Separation of milk fat globules via microfiltration: Effect of diafiltration media and opportunities for stream valorization.

Milk fat globule membranes (MFGM) sourced in buttermilk have gained recent interest given their nutritional value and functional properties. However, production of isolated MFGM has been challenging given their size similarity with casein micelles, which limits attempts toward fractionation by size exclusion techniques. Therefore, the hypothesis underpinning this study is that the removal of proteins from cream before butter-making facilitates MFGM isolation. As such, milk fat globules were separated from raw whole milk via microfiltration (1.4-µm pore diameter and 0.005-m2 filtration surface area) by using 3 diafiltration media; namely, skim milk ultrafiltration permeate, saline, and water. Their effects on the stability of the milk fat globules and protein permeation was elucidated. Whereas a substantial reduction in protein concentration was achieved with all diafiltration media (~90% reduction), water and saline produced negligible membrane fouling with better filtration performance. Moreover, diafiltration with skim milk ultrafiltration permeate exhibited reduced permeate flux. Colloidal stability of the resultant milk decreased with all diafiltration solutions due to changing composition and reduced apparent viscosity. Overall, microfiltration was found to be an efficient method for separation of milk fat globules from whole milk, leading to increased MFGM fragment concentration in buttermilk dry matter, thus making it more suitable for industrial utilization.

Epithelial-mesenchymal interactions in breast cancer: evidence for a role of nuclear localized ß-catenin in carcinoma-associated fibroblasts.

AIMS: Characteristics of the stroma around tumours are critical in defining the behaviour of cancers. ß-Catenin is well established as a critical regulator of carcinogenesis, acting as a transcriptional co-activator in the nuclei of epithelial cancer cells. We have examined the prevalence and influence of nuclear ß-catenin within the stromal fibroblasts of breast cancer. METHODS AND RESULTS: We examined ß-catenin expression in 201 breast cancers and adjacent normal tissue. Fibroblasts expressing nuclear ß-catenin were present in a significantly greater proportion of tumour tissues than normal tissues. The presence of fibroblasts with nuclear ß-catenin in tumours correlated with survival; tumours with prevalent positive fibroblasts were associated significantly with relatively good prognoses. Functional studies to examine influences of fibroblasts with nuclear ß-catenin, showed fibroblasts transfected to allow overexpression of ß-catenin were capable of inducing increases in both proliferation and invasion of breast cancer cell lines. CONCLUSION: The presence of fibroblasts with nuclear ß-catenin in tumours is a good prognostic indicator, although in the context of tissue culture models these cells can increase the growth and metastatic potential of cancer cells. These apparently paradoxical observations underline the complexity of epithelial-stromal signalling within tumours and highlight an area for further study.

Stearic acid, trans fatty acids, and dairy fat: effects on serum and lipoprotein lipids, apolipoproteins, lipoprotein(a), and lipid transfer proteins in healthy subjects.

To compare the effects on serum lipoproteins of stearic acid, trans fatty acids, and dairy fat, 80 healthy subjects consumed a dairy fat-based (baseline) diet for 5 wk, then an experimental diet high in either trans fatty acids (8.7% of energy; n = 40) or stearic acid (9.3% of energy; n = 40) for another 5 wk. All diets provided 32.2-33.9% of energy as fat, 14.6-15.8% as saturated plus trans fatty acids, 11.4-12.5% as cis-monounsaturated fatty acids, 2.9-3.5% as polyunsaturated fatty acids, and 200-221 mg cholesterol/10 MJ. Compared with the dairy fat diet, stearic acid and trans fatty acids decreased serum total cholesterol concentrations similarly (by 13% and 12%, respectively, P < 0.001) but the trans fatty acid diet decreased HDL cholesterol (17%) and apolipoprotein (apo) A-I (15%) significantly more than did the stearic acid diet (11% and 12%, respectively). Stearic acid but not trans fatty acids reduced concentrations of LDL cholesterol and apo B significantly (P < 0.001). The trans fatty acid diet increased the ratio of LDL to HDL cholesterol (19%) and of apo B to apo A-I (16%) more than did the dairy fat diet (P < 0.001) but the stearic acid diet had no effect. Lipoprotein(a) concentrations increased with both experimental diets, significantly more with trans fatty acids (30%) than with stearic acid (10%). In conclusion, high amounts of trans fatty acids had more adverse effects on lipoproteins than did equal amounts of stearic acid and dairy fat. Stearic acid reduced LDL cholesterol, did not affect the ratio of LDL to HDL cholesterol, and increased lipoprotein(a), although to a lesser extent than did trans fatty acids. Dietary fats low in both saturated fatty acids and trans fatty acids should be favored.

Mutations, tissue accumulations, and serum levels of p53 in patients with occupational cancers from asbestos and silica exposure.

In order to determine the relationship between mutations, tissue accumulations, and serum levels of p53 in occupational cancers, we used denaturing gradient gel electrophoresis and DNA sequencing of exons 5-9 of the p53 gene, immunohistochemical analysis for tissue identification of mutant p53 protein, and enzyme-linked immunosorbent assay for serum levels of mutant p53 protein to examine for such alteration in a cohort of individuals with workplace exposure to asbestos or silica, and resultant lung cancers or mesotheliomas. DNA analysis detected mutations in 5 of 18 (28%) tumors, and tissue accumulations of protein were detected in 7 of 20 (35%) tumors; the agreement between mutational and immunohistochemical analyses was significant (kappa = 0.62, P = 0.002). Serum elevations of protein were detected in 4 of 11 (36%) cases with available serum samples; the agreement between tissue alterations and serum elevations was also significant (kappa = 0.71, P = 0.017). In addition, based on the analysis of banked samples, serum results tended to be consistent over time prior to the diagnosis of disease (positive predictive value = 0.67, negative predictive value = 0.83). These results suggest that serum levels of p53 are reasonably accurate in reflecting tissue alterations in p53 at the gene and/ or protein level and may be early biomarkers of disease risk.

Serum oncoproteins in asbestosis patients.

Using ELISAs, we determined the concentrations of transforming growth factor alpha (TGF-alpha), the extracellular domain of the erbB-2 receptor (erbB-2 ECD), and mutant p53 protein in stored serum samples of asbestosis patients with and without cancer and control subjects (without asbestosis or cancer). The percentage of individuals in these three groups with increased serum concentrations of TGF-alpha, erbB-2 ECD, and mutant p53, respectively, were: asbestosis patients with cancer, 36%, 16%, 19%; asbestosis patients without cancer, 38%, 19%, 6%; control subjects, 0%, 5%, 10%. Although differences in serum positivity for these oncoproteins were apparent among these groups, the differences did not achieve statistical significance (P > 0.05). In several of the cancer cases, increased concentrations of TGF-alpha, erbB-2 ECD, and mutant p53 were also detected in the stored serum samples collected years before the clinical diagnosis of disease.

Tumour necrosis factor-alpha (TNF-alpha) in patients who have asbestosis and develop cancer.

OBJECTIVES: Concentrations of tumour necrosis factor-alpha (TNF-alpha) were assayed by radioimmunoassay in serum samples collected between 1981 and 1987 from 111 patients with asbestosis who were at a high risk of cancer. Follow up of these patients until 1993 showed that 38 had developed cancer (27 lung, three mesotheliomas, and eight diverse malignancies). RESULTS: The mean serum concentrations of TNF-alpha given in fmol/100 microliters serum in all the cases with cancer (14.1) and the cases with lung cancer (13.6) were significantly higher (P < 0.05) than the mean concentrations in the exposed controls (10.5). A positive increase was considered to be any value that was > 2 SDs above the mean of the exposed controls. 22% (six of 27) of the cases with lung cancer were positive compared with 4% (three of 73) of the exposed controls, a significant difference (P < 0.001). The serum concentrations of TNF-alpha correlated moderately with cancer (r = 0.3), lung cancer (r = 0.3), and Neu oncoproteins and epidermal growth factor receptor (EGFR) (r = 0.3, 0.5 respectively). Also, there was a significant correlation between development of cancer and severity or progression of asbestosis. There was no correlation between the concentrations of TNF-alpha and severity or progression of asbestosis. CONCLUSIONS: These results showed high concentrations of TNF-alpha in the patients who had cancer. TNF-alpha may offer an auxiliary method in early diagnosis of cancers related to asbestosis.

The detection of increased amounts of the extracellular domain of the epidermal growth factor receptor in serum during carcinogenesis in asbestosis patients.

Overexpression of the epidermal growth factor receptor (EGFr) has been implicated in the pathogenesis of a wide variety of human malignancies and may be related to asbestos-induced carcinogenesis. Overexpression of the EGFr can be detected immunologically by quantitation of the extracellular domain (ECD) in the extracellular fluid in vitro and in serum in vivo. An enzyme-linked immunosorbent assay (ELISA) for the EGFr ECD was used to examine banked serum samples of 38 asbestosis patients who subsequently developed cancer, 72 age-sex-race-smoking-asbestos exposure matched asbestosis controls without cancer, and 20 age-sex-race-smoking matched nonasbestosis noncancer controls. The mean serum level for the EGFr ECD in the cancer cases (636 +/- 299 fmol/ml) was statistically significantly elevated (P < 0.05) in comparison to the mean level in the asbestosis controls (546 +/- 147 fmol/ml) or the nonasbestosis controls (336 +/- 228 fmol/ml). Defining a positive elevation of the serum EGFr ECD as any value more than 2 standard deviations above the nonasbestosis control mean, 7 (18%) of the cancer cases were positive compared to 4 (6%) of the asbestosis controls and one (5%) of the nonasbestosis controls. In addition, all of these cancer cases had positive serum samples prior to the time of disease diagnosis (average = 5.1 years). These results suggest that serum EGFr ECD may be elevated at an early stage of carcinogenesis in some asbestosis patients and that further prospective study of the utility of this biomarker is warranted.

Serum levels of growth-factor receptors, egfr and Neu in asbestosis patients - a follow-up-study.

A cohort of asbestosis patients at high risk for cancer was investigated by the means of epidermal growth factor receptor and Neu oncoprotein levels in serum. The samples were collected between 1981 and 1987 from 111 asbestosis patients. X-ray pictures, taken three times during this period of time, were evaluated according to the ILO classification to deduce the severity of the disease (stable, unstable) and compared side by side to determine the progression of asbestosis. Follow-up of these patients until 1993 showed that 38 patients had developed cancer (27 lung, 3 pleural mesothelioma, 8 diverse malignancies). As determined by ELISA for EGFR and two different assays for Neu oncoprotein (Neu, nNeu) there was a statistically significant difference between EGFR levels in all the cancer and the non-lung cancer cases compared with the controls. No statistically significant differences were detected between Neu levels in the cancer cases and the controls. Protein concentrations showed no change prior the diagnosis of cancer. The serum levels of the proteins correlated with each other (EGFR-Neu, r=0.3; Neu-nNeu, r=0.7). Among 9 factors that were analyzed by multiple regression analysis progressive asbestosis was statistically significantly associated with the elevation of Neu and smoking with the elevation of nNeu. In logistic regression analysis only unstable asbestosis associated with cancer (OR = 6.25, p=0.0012). This study shows a coherence between the severity of asbestosis and development of cancer. Furthermore, the Neu concentration in the serum was elevated in both unstable and progressive asbestosis cases and may offer tools, supplementary to X-rays, for clinical monitoring of the pneumoconiosis patients.

Decreased gastric acid secretion and bacterial colonization of the stomach in severely malnourished Bangladeshi children.

To assess the effect of malnutrition on gastric acidity and gastric bacterial colonization, we studied 35 severely malnourished Bangladeshi children before (0 wk) and after (3 wk) they received nutritional rehabilitation for 3 wk. These results were compared with those obtained from a similarly examined group of 20 better-nourished Bangladeshi children. Gastric acid output, both basal and after betazole stimulation, was significantly lower in the malnourished group at 0 wk compared with the better-nourished children (p less than 0.01): basal 0.22 vs. 0.52 mEq HCl/h and stimulated 0.90 vs. 2.5 mEq HCl/h. Both the concentration of acid and the rate at which gastric juice was secreted were decreased in the malnourished group but serum gastrin levels were not significantly different. After 3 wk, the malnourished children had improved from 61% (+/- 9.0%; SD) to 81% (+/- 8.1%) of expected weight-for-height and were not significantly different than the better-nourished group (86% +/- 11%). Nevertheless, gastric acid concentration remained depressed in the 3-wk group, although the rate of gastric juice secretion equaled levels observed in the better-nourished group. None of the better-nourished children had detectable gram-negative bacterial colonization of their gastric juice. In contrast, 26 of 32 (81%) malnourished children at 0 wk were colonized--even after betazole stimulation, 11 of 33 (33%) gastric juice samples yielded viable organisms--suggesting that the decrease in gastric acid output greatly reduced the gastric acid barrier. Interestingly, only 9 of 20 (45%) better-nourished children had gastric juice with basal pH values below 4.0, suggesting that the gastric acid barrier may be an intermittent defense factor in Bangladeshi children.