A combination of probiotics and whey proteins enhances anti-obesity effects of calcium and dairy products during nutritional energy restriction in aP2-agouti transgenic mice.

Lactobacillus rhamnosus GG, Lactobacillus paracasei TMC0409, Streptococcus thermophilus TMC1543 and whey proteins were used to prepare fermented milk. For the experiment aP2- agouti transgenic mice were pre-treated with a high-sucrose/high-fat diet for 6 weeks to induce obesity. The obese mice were fed a diet containing 1·2% Ca and either non-fat dried milk (NFDM) or probiotic-fermented milk (PFM) with nutritional energy restriction for 6 weeks. The animals were examined after the treatment for changes in body weight, fat pad weight, fatty acid synthase (FAS) activity, lypolysis, the expression levels of genes related to lipid metabolism, insulin sensitivity in adipocytes and skeletal muscle and the presence of biomarkers for oxidative and inflammatory stress in plasma. It was found that the PFM diet significantly reduced body weight, fat accumulation, and adipocyte FAS activity, and increased adipocyte lipolysis as compared with the effects of the NFDM diet (P<0·05). The adipose tissue gene expression of 11ß-hydroxysteroid dehydrogenase 1 (11ß-HSD1) was significantly suppressed in mice that were fed PFM as compared with those that were fed NFDM (P<0·05). PFM caused a greater up-regulation of skeletal muscle PPARα, PPARδ, uncoupling protein 3 (UCP3) and GLUT4 expression and a significant decrease in the plasma concentration of insulin, malondialdehyde, TNF-α, monocyte chemotactic protein-1 and C-reactive protein as compared with the effects of NFDM (P<0·05). Fermentation of milk with selected probiotics and supplementation of milk with whey proteins may thus enhance anti-obesity effects of Ca and dairy products by the suppression of adipose tissue lipogenesis, activation of fat oxidation in skeletal muscle and reduction of oxidative and inflammatory stress.

Advanced application of porcine intramuscular adipocytes for evaluating anti-adipogenic and anti-inflammatory activities of immunobiotics.

We previously established a clonal porcine intramuscular preadipocyte (PIP) line and we were able to establish a protocol to obtain functional mature adipocytes from PIP cells. We hypothesized that both PIP cells and mature adipocytes are likely to be useful in vitro tools for increasing our understanding of immunobiology of adipose tissue, and for the selection and study of immunoregulatory probiotics (immunobiotics) able to modulate adipocytes immune responses. In this study, we investigated the immunobiology of PIP cells and mature adipocytes in relation to their response to TNF-α stimulation. In addition, we evaluated the possibility that immunobiotic microorganisms modify adipogenesis and immune functions of porcine adipose tissue through Peyer's patches (PPs) immune-competent cells. We treated the porcine PPs immune cells with different probiotic strains; and we evaluated the effect of conditioned media from probiotic-stimulated immune cells in PIP cells and mature adipocytes. The Lactobacillus GG and L. gasseri TMC0356 showed remarkable effects, and were able to significantly reduce the expression of pro-inflammatory factors and negative regulators (A20, Bcl-3, and MKP-1) in adipocytes challenged with TNF-α. The results of this study demonstrated that the evaluation of IL-6, and MCP-1 production, and A20 and Bcl-3 down-regulation in TNF-α-challenged adipocytes could function as biomarkers to screen and select potential immunobiotic strains. Taking into consideration that several in vivo and in vitro studies clearly demonstrated the beneficial effects of Lactobacillus GG and L. gasseri TMC0356 in adipose inflammation, the results presented in this work indicate that the PIP cells and porcine adipocytes could be used for the screening and the selection of new immunobiotic strains with the potential to functionally modulate adipose inflammation when orally administered.

Lactobacillus GG-fermented milk prevents DSS-induced colitis and regulates intestinal epithelial homeostasis through activation of epidermal growth factor receptor.

BACKGROUND: Fermented milk is considered one of the best sources for efficient consumption of probiotic strains by hosts to promote good health. The purpose of this study was to investigate the effects of orally administering LGG-fermented milk (LGG milk) on intestinal inflammation and injury and to study the mechanisms of LGG milk's action. METHODS: LGG milk and non-LGG-fermented milk (non-LGG milk) were administered through gavage to mice before and during dextran sodium sulfate (DSS)-induced intestinal injury and colitis. Inflammatory/injury score and colon length were assessed. Intestinal epithelial cells were treated with the soluble fraction of LGG milk to detect its effects on the epidermal growth factor receptor (EGFR) and its downstream target, Akt activation, cytokine-induced apoptosis, and hydrogen peroxide (H2O2)-induced disruption of tight junctions. RESULTS: LGG milk treatment significantly reduced DSS-induced colonic inflammation and injury, and colon shortening in mice, compared to that in non-LGG milk-treated and -untreated mice. The soluble fraction of LGG milk, but not non-LGG milk, stimulated the activation of EGFR and Akt in a concentration-dependent manner, suppressed cytokine-induced apoptosis, and attenuated H2O2-induced disruption of tight junction complex in the intestinal epithelial cells. These effects of LGG milk were blocked by the EGFR kinase inhibitor. LGG milk, but not non-LGG milk, contained two soluble proteins, p40 and p75, that have been reported to promote survival and growth of intestinal epithelial cells through the activation of EGFR. Depletion of p40 and p75 from LGG milk abolished the effects of LGG milk on prevention of cytokine-induced apoptosis and H2O2-induced disruption of tight junctions. CONCLUSIONS: These results suggest that LGG milk may regulate intestinal epithelial homeostasis and potentially prevent intestinal inflammatory diseases through activation of EGFR by LGG-derived proteins.

Evaluation of the age-related changes in movement smoothness in the lower extremity joints during lifting.

The purpose of this study was to analyze age-related movement smoothness changes in the lower extremity joints during load lifting. A total of 10 young and 13 elderly subjects participated in the study. Infrared reflective markers were attached to body landmarks in each subject. While the subjects stood on force plates and lifted a box, the marker displacements and ground reaction forces were measured using a 3D motion analysis system. The jerk square mean value (JSM) was defined as the lower extremity joint movement smoothness index during lifting. JSM represented the average of the square of the joint angle third derivative value, according to the jerk third derivative of the position data. Each subject's JSM values were calculated for the hip, knee and ankle joints. Movement smoothness appeared to decrease as JSM increased. Multiple regression analyses were performed for dependent variables (hip, knee and ankle joint JSM values) and independent variables (age, hand grip strength, sex difference and lifting duration). The level of significance was set at p<0.05. For the hip joint JSM, the regression coefficient for age was significantly positive and that for lifting duration was significantly negative. For the knee joint JSM, the regression coefficient for lifting duration was significantly negative. For the ankle joint JSM, the regression coefficients for age and hand grip strength were significantly positive and that for lifting duration was significantly negative. These results suggest that movement smoothness in the hip and ankle joints during lifting decreases with advancing age.

Noninvasive estimation of muscle fiber conduction velocity distribution using an electromyographic processing technique.

BACKGROUND: Electromyography (EMG) is useful in investigating muscle activation; however, noninvasive evaluation of surface EMG is limited due to its complicated waveform. This study investigated muscle structure and activation using an analysis technique for surface EMG. MATERIAL/METHODS: Surface array electrodes were used in 17 healthy male subjects to record eight-channel EMGs from each biceps brachii muscle during voluntary isometric contraction with a 1-kg weight band with the subjects seated. The peaks detected by referenced EMGs were normalized and averaged as averaged pulses (APs) and the innervation zone (IZ) was estimated from the APs. Muscle fiber conduction velocities (MFCVs), estimated by the time difference of the peaks (method P) and by cross-correlation (method CC) by APs, were compared. Time periods with positive values around the central peak in AP (PP) were measured and the contribution of MFCVs by motor unit action potentials (MUAPs) was estimated. RESULTS: IZs were estimated in 12 subjects. Near the IZ, correlation between MFCVs by methods P and CC was lower than in other locations; MFCV was significantly larger by method P than by method CC in the vicinity of the IZ. PP of the comparison AP was significantly larger than that of the reference AP. The distribution of the MFCVs by different MUAPs was verified by computer simulation. CONCLUSIONS: Surface EMG was used to demonstrate the diversity of MFCVs, with some increased MFCVs, for several MUAPs in the vicinity of the IZ. This method could be applied to the evaluation of neuromuscular disorders.

Effect of occlusion status on the time required for initiation of recovery in response to external disturbances in the standing position.

BACKGROUND: To examine whether occlusion status contributes to improvement of postural balance. METHODS: Thirty healthy adolescents (15 males and 15 females; mean age, 20.3; standard deviation (SD) 1.6 years) with no equilibrium or stomatognathic function abnormalities were examined. Occlusion is a term meaning "jaw clenching." Occlusion status was evaluated by measuring masseter activity using the EMG system. Balancing ability was evaluated using the EquiTest system, which measures sway of the center of gravity produced by rapid movement of force plates as an external disturbance (three intensity levels). The time required for initiation of recovery after application of the disturbance was calculated by measuring displacement of the center of foot pressure. Data were compared according to occlusion status. FINDINGS: Little difference in latency was observed following a small disturbances; however, the greater the disturbance the shorter the latency with occlusion, while without occlusion, latency increased with increasing disturbance. A statistically significant interaction (P<0.001) between occlusion and external disturbance was also found. INTERPRETATION: This study suggested that occlusion contributes to maintenance of postural balance and improvement of stability when unexpected sway occurs in the standing position.

CAD/CAM evaluation of the fit of trans-tibial sockets for trans-tibial amputation stumps.

The purpose of this study was to objectively evaluate the fit of sockets for trans-tibial stumps in order to establish a guideline for use in the automated production of prosthetic sockets. Subjects were 24 trans-tibial amputees. Using a CAD/CAM system, 11 parameters regarding the 3D shape of the stumps and the sockets were objectively evaluated. A correlation was found between the activity level and the upper and lower volumes of the socket, and between the cause of amputation and the upper volume of the socket. It was considered desirable to make the lower part of the socket looser for patients with lower activity levels, to make the upper part tighter for patients with higher activity levels, and to make the upper part looser for amputation patients with peripheral circulatory diseases.

Effect of orally administered non-viable Lactobacillus cells on murine humoral immune responses.

BALB/c mice were immunized intraperitoneally with the food antigen ovalbumin (OVA) while they were fed with Lactobacillus GG heated killed cell preparation. The oral administration of Lactobacillus GG did not appear to modify the antigen-augmented serum IgE in the tested mice but significantly augmented serum OVA specific IgG in the tested mice fed with a diet containing 0.1% Lactobacillus GG as the non-viable cell preparation (P< 0.05). The fecal OVA specific IgA of the tested mice fed with nonviable Lactobacillus GG cells was also significantly elevated (P< 0.05) compared to those from OVA immunized mice. The spleen cells of mice fed with non-viable Lactobacillus GG cells secreted more IL-6 (P< 0.01). These results suggest that the non-viable Lactobacillus GG can augment the systemic and mucosal immune responses in a host animal favoring secretory IgA but not IgE in an adjuvant-like manner.

Strong immunostimulation in murine immune cells by Lactobacillus rhamnosus GG DNA containing novel oligodeoxynucleotide pattern.

Whole cells, cell wall components and some soluble factors from Lactobacillus rhamnosus GG (LGG) are known to invoke immune responses as they interact with animal and human immune cells. In the present study, we found that chromosomal DNA from LGG is a potent inducer of splenic B cell proliferation, CD86/CD69 expression and cytokine production in mice. In the genomic DNA of LGG we discovered TTTCGTTT oligodeoxynucleotide (ODN) ID35, which has a potent activity in a number of immunostimulatory assays. Phosphorothioate backbone is not required for the activity of ID35. The ODN ID35 showed levels of activity comparable with those induced by the murine prototype ODN 1826 in B cell proliferation, CD86/CD69 expression, interleukin (IL)-6, IL-12, IL-18, interferon gamma (IFN-gamma) and tumour necrosis factor alpha (TNF-alpha) mRNA expression and IFN-gamma/IL-12p70 protein production assays. Additionally, ID35 appeared to be equally active in both murine and human immune cells. These stimulatory effects are due to TTTCGTTT motif located in the 5' end of ID35. In this study we demonstrate for a first time that, DNA from LGG is a factor of immunobiotic activity. Furthermore, ODN ID35 is the first ODN, with such a strong immunostimulatory activity to be found in immunobiotic bacterial DNA.

Antidiabetic effect of Lactobacillus GG in streptozotocin-induced diabetic rats.

Neonatally streptozotocin-induced diabetic (n-STZ) rats were given food containing Lactobacillus GG cells (GG) or a control diet (control), from 9 to 18 weeks of age. The GG cells significantly lowered the blood hemoglobin A(1C) (HbA(1C)) level and improved glucose tolerance in n-STZ rats (p<0.05). In the GG group, the serum insulin level at 30 min after glucose loading was significantly higher than in the control group (p<0.05).

Cytokine production by the murine macrophage cell line J774.1 after exposure to lactobacilli.

Eleven strains of lactobacilli were tested for their ability to induce the murine macrophage-like cell line J774.1 to secrete cytokines. Some of the bacteria tested induce the production of interleukin(IL) 6, IL-12, and tumor necrosis factor a (TNF-alpha) by J774.1 cells. Seven strains also induced the production of IL-10. However, no IL-1beta was produced. Lactobacillus acidophilus TMC 0356 significantly induced the production of more IL-6, IL-10, IL-12, and TNF-alpha than the other bacteria tested (p < 0.0001; ANOVA). These results suggest that lactobacilli can activate macrophages to secrete both inflammatory and anti-inflammatory cytokines. Selected strains might be used to bring about pro or antiinflammatory immune reactions.

Adhesion of lactic acid bacteria to caco-2 cells and their effect on cytokine secretion.

Cytokines secreted by human enterocytes play a critical role in mucosal and systemic immunity. Intestinal microorganisms can influence this secretion. In the present study, 30 strains of lactic acid bacteria were characterized for their adhesion to Caco-2 cells and their potential to stimulate proinflammatory cytokine secretion by this cell line. The bacteria adhered in a strain-dependent manner to Caco-2 cells. Contact with lactobacilli did not result in the production of IL-6 or IL-8. A slight IL-6 and IL-8 production by a Caco-2 cell was detected after exposure to 8 of the tested Bifidobacterium strains. No correlation was found between adhesion and cytokine induction among the bacteria tested. This indicates that lactic acid bacteria, even those with strong adhesive properties, are not very likely to trigger an inflammatory response in human enterocytes.

Stimulation of the secretion of pro-inflammatory cytokines by Bifidobacterium strains.

To characterize the ability of bifidobacteria to affect the production of macrophage-derived cytokines, a murine macrophage-like cell line, J774.1, was cultured in the presence of 27 strains of heat-inactivated bifidobacteria. Bifidobacterium adolescentis and B. longum, known as adult-type bifidobacteria, induced significantly more pro-inflammatory cytokine secretion, IL-12 and TNF-alpha, by J774.1 cells, than did the infant-type bifidobacteria, B. bifidum, B. breve, and B. infantis (P<0.01). In contrast, B. adolescentis did not stimulate the production of anti-inflammatory IL-10 from J774.1 cells as the other tested bacteria did. The results suggest that the adult-type bifidobacteria, especially B. adolescentis, may be more potent to amplify but less able to down-regulate the inflammatory response.