Hyunjoon Park

Functional properties of Lactobacillus strains isolated from kimchi

The objective of this study was to evaluate the functional properties of lactic acid bacteria (LAB) from kimchi, a traditional Korean fermented vegetable product generally consumed raw as a side-dish with practically every meal. Twelve mild acid producing facultatively heterofermentative Lactobacillus strains were selected for their potential as starter cultures for fermentation of kimchi, and evaluated for their functional properties. Eleven strains were identified as Lactobacillus sakei and one as Lactobacillus plantarum. The strains identified as L. sakei differed in some physiological features; of particular interest was the fact that 9 of these strains produced L(+) lactic acid from glucose in presence of acetate. All strains were able to survive gastrointestinal conditions simulating stomach and duodenum passage. In addition, they showed higher adherence to HT-29 cells than Lactobacillus rhamnosus GG, a commercial probiotic strain used worldwide. These strains also showed antimicrobial activity against a number of food-borne pathogens. Their ability to lower cholesterol was demonstrated by BSH (bile salt hydrolytic) activity, and cholesterol assimilation tests in vitro. The results suggest the probiotic potential of these strains for use in kimchi fermentation.

Modulation of the murine microbiome with a concomitant anti-obesity effect by Lactobacillus rhamnosus GG and Lactobacillus sakei NR28

The microbiota of the gastrointestinal tract (GIT) constitutes the major part of the total human microbiome and is considered to be an important regulator of human health and host metabolism. Numerous investigations in recent years have focused on the connection between the human microbiota and metabolic diseases such as obesity, type II diabetes and atherosclerosis. Yet, little is known about the impact of probiotic consumption on the GIT microbial population and the potential effect on chronic diseases. In this study, the modulation of the microbial community in the murine small intestine resulting from probiotic feeding was investigated and was found to be associated with an anti-obesity effect. Changes in the microbiota of the mouse faeces and small intestine were monitored using quantitative real-time PCR and by following the mRNA expression levels of various obesity-related biomarkers following probiotic feeding in a mouse model. Lactobacillus rhamnosus GG and Lactobacillus sakei NR28 (a putative probiotic strain isolated from kimchi) were administered at a daily level of approximately 1×10(8) viable bacteria per mouse (C57BL/6J mice) for up to three weeks. Feeding these strains resulted in a significant reduction of epididymal fat mass, as well as obesity-related biomarkers like acetyl-CoA carboxylase, fatty acid synthase, and stearoyl-CoA desaturase-1 in the liver. The total number and ratio of the microbial groups, i.e. Firmicutes, Bacteroidetes, Clostridium cluster I and XIVab, and Lactobacillus spp. were modulated in the small intestine, and the Firmicutes:Bacteroidetes ratio was decreased. In contrast, no noticeable effect of probiotic feeding could be detected on the faecal microbiota, neither quantitatively, nor with regard to the bacterial groups (Firmicutes, Bacteroidetes, Clostridium cluster I and XIVab, and Lactobacillus spp.) studied.

Selection of functional lactic acid bacteria as starter cultures for the fermentation of Korean leek (Allium tuberosum Rottler ex Sprengel.).

The purpose of this research was to find safe and suitable starter cultures for the fermentation of Korean leek (Allium tuberosum Rottler), also known as garlic chives or Oriental garlic. This traditional herb has several functional properties and a strong flavour; its leaves are used as food material. Eighteen strains of lactic acid bacteria (LAB) were isolated from well-fermented leek kimchi. Controlled fermentation of the leek leaves was conducted with 2 strains (Weissella confusa LK4 and Lactobacillus plantarum LK8), selected as potential starter cultures on the basis of their safety properties, and on the pH, total titratable acidity (TTA), and viable cell numbers [colony forming units (CFUml(-1))] achieved during the fermentation. Microbial dynamics was also followed during fermentation by using PCR-DGGE (Denaturing Gradient Gel Electrophoresis) on DNA level. To analyse bioactive compounds such as thiols and allicin (diallyl thiosulfinates), the total flavonoid and polyphenolic contents were determined by colorimetric methods. Functional properties were assessed on the basis of anti-oxidative capacities by determining the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging effect, and ferric reducing antioxidant power (FRAP). W. confusa LK4 rapidly increased during the first stage of leek fermentation, and was mainly responsible for accelerated fermentation during the early period in contrast to L. plantarum LK8, a stronger acid producer during the later stages of fermentation. After 48 h fermentation, leeks fermented with W. confusa LK4 showed the highest radical scavenging effects and reducing ability. The detectable amount of allicin of fermented leeks decreased relative to the change in pH, whereas the concentration of thiols significantly increased. Total flavonoid and poly-phenolic contents changed during fermentation and showed correlation with anti-oxidant effects. We therefore suggest the suitability of W. confusa LK4 as a potential starter culture for fermentation of leeks.

The probiotic Lactobacillus rhamnosus BFE5264 and Lactobacillus plantarum NR74 promote cholesterol efflux and suppress inflammation in THP‐1 cells

BACKGROUND The balance between the rate of cholesterol uptake/accumulation and the rate of cholesterol efflux is reflected in the amount of lipid accumulation in macrophages. Based upon the fact that liver X receptors (LXRs) play a role in cholesterol efflux, we studied the effects of probiotics on cholesterol efflux and anti-inflammatory action in macrophages. We confirmed changes in LXR expression by treatment of LXR-transfected CHO-K1 cells with lactic acid bacteria (LAB), and co-cultured THP-1 cells with LAB to investigate changes in cholesterol efflux and inflammation. RESULTS The experiment with CHO-K1 cells showed upregulation of LXR-β by LAB. Treatment of THP-1 cells with LAB promoted LXR expression in THP-1, which eventually led to significant upregulation of ABCA1 and ABCG1 expression. The treatment with live LAB also significantly promoted cholesterol efflux. LAB suppressed expression of interleukin (IL)-1β and tumor necrosis factor (TNF)-α, which resulted from activation of LXR. CONCLUSION Our study shows that Lactobacillus rhamnosus BFE5264 and Lactobacillus plantarum NR74 activated LXR and induced cholesterol efflux by promoting expression of ABCA1 and ABCG1. Both strains also suppressed proinflammatory cytokines including IL-1β and TNF-α. This study could account for the observation that LAB may block foam cell formation by cholesterol efflux and immune modulation.

Anti-obesity and antioxidative effects of purple sweet potato extract in 3T3-L1 adipocytes in vitro.

The purpose of the current study was to determine the anti-obesity and anti-inflammatory effects of an extract of purple sweet potatoes (PSPs) on 3T3-L1 adipocytes. For this purpose, differentiated 3T3-L1 adipocytes were treated with a PSP extract at concentrations of 1,000, 2,000, and 3,000 μg/mL for 24 hours. Then, we measured the changes in the sizes of the adipocytes, the secretion of leptin, and the mRNA/protein expression of lipogenic, inflammatory, and lipolytic factors after the treatment with the PSP extract. The PSP extract diminished leptin secretion, indicating that growth of fat droplets was suppressed. The extract also suppressed the expression of mRNAs of lipogenic and inflammatory factors and promoted lipolytic action. The antioxidative activity of the PSP extract was also measured using three different in vitro methods: 1,1-diphenyl-2-picrylhydrazyl free radical scavenging activity, ferric reducing ability potential assay, and chelating activity of transition metal ions. Taken together, our study shows that PSP extract has antilipogenic, anti-inflammatory, and lipolytic effects on adipocytes and has radical scavenging and reducing activity.

Reduction in cholesterol absorption in Caco-2 cells through the down-regulation of Niemann-Pick C1-like 1 by the putative probiotic strains Lactobacillus rhamnosus BFE5264 and Lactobacillus plantarum NR74 from fermented foods.

Hypercholesterolaemia is a major risk factor related to atherosclerosis, and it may be influenced by our diet. This study addresses the impact of Lactobacillus rhamnosus BFE5264 (isolated from Maasai fermented milk) and Lactobacillus plantarum NR74 (from Korean kimchi) on the control of cholesterol absorption through down-regulation of Niemann-Pick C1-like 1 (NPC1L1) expression. Caco-2 enterocytes were treated with the live, heat-killed (HK) bacteria, bacterial cell wall extracts and metabolites; mRNA level and protein expression were measured. Caco-2 cells showed lower NPC1L1 expression in the presence of the live test strains than the control, elucidating down-regulation of cholesterol uptake, and were compared well with the positive control, L. rhamnosus GG. This effect was also observed with HK bacteria and cell wall fractions but not with their metabolites. The potential of some Lactobacillus strains associated with traditional fermented foods to suppress cholesterol uptake and promote its efflux in enterocytes has been suggested from these data.

Lactobacillus plantarum HAC01 regulates gut microbiota and adipose tissue accumulation in a diet-induced obesity murine model

The functional features of Lactobacillus plantarum HAC01 (HAC01), isolated from fermented Korean kimchi, were studied with regard to the fat mass, immunometabolic biomarkers and dysbiosis in a diet-induced obesity (DIO) murine model. L. rhamnosus GG (LGG) served as reference strain and a PBS-treated group as control. The administration of L. plantarum HAC01 resulted in reduction of the mesenteric adipose depot, the conjunctive tissue closely associated with the gastrointestinal tract, where lipid oxidative gene expression was upregulated compared to the control group. Metagenome analysis of intestinal microbiota showed that both strains HAC01 and LGG influenced specific bacterial families such as the Lachnospiraceae and Ruminococcaceae rather than the phyla Firmicutes and Bacteroidetes as a whole. The relative abundance of the Lachnospiraceae (phylum Firmicutes) was significantly higher in both LAB-treated groups than in the control. Comparing the impact of the two Lactobacillus strains on microbial composition in the gut also suggests strain-specific effects. The study emphasises the need for deeper studies into functional specificity of a probiotic organism at the strain level. Alleviation of obesity-associated dysbiosis by modulation of the gut microbiota appears to be associated with “indicator” bacterial taxa such as the family Lachnospiraceae. This may provide further insight into mechanisms basic to the mode of probiotic action against obesity and associated dysbiosis.

Lactobacillus rhamnosus BFE 5264 and Lactobacillus plantarum NR74 Promote Cholesterol Excretion Through the Up-Regulation of ABCG5/8 in Caco-2 Cells.

The effect of two putative probiotic strains, Lactobacillus rhamnosus BFE5264 and Lactobacillus plantarum NR74, on the control of cholesterol efflux in enterocytes was assessed by focusing on the promotion of ATP-binding cassette sub-family G members 5 and 8 (ABCG5 and ABCG8). Differentiated Caco-2 enterocytes were treated with live bacteria, heat-killed bacteria, a bacterial cell wall fraction, and metabolites and were subjected to cholesterol uptake assay, mRNA analysis, and protein analyses. Following LXR-transfection by incubation with CHO-K1 cells in DNA-lipofectin added media, the luciferase assay was conducted for LXR analysis. Treatment of Caco-2 cells with L. rhamnosus BFE5264 (isolated from traditional fermented Maasai milk) and L. plantarum NR74 (isolated from Korean kimchi) resulted in the up-regulation of LXR, concomitantly with the elevated expression of ABCG5 and ABCG8. This was associated with the promotion of cholesterol efflux at significantly higher levels compared to the positive control strain L. rhamnosus GG (LGG). The experiment with CHO-K1 cells confirmed up-regulation of LXR-beta by the test strains, and treatment with the live L. rhamnosus BFE5264 and L. plantarum NR74 strains significantly increased cholesterol efflux. Heat-killed cells and cell wall fractions of both LAB strains induced the upregulation of ABCG5/8 through LXR activation. By contrast, LAB metabolites did not show any effect on ABCG5/8 and LXR expression. Data from this study suggest that LAB strains, such as L. rhamnosus BFE5264 and L. plantarum NR74, may promote cholesterol efflux in enterocytes, and thus potentially contribute to the prevention of hypercholesterolemia and atherosclerosis.

Influence of gastrointestinal stress on autoinducer-2 activity of two Lactobacillus species

Quorum sensing is a bacterial communication signalling system that regulates the expression of certain target genes with autoinducers in a cell density-dependent manner. The universal luxS-mediated quorum sensing using the autoinducer-2 (AI-2) signal is present in a wide variety of bacteria with only sparse information on probiotic lactobacilli. Effective probiotics should exhibit tolerance and adaptation to stress conditions typical of the GIT. Adhesion to human intestinal epithelial cells and competitive exclusion of pathogens are also considered important. The AI-2 signal system plays an important role in the response of probiotic lactobacilli to the surrounding environment. Intraspecies-related changes in quorum signalling in the GIT were determined by monitoring the AI-2 activity of two strains each of Lactobacillus rhamnosus and L. plantarum under various stress conditions. Modulation of the AI-2 activity of all the strains was induced by stress responses to pH, bile acid, temperature, osmotic pressure and starvation, and was both species- and strain-specific. AI-2 inhibition correlated with a reduction in the stress-related genes of L. rhamnosus. We therefore suggest that AI-2 quorum signalling of probiotic lactobacilli may represent one way of adapting to the hosts ecosystem and of interacting within the intestinal environment.

Development of putative probiotics as feed additives: validation in a porcine-specific gastrointestinal tract model

Enforced restrictions on the use of antibiotics as growth promoters (AGPs) in animal production have prompted investigations into alternative feed additives in recent decades. Probiotics are currently the main feed additive used in livestock. However, the selection of probiotic candidates relies on human-based methods and little is known about the verification criteria for host-specific selection. We investigated the probiotic potential of Lactobacillus salivarius strains isolated from fed pig feces for their use as porcine feed additives. Two methods were developed that simulated the pig gastrointestinal (GI) tract and the intestinal epithelium, and these were compared with human-based in vitro methods and used for selecting porcine probiotics. Lactobacillus salivarius strain LS6 was identified as a promising probiotic strain for potential use as a porcine feed additive. This strain prevented disruption of the epithelial integrity of pig small intestine (PSI) cells by inhibiting the adherence of enterotoxigenic Escherichia coli K88. It also showed high survival rates in the in vitro pig GI tract model and good adhesion to PSI cells. We propose that host target-specific screening and validation methods are important tools in the development of effective probiotic feed additives, and this approach may support future-oriented agriculture.