Safety evaluation of the excessive intake of Bacillus subtilis C-3102 in healthy Japanese adults: A randomized, placebo-controlled, double-blind, parallel-group, comparison trial.

OBJECTIVE: Continuous intake of Bacillus subtilis C-3102 (B. subtilis C-3102) has been reported to modulate the gut microbiota and increase the bone mineral density of the femur in healthy adults. This study aimed to evaluate the safety of excessive B. subtilis C-3102 intake through a randomized, placebo-controlled, double-blind, parallel-group study. METHOD: A total of 69 individuals provided an informed consent, and 44 subjects who met the inclusion criteria were allocated to either the B. subtilis C-3102 (C-3102 group, n = 22) or the placebo group (P group, n = 22). All subjects took 18 tablets containing either containing B. subtilis C-3102 or placebo per day for 4 weeks with water and without chewing. Subjects in the C-3102 group consumed 4.8 × 1010 colony forming units (cfu) per day. Physical examination, urinalysis, blood analysis, records of subjective symptoms, and a medical questionnaire administered by a clinical trial physician were performed to determine the safety of test tablets. Furthermore, bone mineral density was measured. RESULTS: The final analysis included data from 22 subjects (9 men, 13 women; age, 46.1 ±â€¯13.8 years) in the C-3102 group and 22 subjects (9 men, 13 women; age, 46.1 ±â€¯13.5 years) in the P group. The results revealed no medical-related problems in both C-3102 and P groups. CONCLUSION: This study proved the safety of 4-week continuous consumption of an excessive amount of B. subtilis C-3102 tablets.

Morphological analysis on the distribution of membrane lipids and a membrane protein, NAP-22, during neuronal development in vitro.

Specific localization of membrane proteins based on the interactions with membrane lipids at various microdomains (MDs) is under active investigation, since the elucidation of the molecular mechanism of the interactions could reveal a novel concept of cell organization. Due to the strong interactions not only between lipids but also between lipids and proteins, these MDs are considered to be recovered in a detergent-resistant low-density membrane fraction (DRM) after detergent extraction and density-gradient centrifugation. Neurons take well-developed membrane systems during maturation and specific localization of various membrane components, not only proteins but also lipids, is essential for the establishment of the nervous system. In previous studies, we showed that NAP-22 is a major protein of neuronal DRM and binds liposomes in a cholesterol-dependent manner. In this study, we analyzed the localization of membrane lipids during neuronal maturation in vitro and compared their distribution with that of NAP-22. In an attempt to detect DRM-associated lipids, we observed the staining patterns of neurons treated with Triton-X-100 at 4 degrees C before fixation. Our results showed the less staining patterns of cholesterol and sphingomyelin at the axonal tips and a different staining pattern of two gangliosides, GM(1) and GD(3). The enrichment of cholesterol at the NAP-22 localizing spots was observed after the treatment of the detergent. Since the application of maitotoxin, a calcium ion channel, caused the diminution of NAP-22 and cholesterol positive spots, the distribution of these molecules are considered under the calcium regulation.

Immunoglobulin E-independent activation of mast cell is mediated by Mrg receptors.

Mast cells play a central role in inflammatory and allergic reactions by releasing inflammatory mediators through two main pathways, immunoglobulin E-dependent and -independent activation. In the latter, mast cells are activated by a diverse range of basic molecules, including peptides and amines such as substance P, neuropeptide Y, and compound 48/80. These secretagogues are thought to activate the G proteins in mast cells through a receptor-independent mechanism. Here, we report that the basic molecules activate G proteins through the Mas-related gene (Mrg) receptors on mast cells, leading to mast cell degranulation. We suggest that one of the Mrg receptors, MrgX2, has an important role in regulating inflammatory responses to non-immunological activation of human mast cells.

Molecular basis of non-self recognition by the horseshoe crab tachylectins.

The self/non-self discrimination by innate immunity through simple ligands universally expressed both on pathogens and hosts, such as monosaccharides and acetyl group, depends on the density or clustering patterns of the ligands. The specific recognition by the horseshoe crab tachylectins with a propeller-like fold or a propeller-like oligomeric arrangement is reinforced by the short distance between the individual binding sites that interact with pathogen-associated molecular patterns (PAMPs). There is virtually no conformational change in the main or side chains of tachylectins upon binding with the ligands. This low structural flexibility of the propeller structures must be very important for specific interaction with PAMPs. Mammalian lectins, such as mannose-binding lectin and ficolins, trigger complement activation through the lectin pathway in the form of opsonins. However, tachylectins have no effector collagenous domains and no lectin-associated serine proteases found in the mammalian lectins. Furthermore, no complement-like proteins have been found in horseshoe crabs, except for alpha(2)-macroglobulin. The mystery of the molecular mechanism of the scavenging pathway of pathogens in horseshoe crabs remains to be solved.

Molecular basis of non-self recognition by the horseshoe crab lectins.

The target molecules of innate immunity are not proteins of direct gene products but the molecular arrays or patterns on pathogens, called pathogen-associated molecular patterns (PAMPs). The self/non-self discrimination by innate immunity through simple ligands universally expressed both on pathogens and hosts, such as monosaccharides and the acetyl group, probably depends on the density or clustering patterns of the ligands. The specific recognition by the horseshoe crab lectins, named tachylectins, with a propeller-like fold or a propeller-like oligomeric arrangement is reinforced by the short distance between the individual binding sites that interact with PAMPs. There is virtually no conformational change in the main or side chains of tachylectins upon binding with the ligands. This low structural flexibility of the propeller structures must be very important for specific interaction with PAMPs.