This study was undertaken to evaluate whether exosomes from human adipose-derived stem cells (ASC-exo) can stimulate the regeneration of human dermal fibroblasts (HDFs). Immunoblotting and FACS analyses showed that ASC-exo was positive for exosome markers. Fluorescence tracking revealed that the contents of ASCexo were transferred into the HDFs. ASC-exo treatment also stimulated the proliferation and migration of HDFs in a dose-dependent manner. Similarly, the expression levels of genes involved in skin cell proliferation were increased by ASC-exo. Microarray analysis showed an enrichment of microRNAs that have regenerative function. We suggest that the ASC-exo can stimulate skin cell proliferation. | BACKGROUNDIt has been well reported that adipose tissue-derived stem cell (ASC) secretes various soluble factors, that is growth factors, cytokines, exosomes that can rescue damaged cells.[1] Exosome, a 30-to 150-nm-sized extracellular vesicle that is secreted from most cell types, is formed within endosomal compartments and released into the extracellular milieu.[2] It has been demonstrated that exosomes possess similar functional properties of mesenchymal stem cells (MSCs) from which they are derived, [3] and due to such functions, exosomes are regarded now as a communicator between tissues.[4] Functionally, exosomes can reduce the immune recognition, so that the integrity of cell membrane can be maintained. [5] A study of stem cell transplantation therapy demonstrated that the role of MSCs in cell-tocell communication was exerted through a paracrine mechanism and that exosomes play a major role in this process.[6] Other findings also described that the conditioned media of ASC (ASC-CM) [7] or exosomes [8] were able to promote the migration of dermal fibroblasts during the process of wound healing. [7] Although the mechanism of the exosomes' role on proliferation and migration of fibroblasts was demonstrated, [8] no study has been conducted to identify the expressional changes of microRNAs, which play major role in various cellular responses, including proliferation. | QUESTIONS ADDRESSEDWe asked whether ASC-derived exosomes can stimulate the skin dermal fibroblasts' proliferation and migration, and by what mechanism these exosomes play such functions. | EXPERIMENTAL DESIGNSee supplementary information. | RESULTSTransmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA) showed that ASC-exo isolated from the supernatants (Fig. S1A). Western blot analysis revealed that ASC-exo expresses exosome markers HSP70, CD63and CD9 (Fig. S1B), while these markers were absent in ASC lysate.Flow cytometry analysis also showed that ASC-exos were positive for CD9, CD63 and CD81 (Fig. S1C). We next asked whether the contents of ASC-exos could be transmitted to HDFs. After co-incubation, for 24 hours, of PKH26-labelled ASC-exos with HDFs, it was found that red fluorescence of PKH26 was localized within the cytoplasm of HDFs (Fig. S1D), showing that ASC-exo can be internalized intoHDFs. Also, quantitati...

Kefir is a fermented product from yeast and lactic acid bacteria, and has been associated with various health benefits including relieving inflammatory bowel disease. Recently, it has been shown that gram-positive bacteria produce extracellular vesicles (EV). The EV could be appearing as potentially important mediators of cell to cell interaction. In this study, we explored the role of kefir grain Lactobacillus-derived EV in modulating inflammation responses via alleviating the production of inflammatory cytokines in tumor necrosis factor-α (TNF-α)-induced inflammation in Caco-2 cells and the 2,4,6-trinitrobenzene sulfonic acid-induced inflammatory bowel disease mouse model. Kefir-derived Lactobacillus EV were isolated by ultracentrifugation of the culture medium of 3 different kefir-derived strains (i.e., Lactobacillus kefir, Lactobacillus kefiranofaciens, and Lactobacillus kefirgranum). Nanoparticle tracking analysis showed that the size of isolated kefir-derived Lactobacillus EV was within 80 to 400 nm, and kefir-derived Lactobacillus EV uptake into recipient Caco-2 cells was confirmed by fluorescence labeling. Treatment of each kefir-derived Lactobacillus EV onto TNF-α-stimulated Caco-2 cells significantly reduced the level of both mRNA expression and secretion of IL-8, and Western blot analysis revealed that such an effect was related to inhibition of TNF-α signaling mediated by reducing the phosphorylation of p65, a subunit of NF-kB. Subsequent administration of kefir-derived Lactobacillus EV into inflammatory bowel disease-induced mice significantly alleviated the body weight loss and rectal bleeding, and enhanced stool consistency. Histological examination showed that kefir-derived Lactobacillus EV substantially reduced the infiltration of transmural leukocytes and loss of goblet cells within the colon, and the serum level of myeloperoxidase was significantly lower in the EV-treated group than control group. Our study demonstrates that kefir-derived Lactobacillus EV can be potentially used for developing innovative strategies for alleviating inflammatory bowel disease.

Systemic lupus erythematosus (SLE) is a chronic multisystemic autoimmune disease with an unknown etiology. Recently, it has been elucidated that dysregulated histone deacetylase (HDAC) activity is related to the pathogenesis of inflammatory and autoimmune diseases. Broad-spectrum HDAC inhibitors are effective for the treatment of allergy, cancer, and autoimmune diseases, but they have several adverse side effects. Thus, the purpose of this study was to evaluate the effects of a novel HDAC 6-specific inhibitor, CKD-506, in a murine SLE model. CKD-506 significantly improved survival rate and significantly decreased the incidence of severe proteinuria, blood urea nitrogen, kidney inflammation, and glomerular infiltration of IgG and C3. In addition, CKD 506 reduced the proportions of CD138+ plasma cells, CD4−CD8− T cells, and CD25+ cells and the Th1/Th2 ratio in the spleen. CKD-506 significantly reduced inflammatory cytokines such as IL-10, IL-15, IL-17, TNF-α, and IFN-inducible protein (IP-10) and significantly increased TGF-β in serum. CKD-506 also significantly reduced IFN-γ, IL-1β, IL-4, IL-6, IP-10, MCP-1, and CCL4 levels in kidney. CKD-506 decreased the production of various pro-inflammatory cytokines and chemokines in the serum and kidneys, resulting in inhibition of cell migration and suppression of lupus nephritis without adverse effects.

Calf diarrhea caused by infectious agents is associated with economic losses in the cattle industry. The purpose of this study was to identify the causative agents and epidemiological characteristics of diarrhea in Korean native calves (KNC). In total, 207 diarrheal KNC aged less than 7 months were investigated. Fecal samples collected from the rectum were examined for causative agents using polymerase chain reaction (PCR) or real-time PCR and the number of oocysts were counted. Fourteen causative agents were detected from 164 of the 207 diarrheal KNC. Rotavirus was the most common agent (34.8%), followed by Eimeria spp. (31.7%), Escherichia coli (22.0%), Giardia spp. (14.0%), Clostridium difficile (9.8%), bovine viral diarrhea virus (8.5%), coronavirus (7.9%), Cryptosporidium spp. (7.3%), torovirus (6.7%), parvovirus (5.5%), norovirus (4.9%), kobuvirus (1.8%), adenovirus (1.2%), and Salmonella spp. (0.6%). About 95 (57.9%) of 164 calves were infected with a single causative agent and 42.1% were infected by multiple agents. No significant difference was observed in mortality between calves infected with a single agent and multiple agents. The occurrence of diarrhea caused by rotavirus, Eimeria spp., kobuvirus, and Giardia spp. was significantly different based on onset age, and the prevalence of diarrhea caused by rotavirus or C. difficile was significantly different between seasons. This study help the understanding of KNC diarrhea for the development of an effective strategy for disease prevention and control, especially in Eastern provinces of South Korea.

Abbreviations: AMPK, AMP-activated protein kinase; HFD, high-fat diet; ND, normal diet; PPARγ, peroxisome proliferator-activated receptor γ; FAS, fatty acid synthase; ACC1, acetyl-CoA carboxylase 1; SCD1, stearyl-CoA desaturase 1; RXRα , retinoid X receptor α ; BADGE, bisphenol A diglycidyl ether; ADD1, adipocyte differentiation and determination factor 1; SREBP1c, sterol regulatory element binding protein 1c; ACO, acyl-CoA oxidase; CPT1, carnitine palmitoyltransferase 1; C/EBP, CCAAT/enhancer-binding protein; AICAR, aminoimidazole carboxamide ribonucleotide AbstractLysimachia foenum-graecum has been used as an oriental medicine with anti-inflammatory effect. The anti-obesity effect of L. foenum-graecum extract (LFE) was first discovered in our screening of natural product extract library against adipogenesis. To characterize its anti-obesity effects and to evaluate its potential as an anti-obesity drug, we performed various obesity-related experiments in vitro and in vivo. In adipogenesis assay, LFE blocked the differentiation of 3T3-L1 preadipocyte in a dose-dependent manner with an IC50 of 2.5 μg/ml. In addition, LFE suppressed the expression of lipogenic genes, while increasing the expression of lipolytic genes in vitro at 10 μ g/ml and in vivo at 100 mg/kg/day. The anti-adipogenic and anti-lipogenic effect of LFE seems to be mediated by the inhibition of PPARγ and C/EBPα expression as shown in in vitro and in vivo, and the suppression of PPARγ activity in vitro. Moreover, LFE stimulated fatty acid oxidation in an AMPK-dependent manner. In high-fat diet (HFD)-induced obese mice (n = 8/group), oral administration of LFE at 30, 100, and 300 mg/kg/day decreased total body weight gain significantly in all doses tested. No difference in food intake was observed between vehicle-and LFE-treated HFD mice. The weight of white adipose tissues including abdominal subcutaneous, epididymal, and perirenal adipose tissue was reduced markedly in LFE-treated HFD mice in a dose-dependent manner. Treatment of LFE also greatly improved serum levels of obesity-related biomarkers such as glucose, triglycerides, and adipocytokines leptin, adiponectin, and resistin. All together, these results showed anti-obesity effects of LFE on adipogenesis and lipid metabolism in vitro and in vivo and raised a possibility of developing LFE as anti-obesity therapeutics.