Effect of heat-killed Lactobacillus brevis SBC8803 on cutaneous arterial sympathetic nerve activity, cutaneous blood flow and transepidermal water loss in rats.

AIM: To evaluate the efficacy of the effects of heat-killed Lactobacillus brevis SBC8803 (HK-SBC8803) on the standard physiological markers of skin health of cutaneous arterial sympathetic nerve activity (CASNA), cutaneous blood flow and transepidermal water loss (TEWL) and to determine whether SBC8803 targets serotonin 5-HT3 receptors in rats. METHODS AND RESULTS: A set of three experiments were conducted to examine the effects of SBC8803 on CASNA, cutaneous blood flow and TEWL using Wistar and hairless rats. Two additional experiments further attempted to determine whether HK-SBC8803 was targeting the serotonin 5-HT3 receptors by pretreatment with the 5-HT3 antagonist granisetron. Administration of HK-SBC8803 in the first three experiments caused marked inhibition of CASNA and significant elevation of cutaneous blood flow under urethane anaesthesia as well as significant decrease in TEWL on the dorsal skin of conscious hairless rats. Pretreatment with granisetron decreased the effects of HK-SBC8803 on CASNA and cutaneous blood flow. CONCLUSIONS: These findings suggest that HK-SBC8803 reduces CASNA, increases cutaneous blood flow and decreases TEWL and that 5-HT3 receptors may be involved in CASNA and cutaneous blood flow responses. SIGNIFICANCE AND IMPACT OF THE STUDY: HK-SBC8803 could be a useful substance in the treatment/prevention of skin problems, specifically chapped or dry skin.

Oral administration of heat-killed Lactobacillus brevis SBC8803 elevates the ratio of acyl/des-acyl ghrelin in blood and increases short-term food intake.

It is known that gastrointestinal microbiota, probiotics and heat-killed microbes can regulate intestinal immunity; however, their effect on the secretion of gastrointestinal hormones is unclear. The secretion of gastrointestinal hormones can be mediated by the elevation of intracellular Ca2+ concentration, suggesting that these hormones may act through common mechanisms. We have previously shown that heat-killed Lactobacillus brevis SBC8803 (hk-SBC8803) induced the secretion of serotonin and elevated intracellular Ca2+ concentration in serotonin-producing RIN-14B cells, suggesting that hk-SBC8803 could potentially cause the same effect on other gastrointestinal hormones, including hunger hormone ghrelin. Here, we tested this hypothesis by treating cultured cells and experimental animals with hk-SBC8803 and assessing ghrelin secretion, expression of ghrelin-related genes, and food intake. The results indicated that hk-SBC8803 treatment for 30 min significantly upregulated the secretion of acyl ghrelin (active form) (P=0.046) and mRNA expression of the Syt3 (synaptotagmin 3) gene related to ghrelin exocytosis (P<0.05) in primary mouse stomach cells. In addition, oral administration of 500 mg/kg hk-SBC8803 to rats tended to upregulate acyl ghrelin concentration (P=0.10) and significantly increased the ratio of acyl to des-acyl (inactive) ghrelin (P=0.027) in blood, which corresponded to a tendency of stimulating food intake (P=0.087) at 30 min post-treatment. However, when in order to minimise individual differences we normalised the data on food intake to those on one-day food intake prior to food deprivation, the resultant food intake ratio showed a significant increase (by 5% compared to control; P=0.032) at 30 min after hk-SBC8803 treatment, indicating that hk-SBC8803 administration stimulated rats to take more food during the first meal after fasting. These results suggest that hk-SBC8803 induces short-term ghrelin secretion and transiently increases appetite, which is an important effect for individuals with low energy intake.

Effect of dietary heat-killed Lactobacillus brevis SBC8803 (SBL88™) on sleep: a non-randomised, double blind, placebo-controlled, and crossover pilot study.

We previously reported that dietary heat-killed Lactobacillus brevis SBC8803 affects sleep rhythms in mice. The present study evaluated the effect of consumption of heat-killed SBC8803 on sleep architecture in humans. A non-randomised, placebo-controlled, double blind, and crossover pilot study was conducted using volunteers who scored at a slightly high level (i.e. ≥6) on the Athens Insomnia Scale (AIS). Male subjects (n=17; age 41-69 y) consumed placebo or SBC8803 capsules (25 mg/day of heat-killed SBC8803) for 10 days. Electroencephalograms (EEG) were recorded using a mobile, one-channel system, providing objective data on sleep. Subjects' sleep journals and administration of the AIS provided subjective data on sleep. Three subjects were excluded from the statistical analysis. Analysis of the remaining 14 volunteers revealed no significant differences between placebo and SBC8803 consumption in either the AIS or the sleep EEG. The sleep journals revealed an improvement in 'waking' for the SBC8803 consumption periods (P=0.047), and there was a marginally significant effect on 'drowsiness during the following day' (P=0.067). Effects on the EEG delta power value (µV(2)/min) were revealed by a stratified analysis based on age, AIS, and the Beck Depression Inventory (BDI). Specifically, effects were found among subjects in their 40s who consumed the SBC8803 capsules (P=0.049) and among subjects with a BDI score less than the all-subjects average (13.3) (P=0.045). A marginally significant effect was found among subjects with an AIS score less than the all-subjects average (11.6) (P=0.065). The delta power value of 5 subjects with both BDI and AIS scores less than the average increased significantly (P=0.017). While the number of subjects was limited, a beneficial effect on sleep due to consumption of heat-killed L. brevis SBC8803 was found in subjects with slightly challenged sleep.

The serotonin receptor mediates changes in autonomic neurotransmission and gastrointestinal transit induced by heat-killed Lactobacillus brevis SBC8803.

Lactobacilli exhibit several health benefits in mammals, including humans. Our previous reports established that heat-killed Lactobacillus brevis SBC8803 (SBC8803) increased both efferent gastric vagal nerve activity and afferent intestinal vagal nerve activity in rats. We speculated that this strain could be useful for the treatment of gastrointestinal (GI) disorders. In this study, we examined the effects of SBC8803 on peristalsis and the activity of the efferent celiac vagal nerve innervating the intestine in rats. First, we examined the effects of intraduodenal (ID) administration of SBC8803 on efferent celiac vagal nerve activity (efferent CVNA) in urethane-anesthetised rats using electrophysiological studies. The effects of intravenous injection of the serotonin 5-HT3 receptor antagonist granisetron on changes in efferent CVNA due to ID administration of SBC8803 were also investigated. Finally, the effects of oral gavage of SBC8803 on GI transit were analysed using the charcoal propulsion method in conscious rats treated with or without granisetron. ID administration of SBC8803 increased efferent CVNA. Pretreatment with granisetron eliminated SBC8803-dependent changes in efferent CVNA. Furthermore, oral gavage of SBC8803 significantly accelerated GI transit, while pretreatment with granisetron inhibited GI transit. Our findings suggested that SBC8803 increased efferent CVNA and GI transit of charcoal meal via 5-HT3 receptors. Moreover, SBC8803 enhanced the activity of efferent vagal nerve innervating the intestine and promoted peristalsis via 5-HT3 receptors.

Chrymutasins: novel-aglycone antitumor antibiotics from a mutant of Streptomyces chartreusis. I. Taxonomy, mutation, fermentation, isolation and biological activities.

Three novel antibiotics, named chrymutasins A, B and C, were isolated from the fermentation products of a mutant strain obtained by NTG (N-methyl-N'-nitro-N-nitrosoguanidine) treatment. The mutant strain produced the chrymutasins, which differed in the aglycone moiety from chartreusin, and related compounds. The production of these compounds needed a characteristically long fermentation period. The antitumor activity of chrymutasin A is better in vivo than that of chartreusin, the cytotoxic activity against cell lines in vitro is equivalent, and the antimicrobial spectrum is narrower.

Chrymutasins: novel-aglycone antitumor antibiotics from a mutant of Streptomyces chartreusis. II. Characterization and structural elucidation.

Chrymutasins A, B and C are glycosidic antibiotics produced by a mutant of the chartreusin producer-organism Streptomyces chartreusis. We report here the structure elucidation of these compounds. The sugar moieties involved were determined by comparison with the related chartreusins. The structure of the aglycone, the same in all three compounds, was elucidated by NMR, incorporation studies of labeled compounds and synthesis of derivatives. The chrymutasin aglycone differs from that of chartreusin by a single carbon and an amino group.

Novel antitumor antibiotics, saptomycins. I. Taxonomy of the producing organism, fermentation, HPLC analysis and biological activities.

Streptomyces sp. HP530 was found to produce novel antitumor antibiotics, saptomycins, closely related to the pluramycin-group and was further found to mutate frequently. The natural mutant produced several new saptomycins as determined by HPLC analyses. We isolated saptomycins A, B, C1, C2 and F from the parent strain and saptomycins D, E, G and H from the mutant. The saptomycins showed antimicrobial activities and potent antitumor activities against human or murine tumor cell lines in vitro and against Meth A fibrosarcoma in vivo. In particular, saptomycin D was most effective component in vivo of all saptomycins.

Novel antitumor antibiotics, saptomycins. II. Isolation, physico-chemical properties and structure elucidation.

A complex of novel antitumor antibiotics related to the pluramycin-group was isolated from the fermentation of actinomycete, named Streptomyces sp. HP530. The producing strain mutated frequently. The products isolated from the parent strain were designated saptomycins A, B, C1, C2 and F, while those of the mutant were named saptomycins D, E, G and H. These structures were elucidated by several NMR spectral analyses and other spectroscopic experiments.

Novel cytocidal compounds, oxopropalines from Streptomyces sp. G324 producing lavendamycin. I. Taxonomy of the producing organism, fermentation, isolation and biological activities.

In the course of our investigation aimed at the discovery of novel antitumor antibiotics from microorganisms, Streptomyces sp. G324 was found to produce the antitumor antibiotic, lavendamycin, and also, to yield the novel beta-carboline compounds, oxopropalines. We isolated five compounds as oxopropalines A, B, D, E and G. Oxopropalines B, D and G showed cytocidal activities against human or murine tumor cell lines in vitro.

Novel cytocidal compounds, oxopropalines from Streptomyces sp. G324 producing lavendamycin. II. Physico-chemical properties and structure elucidations.

Novel cytocidal compounds designated oxopropalines A, B, D, E and G were isolated from the fermentation of an actinomycete named Streptomyces sp. G324, a strain that also produced an antitumor antibiotic, lavendamycin. All these compounds possessed a beta-carboline chromophore. The structures of the oxopropalines were elucidated by several NMR spectral analyses and other spectroscopic experiments.

Enzymatic formation of glidobactamine: a peptide nucleus of glidobactins A, B and C, new lipopeptide antitumor antibiotics.

Glidobactin deacylating activity was found in a bacterial strain of Pseudomonas sp. Glidobactamine, a key intermediate for acyl analogues of glidobactin, was isolated from the enzymatic degradation products of glidobactins after treatment using a column of fibrous active gel on which the cells of the Pseudomonas strain were immobilized. The chemical structure of glidobactamine was confirmed as the intact peptide moiety of glidobactins by chemical reformation of glidobactin A from glidobactamine and 2,4-dodecadienoic acid which is the constitutive fatty acid of glidobactin A.

A new rapid technique for detection of microorganisms using bioluminescence and fluorescence microscope method.

The MicroStar-RMDS-SPS (RMDS; R apid M icrobe D etection S ystem) enables detection and determination of the number of microorganisms trapped on a membrane filter based on the ATP-bioluminescence method. However, this system could not provide information about the identity of contaminating species based on the measurement results. Therefore, we developed a new technique for observing microcolonies of contaminants using a fluorescence microscope (Micro Scanner). The coordinates in the image data of the RMDS were converted to the coordinates of the sample stage of the microscope, and a microcolony could be easily observed. A single yeast cell and a microcolony consisting of 20-30 lactic acid bacterial cells could be observed on a membrane filter after measurement using the RMDS.

Classification and identification of strains of Lactobacillus brevis based on electrophoretic characterization of D-lactate dehydrogenase: relationship between D-lactate dehydrogenase and beer-spoilage ability.

The cell-free extracts of 60 strains which were identified phenotypically as being those of Lactobacillus brevis, including 48 isolates from the environment and 12 reference strains, were applied to polyacrylamide gel electrophoresis for extracting their NAD-dependent D- and L-lactate dehydrogenases (LDH). These strains were divided into 5 groups, i.e., Groups A, B, C, D, and E, on the basis of the electrophoretic mobilities of their D-LDH. The strains showed variations in their carbohydrate fermentation patterns. No relationship between the profile of D-LDH and the carbohydrate fermentation pattern was recognized. However, there appeared to be a relationship between the D-LDH profile and the beer-spoilage ability, because 40 out of 44 beer-spoilage strains identified as L. brevis were classified to Group B. We purified D-LDHs from the so-called complete beer-spoilage strain SBC 8002 of LDH Group B and from the non beer-spoilage strains JCM 1059T of LDH Group A and AHU 1508 of LDH Group C. Although the purified D-LDHs had the same molecular weight (84 kDa), each possessed a different optimum pH, optimum temperature, and isoelectric point. The aforementioned parameter values for the enzyme from the so-called complete beer-spoilage strain SBC 8002 of LDH Group B were 10.0, 50 degrees C, and 4.1, respectively; this strain was discriminated from the D-LDHs of the other two non beer-spoilage strains especially by its optimum temperature (50 degrees C).

Application of a bioluminescence method for the beer industry: sensitivity of MicroStar-RMDS for detecting beer-spoilage bacteria. Rapid Microbe Detection System.

We set up the original operating conditions of the MicroStar-Rapid Microbe Detection System (RMDS) to suppress false positives, which have kept this system from practical. The detection limit of our system was between 6.3 x 10(-16) mol and 3.1 x 10(-16) mol in terms of the amount of ATP, which is approximately equal to the ATP content of one yeast cell or 50 lactic acid bacteria cells. The detection time and the detection count were compared between the RMD method and the conventional plate count method (C.P.C. method) using 23 test samples of beer-spoilage Lactobacillus brevis. Judging from the detection time and detection count, 16-24 hours of cultivation for the RMD method corresponded to 40-96 hours of cultivation for the C.P.C. method. The RMD method reached a useful level for our practical use at the point of sensitivity.