Oral administration of Lactobacillus casei variety rhamnosus partially alleviates TMA-induced atopic dermatitis in mice through improving intestinal microbiota.

AIMS: The purpose of this study was to investigate the effect of Lactobacillus casei variety rhamnosus (LCR35) on Atopic dermatitis (AD)-like symptoms in mice. METHODS AND RESULTS: AD-like skin lesions in BALB/C mice were induced by sensitization and subsequent repeated challenges with trimellitic anhydride (TMA) for 10 days. LCR35 was orally administered to the mice once daily throughout the study. In the TMA-induced AD model, orally administered LCR35 suppressed significantly irritant-related scratching behaviour and skin dehydration as well as apparent severity of AD. LCR35 also significantly decreased serum levels of IgE and IL-4, but not IFN-γ, implying the restoration of TMA-induced disruption of Th1/Th2 balance. Quantitative real-time PCR targeting hypervariable regions of 16S rDNA gene of faecal microbiota indicated that the LCR35 treatment increased the population of Bifidobacterium, Lactobacilli, Enterococcus and Bacteroides fragilis group, but decreased those of Clostridium coccoides group. CONCLUSIONS: LCR35 has the ability to suppress the development of AD in mice, possibly through the modulation of Th1/Th2 balance and gut microbiota. SIGNIFICANCE AND IMPACT OF THE STUDY: LCR35 has a strong potential as a probiotic for preventing AD.

Asarone inhibits adipogenesis and stimulates lipolysis in 3T3-L1 adipocytes.

Asarone is a molecule found in certain plants such as Acorus calamus, the root of which is used in traditional medicine to treat diabetes. We determined the molecular mechanism underlying the anti-diabetic activity of asarone. Treatment of asarone significantly inhibited the differentiation of 3T3-L1 preadipocytes through suppression of expression of the transcription factors, CCAAT/enhancer binding protein-alpha and peroxisome proliferator activated receptor-gamma, which activate adipogenesis. Intracellular triglyceride levels were reduced by asarone in a dose-dependent manner and asarone treatment stimulated the phosphorylation of hormone-sensitive lipase. Together, the present findings indicate that asarone inhibits adipogenesis by down-regulation of PPARgamma and C/EBPalpha and reduces lipid accumulation by stimulation of lipolysis through an increase in hormone-sensitive lipase activity.

Wound-healing activity of Astragali Radix in rats.

Inhibition of inflammatory responses, acceleration of basal cell growth and balanced synthesis of the extracellular matrix (ECM) are important in the healing of open cutaneous wounds. To evaluate the wound-healing effects of Astragali Radix (AR) (the root of Astragalus membranaceus [Fisch.]), experimental open wounds were made on the dorsal side of rats under anesthesia. Boiling water extracts of AR, soaked into a hydrophilic foam dressing, were topically applied to the wounds once a day for 11 consecutive days. The healing process was assessed by scoring macroscopic appearance and measuring the area of the open wounds. Molecular aspects of the healing skin area were also investigated via histological observation indicating cell density and linear alignment of the granulation tissue. The AR extracts significantly accelerated cutaneous wound healing by suppressing inflammation and stimulating basal cell growth in the wound area compared to epidermal growth factor as a positive control. Promotion of basal cell proliferation and angiogenesis by the AR extracts was remarkable in the early stages of wound healing, resulting in a significant reduction in the duration of the wound-healing process. We conclude that extracts of AR could be useful in enhancing cutaneous wound healing.

Human neutrophil lactoferrin trans-activates the matrix metalloproteinase 1 gene through stress-activated MAPK signaling modules.

It has been proposed that human neutrophil lactoferrin (Lf) could be involved in gene expression as a DNA-binding protein after its translocation into the nucleus. However, the molecular basis of Lf action has not been defined, and Lf-regulated target genes have not been identified. We report here that overexpressed Lf functions as a specific trans-activator of matrix metalloproteinase 1 (MMP1) gene, and that induction of this AP-1-responsive gene is mediated via the stress-activated MAPK signaling modules. Transactivation of the MMP1 promoter by overexpressed Lf requires the presence of an AP-1 binding site. In gel shift experiments, Lf did not interact directly with AP-1-containing fragments of the MMP1 promoter. However, nuclear extracts from Lf-expressing cells contained increased levels of proteins that bound to AP-1 elements. This Lf-induced AP-1 DNA binding activity was reduced by a p38 MAPK inhibitor. Inhibitors of the MEK kinases had little effect on Lf-induced AP-1. However, expression of dominant-negative MKK4 or JNK1 inhibited Lf-induced gene expression. The JNK activity stimulated by Lf correlates with the enhanced AP-1 binding ability. These findings demonstrate that the Lf-induced activation of AP-1 is mediated via JNK and p38 MAPK pathways.

The yiaE gene, located at 80.1 minutes on the Escherichia coli chromosome, encodes a 2-ketoaldonate reductase.

An open reading frame located in the bisC-cspA intergenic region, or at 80.1 min on the Escherichia coli chromosome, encodes a hypothetical 2-hydroxyacid dehydrogenase, which was identified as a result of the E. coli Genome Sequencing Project. We report here that the product of the gene (yiaE) is a 2-ketoaldonate reductase (2KR). The gene was cloned and expressed with a C-terminal His tag in E. coli, and the protein was purified by metal-chelate affinity chromatography. The determination of the NH2-terminal amino acid sequence of the protein defined the translational start site of this gene. The enzyme was found to be a 2KR catalyzing the reduction of 2, 5-diketo-D-gluconate to 5-keto-D-gluconate, 2-keto-D-gluconate (2KDG) to D-gluconate, 2-keto-L-gulonate to L-idonate. The reductase was optimally active at pH 7.5, with NADPH as a preferred electron donor. The deduced amino acid sequence showed 69.4% identity with that of 2KR from Erwinia herbicola. Disruption of this gene on the chromosome resulted in the loss of 2KR activity in E. coli. E. coli W3110 was found to grow on 2KDG, whereas the mutant deficient in 2KR activity was unable to grow on 2KDG as the carbon source, suggesting that 2KR is responsible for the catabolism of 2KDG in E. coli and the diminishment of produced 2KDG from D-gluconate in the cultivation of E. coli harboring a cloned gluconate dehydrogenase gene.

Characterization and evaluation of a pta (phosphotransacetylase) negative mutant of Escherichia coli HB101 as production host of foreign lipase.

In order to evaluate the pta(phosphotransacetylase) (-) mutant of Escherichia coli as a potential host of foreign lipase expression, the pta(-) mutant HB101 was constructed for the purpose of blocking the acetate synthetic pathway. Since acetate is known as a major inhibitory by-product of cell growth and foreign protein production, the growth characteristics and expression kinetics of the microbial lipase of the pta(-) E. coli mutant were investigated. The growth rate was considerably decreased (about 30%) when grown on M9 minimal media containing glucose, mannose or glycerol. Growth retardation was not observed when a gluconeogenic carbon source (acetate, malate or succinate) was utilized. It should be noted that the growth rate of the mutant was enhanced (about 20%) in modified M9 media including a gluconeogenic carbon source and NZ-amine. Growth inhibition of the pta(-) mutant by menadione, a representative redox-cycling drug, was more pronounced than that of the parental type of E. coli. Furthermore, the inhibition effect was more pronounced in glucose minimal medium, whereas the menadione sensitivity was not observed when a gluconeogenic carbon source was used as a sole carbon source or the lactate dehydrogenase gene from Lactobacillus casei was introduced in the pta(-) mutant. Therefore, it is suggested that the growth deficiency of the pta(-) mutant is closely related to the intracellular redox balance. When the pseudomonad lipase was expressed in the pta(-) mutant, a comparable expression rate and yield to the parental type strain was observed. High-cell-density culture of the mutant was easy to achieve even under the fluctuating conditions of residual glucose concentration.