomas. Vitamin E had little apparent affect on tumor multiplicity apart from a reduction in adenocarcinomas in females and adenomas in males. The data do not support the view that cabbage and vitamin E are protective against colon cancer.

In one experiment Swiss mice were maintained on a 16 or 23% fat diet (laboratory chow with added fat, principally corn oil) or on laboratory chow alone (5.5% fat). In another experiment C57BL/1 mice were given a 23% fat diet (as above) or a low-fat diet (67% laboratory chow, 1.9% corn oil, and 31% starch; 5.5% fat). Colon mucosal samples were analyzed for several enzyme activities. In Swiss mice the analyses revealed the following: 1) Ouabain-insensitive ATPase was unaltered in male mice, but it rose significantly in females fed a high-fat diet (this effect was seen when a resuspended high-speed pellet was analyzed but not seen with the initial homogenate); 2) 5'-nucleotidase activity showed a significant stepwise increase with dietary fat; 3) nonspecific esterase activity tended to rise with a high-fat diet (not significant); 4) beta-glucuronidase levels were not altered by diet fat; and 5) ornithine decarboxylase levels were not altered by diet fat. In C57BL/1 mice analyses were done on ouabain-insensitive ATPase, 5'-nucleotidase, nonspecific esterase, and beta-glucuronidase, but no diet effects were seen. Fecal reductase activity was measured with the use of 2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride hydrate). A high-fat diet did not affect the activity in C57BL/1 mice, but it caused a significant rise in Swiss mice.

Strategies of subcellular fractionation are reviewed from the perspective of their utility in the analysis of peroxisomes. The considerable potential inherant in the method of rate dependent banding in zonal rotors is emphasized. The use of various density gradient solutes is considered.

A total of 89 subjects including 30 breast cancer patients with distal metastases, 29 patients with benign breast disease, and 30 healthy subjects were studied. Serum samples from these subjects were obtained from the National Cancer Institute (NCI) Breast Cancer Serum Bank, Bethesda. Serum concentrations of vitamin A and its transport proteins (prealbumin and retinol-binding protein [RBP]), beta-carotene, vitamin E, and selenium were determined. For each of these parameters the mean for the breast cancer patients was lower than that of the healthy subjects. The differences between healthy subjects and patients with either breast cancer or benign breast disease were, however, statistically significant only in the case of RBP (p less than 0.05). In the case of vitamin A and its transport proteins these differences were reduced by comparing the cancer patients with the benign breast disease patients rather than with the healthy controls. This indicates that the low serum levels for those three parameters may be merely a consequence of disease in general rather than a feature of cancer per se.

The epidemiological evidence associating dietary lipids, refined carbohydrates and coronary heart disease (CHD) is evaluated. Population studies often show a high correlation between dietary lipid and heart disease mortality. It is argued that most of this association is secondary to the high correlation that refined carbohydrates have with dietary lipids, on the one hand, and with CHD on the other. This becomes apparent only when examples are found of a weak or negative correlation between dietary lipids and refined carbohydrates. Evidence from diet and drug intervention studies support the hypothesis that dietary lipid is only of secondary importance in CHD. It is concluded that refined carbohydrates are of primary importance.

Adipose tissues and other tissues of the pig have been examined for the presence of the mitochondrial "uncoupling protein," characteristic of brown adipose tissue, in order to assess whether brown fat is present in this species. Mitochondria were prepared from various tissues and the proteins separated on the basis of molecular weight by sodium dodecyl sulphate – polyacrylamide gel electrophoresis. Immunoblotting procedures were then used to probe for uncoupling protein, employing a rabbit anti-(rat uncoupling protein) serum. Pigs were examined at 4 days, 4 weeks, and 8 weeks of age. No evidence for the presence of uncoupling protein was found at any of these ages. The protein was, however, readily detected in brown adipose tissue from rats, mice, golden hamsters, guinea pigs, Richardson's ground squirrel, and lambs. An additional group of pigs was acclimated to the cold (10 °C) for a period of 10 days prior to the examination of tissues, but again uncoupling protein was not detected in any tissue. These results indicate that uncoupling protein is either absent from adipose tissues of the pig or is present at such a low concentration that it is unlikely to support thermogenesis. It is concluded that the pig does not contain adipose tissue that is functionally "brown;" adipose tissues in this species appear to be exclusively "white."Key words: brown adipose tissue, white adipose tissue, uncoupling protein, thermogenesis, immunoblotting.

The effect of dietary beta-carotene on colon carcinogenesis induced by 1,2-dimethylhydrazine [(DMH) CAS: 540-73-8] was studied in female inbred Swiss Webster (ICR) mice. At age 10 weeks and continuing throughout the experiment, mice received diets consisting mainly of natural foods (laboratory chow) and containing 2 or 22 mg beta-carotene/kg. At age 15 weeks they received 7 weekly sc injections of DMH (total dose: 196 mg DMH X diHCI/kg body wt). When autopsied 31 weeks after the first DMH injection, the incidence (percent of mice with tumors) and multiplicity (number of tumors/tumor-bearing mouse) of colon tumors were reduced by half in the mice supplemented with beta-carotene. There was a much greater decrease in adenocarcinomas than in adenomas. Mice observed for 13 additional weeks revealed that the mortality rate, due largely or wholly to colon cancer, was only about half in supplemented mice. Mice sacrificed 12 weeks after the first dose of DMH (i.e., well before tumors appeared) showed mild colon mucosal hyperplasia. beta-Carotene supplementation, however, did not alter this, indicating that the protective effect against colon cancer may have occurred at a late stage of carcinogenesis.

The influence of dietary selenium and cabbage on the formation of colon tumors in female Swiss mice treated with 1,2-dimethylhydrazine [(DMH) CAS: 540-73-8] was reported. Mice received a control diet (laboratory chow), the control diet plus selenium in the drinking water (1 mg/liter), or the control diet with added cabbage (12.8 g/100 g diet). They also received 8 weekly sc injections of DMH. The experiment was divided into two time periods: a) from 5 weeks before the first injection until 3 days after the last one (initiation period), and b) the subsequent 19.5 weeks until sacrifice of the mice (promotion period). Selenium had a strong protective effect when given during the initiation period; adenomas were reduced to a much greater extent than adenocarcinomas. The only effect of selenium supplementation in the promotion period was a small decrease in adenomas. Cabbage apparently had two opposing actions. It increased tumor incidence, particularly adenocarcinomas, if given in the initiation period, but it reduced adenoma formation considerably when given in the promotion period.

A method is described for measuring the deoxyribonucleic acid (DNA) content of small samples of adipose tissue or free fat cells. Lipids and acid-soluble nucleotides are first removed by extraction with a cold diethyl ether-ethanol mixture containing 10 per cent. m/V of trichloroacetic acid. DNA is then measured by hydrolysing the nucleoprotein residue in a 5 per cent. solution of trichloroacetic acid at 90 °C for 20 min, followed by treatment with p-nitro-phenylhydrazine and measurement of the hydrazone at 560 nm.Several aspects of the method have been critically examined in order to determine the optimum conditions. The method is satisfactorily reproducible.