Effect of hydrolysed egg protein on brain tryptophan availability.

Serotonin synthesis critically depends on plasma levels of tryptophan (TRP). Earlier studies have shown that for mood and cognitive benefits to occur, the ratio between TRP and other large neutral amino acids (LNAA) has to be increased by approximately 40 %. The present study investigated the dose-dependent effects of a TRP-rich hydrolysed protein (egg-protein hydrolysate, EPH) on the plasma TRP:LNAA. Moreover, it was investigated whether EPH could increase TRP:LNAA in the presence of 2 g of milk protein (MP). In a randomised double-blind crossover design, plasma amino acids were measured every 30 min for 3·5 h after ingestion of a drink containing either three different doses of 4, 8 and 12 g EPH containing 270, 560 or 800 mg of TRP, respectively, the combination of 4 g EPH and 2 g MP (74 mg TRP), or 4 g MP (148 mg TRP) in twenty healthy subjects with a mean age of 52 years. All three EPH doses caused significant increases of TRP:LNAA above 40 % at 30, 60 and 90 min after consumption in a dose-dependent manner. Compared with the 4 g EPH, the increase in TRP:LNAA in the 4 g EPH with 2 g MP condition was significantly lower at 60 min (63 v. 44 %, P < 0·001) and did not differ significantly at 90 min (58 v. 53 %, P>0·05). The present study showed that a low dose of 4 g EPH with even the addition of 2 g MP was sufficient to increase the ratio of TRP:LNAA above 40 %. Thus, EPH offers a viable ingredient to increase TRP availability.

Hydrolyzed casein decreases postprandial glucose concentrations in T2DM patients irrespective of leucine content.

Lifestyle modifications, including diet, are important in the prevention and management of type 2 diabetes mellitus (T2DM). However, limited information is available on the effects of single doses of meal replacements, particularly with regard to their effect on postprandial glucose. Therefore, a study was performed comparing the effects of a single meal replacement in T2DM patients on postprandial serum glucose, insulin, and glucagon. This randomized, double-blind, partial cross-over study was performed in 36 T2DM patients who continued their oral anti-diabetic medication. Each patient received three out of four treatments separated by 7 days. The treatments were a proprietary casein hydrolysate (insuVida™) alone or with additional leucine, unhydrolyzed casein, or placebo. Blood sampling was done for 4 hr. Treatments were compared using repeated measures ANOVA. Results are given as an estimate of the difference (%) for the 4-hr epoch. Glucose concentrations were lowered by -4.7% by insuVida and insuVida plus added leucine compared to placebo (95% CI: -1.6 to -7.7%), while the effect of unhydrolyzed casein was -1.7% (-4.8 to 1.5%). Addition of leucine to insuVida induced the greatest increase in insulin (i.e., 51.8%; 41.1 to 63.4%). All three treatments increased glucagon concentrations by 14% (8 to 20%) compared to placebo. A single dose of insuVida™ with or without addition of leucine significantly lowered plasma glucose compared to placebo and intact casein in T2DM patients. This is most likely due to an insulinotropic effect of insuVida. The data suggest that this type of intervention may be a viable treatment strategy in T2DM.

Effect of different tryptophan sources on amino acids availability to the brain and mood in healthy volunteers.

RATIONALE: Reduced brain serotonin function is acknowledged as a vulnerability factor for affective disturbances. Since the production of serotonin is limited by the availability of its plasma dietary amino acid precursor tryptophan (TRP), the beneficial effects of tryptophan-rich alpha-lactalbumin whey protein (ALAC) have recently been studied. The effects of ALAC remain rather modest, and alternative protein sources of tryptophan may be more effective. OBJECTIVES: We tested whether hydrolyzed protein (HPROT) has greater effects on the plasma TRP/large neutral amino acids (LNAA) ratio and mood than intact ALAC protein in healthy volunteers. MATERIALS AND METHODS: In a double-blind, randomized cross-over study, plasma amino acids and mood were repeatedly measured in 18 healthy subjects before and after intake of ALAC and HPROT as well as after placebo protein, pure tryptophan, and a tryptophan-containing synthetic peptide. Except for the placebo protein, all interventions contained 0.8 g TRP. RESULTS: Significantly faster and greater increases in plasma TRP/LNAA were found after HPROT than after ALAC. In addition, the effects of HPROT on plasma TRP/LNAA were comparable with the effects of the tryptophan-containing synthetic peptide and even exceeded the effect of pure tryptophan. Sixty minutes after intake, mood was improved only following intake of HPROT and pure tryptophan, whereas longer-lasting mood effects were only found after intake of HPROT. CONCLUSIONS: The use of a tryptophan-rich hydrolyzed protein source may be more adequate to increase brain tryptophan and 5-HT function compared with intact alpha-lactalbumin protein or pure tryptophan.

Protein hydrolysates induce CCK release from enteroendocrine cells and act as partial agonists of the CCK1 receptor.

Protein has been reported to be the most satiating of all macronutrients. Upon gastrointestinal digestion, peptides are generated that stimulate the release of satiety hormones such as cholecystokinin (CCK) from enteroendocrine cells. As such, bioactive peptides could be the target of Functional Food ingredients with satiating effects. We set up an in vitro assay system to investigate if different protein hydrolysates exhibit varying CCK-releasing properties. Soy, pea, potato, casein, and whey protein hydrolysates were incubated with the enteric endocrine cell line STC-1 that endogenously expresses and secretes CCK. Release of CCK was measured by ELISA. All hydrolysates induced CCK release at low concentrations (>0.1 mg.L -1)); however, no significant differences in CCK-releasing properties between the different protein hydrolysates were found, suggesting a generic, nonspecific peptide-sensing mechanism in the STC-1 cells on hydrolyzed protein. As the ELISA exhibits sensitivity to all CCK isoforms possessing the C-terminal CCK octapeptide but varying in biological activity at the CCK 1 receptor (CCK 1R), a secondary module was added to the STC-1 cell assay. Intracellular calcium measurements were performed in CHO-CCK 1R cells. Following exposure of the STC-1 cells to the protein hydrolysates, the medium was tested on the CCK 1R assay. Released CCK was measured with higher sensitivity and lower variability than in the ELISA. Surprisingly, we found that some protein hydrolysates (soy > potato >> casein) also directly stimulated CCK 1R-expressing cells, while whey and pea protein hydrolysates were inactive. As CCK 1R is expressed in the GI tract, direct interaction of CCK 1R with dietary peptides may contribute to their satiety effects. Future experiments developing bioactive ingredients for Functional Foods for weight management could involve isolation of the active, CCK 1R-activating peptides in, for example, soy protein hydrolysates.

Comparison of 3 AT1 receptor binding assays: filtration assay, ScreenReady Target, and WGA Flashplate.

In this article, the study of 3 different angiotensin II type 1 (AT(1)) receptor binding assays in terms of reproducibility, robustness, and feasibility for high-throughput screening (HTS) is described. The following methods were used: a nonhomogeneous filtration assay in a 96-well format using CHO-AT(1) cell membranes and 2 homogeneous assays, which include the commercially available ScreenReady Target for the AT(1) receptor and the wheat germ agglutinin (WGA) Flashplate, which was coated "in-house" with the CHO-AT(1) cell membranes. Receptors were labeled with [(125)I]-Sar(1)-Ile(8)-angiotensin II, and radioligand binding was displaced using the antagonist losartan and the natural agonist angiotensin II. Reproducible K(d), B(max), and K(i) values and good total binding/nonspecific binding (TB/NSB) ratios were obtained with both the ScreenReady Targets and the filtration assay, whereas the WGA Flashplates showed unacceptably high nonspecific binding and high variation when applied as a homogeneous assay. However, when applied as a heterogeneous assay (i.e., when a wash step at the end of the assay is included), the results were significantly better. Interestingly, ligand affinities were consistently lower in Flashplate-based assays than in the filtration assay. This may be due to the immobilization of the receptors onto the solid surface of the plate, affecting their conformation. In terms of reproducibility, robustness, and feasibility for HTS, the authors conclude that the ScreenReady Target plates are most suitable for AT(1) receptor binding screening.

Human adipose cells express CD4, CXCR4, and CCR5 [corrected] receptors: a new target cell type for the immunodeficiency virus-1?

The concept that adipocytes belong to an essential endocrine system with some characteristics of immune cells has recently emerged. The aim of this paper is to present evidence of the expression of CD4, CXCR4, and CCR5 receptors by human adipocytes and to test whether adipose cells support HIV entry. Primary human preadipocytes were cultured and differentiated in vitro. Expression of the three receptors on preadipocytes and adipocytes was demonstrated by reverse transcriptase-polymerase chain reaction, immunocytochemical, and immunohistochemical analysis. Infection of adipose cells to HIV-1 was then investigated. The measurement of the viral p24 antigen in preadipocyte culture medium showed an increase of p24 levels between 24 and 72 h postexposure and then a progressive decrease to reach a low level at 10-15 days. Ten days after the infection test, supernatant of preadipocytes contained infectious particles able to infect the susceptible T-CD4 CEM cell line. The expression of viral proteins by adipocytes was confirmed using a fusion test. The presence of viral DNA was exhibited by gag-specific polymerase chain reaction, supporting the hypothesis of HIV-1 X4- and R5-virus entry in preadipocytes. Adipose cells represent the first cell type that does not belong to the immune system expressing all specific HIV receptors and may represent new HIV-1 target cells.