Carnosol Modulates Th17 Cell Differentiation and Microglial Switch in Experimental Autoimmune Encephalomyelitis.

Medicinal plants as a rich pool for developing novel small molecule therapeutic medicine have been used for thousands of years. Carnosol as a bioactive diterpene compound originated from Rosmarinus officinalis (Rosemary) and Salvia officinalis, herbs extensively applied in traditional medicine for the treatment of multiple autoimmune diseases (1). In this study, we investigated the therapeutic effects and molecule mechanism of carnosol in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). Carnosol treatment significantly alleviated clinical development in the myelin oligodendrocyte glycoprotein (MOG35-55) peptide-induced EAE model, markedly decreased inflammatory cell infiltration into the central nervous system and reduced demyelination. Further, carnosol inhibited Th17 cell differentiation and signal transducer and activator of transcription 3 phosphorylation, and blocked transcription factor NF-κB nuclear translocation. In the passive-EAE model, carnosol treatment also significantly prevented Th17 cell pathogenicity. Moreover, carnosol exerted its therapeutic effects in the chronic stage of EAE, and, remarkably, switched the phenotypes of infiltrated macrophage/microglia. Taken together, our results show that carnosol has enormous potential for development as a therapeutic agent for autoimmune diseases such as MS.

Nutshell Extracts of Xanthoceras sorbifolia: A New Potential Source of Bioactive Phenolic Compounds as a Natural Antioxidant and Immunomodulator.

The nutshell of Xanthoceras sorbifolia, a waste product in the production of edible oil, is rich in health-promoting phenolic acids. However, the individual constituents, bioactivities, and mechanism of action are largely unknown. In this study, 20 phenolic compounds were characterized in nutshell extract (NE) of X. sorbifolia by gas chromatography-mass spectrometry. Four established in vitro studies showed that NE has significant antioxidant potential. Results in vivo indicated that oral administration of NE effectively ameliorated clinical disease severity of experimental autoimmune encephalomyelitis (EAE) and reduced the neuroinflammation and the central nervous system (CNS) demyelination. The underlying mechanism of NE-induced effects involved decreased penetration of pathogenic immunocyte into the CNS, a reduced production of proinflammatory cytokines and factors, and suppressed differentiation of type 1 T helper and type 17 T helper cells through the Janus kinase/signal transducer and activator of transcription pathway. Taken together, our studies showed that X. sorbifolia nutshell, considered a waste material in the food industry, is a novel source of a natural antioxidant and immunomodulator.

miR-23b Suppresses Leukocyte Migration and Pathogenesis of Experimental Autoimmune Encephalomyelitis by Targeting CCL7.

MicroRNAs (miRNAs) are small, non-coding RNAs involved in immune response regulation. Specific miRNAs have been linked to the development of various autoimmune diseases; however, their contribution to the modulation of CNS-directed cellular infiltration remains unclear. In this study, we found that miR-23b, in addition to its reported functions in the suppression of IL-17-associated autoimmune inflammation, halted the progression of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS), by directly inhibiting the migration of pathogenic leukocytes to the CNS. We demonstrated that miR-23b was specifically decreased during the acute phase of EAE and that overexpression of miR-23b resulted in a defect in leukocyte migration and strong resistance to EAE. Furthermore, we found that miR-23b suppressed leukocyte migration of EAE by targeting CCL7, a chemokine that attracts monocytes during inflammation and metastasis. Finally, in the adoptive transfer model, miR-23b reduced the severity of EAE by inhibiting the migration of pathogenic T cells to the CNS rather than diminishing the encephalitogenesis of T cells. Taken together, our results characterize a novel aspect of miR-23b function in leukocyte migration, and they identify miR-23b as a potential therapeutic target in the amelioration of MS and likely other autoimmune diseases.

Effects of total flavonoids from Drynariae Rhizoma prevent bone loss in vivo and in vitro.

Estrogen deficiency is one of the major causes of osteoporosis in postmenopausal women. Drynariae Rhizoma is a widely used traditional Chinese medicine for the treatment of bone diseases. In this study, we investigated the therapeutic effects of the total Drynariae Rhizoma flavonoids (DRTF) on estrogen deficiency-induced bone loss using an ovariectomized rat model and osteoblast-like MC3T3-E1 cells. Our results indicated that DRTF produced osteo-protective effects on the ovariectomized rats in terms of bone loss reduction, including decreased levels of bone turnover markers, enhanced biomechanical femur strength and trabecular bone microarchitecture deterioration prevention. In vitro experiments revealed that the actions of DRTF on regulating osteoblastic activities were mediated by the estrogen receptor (ER) dependent pathway. Our data also demonstrated that DRTF inhibited osteoclastogenesis via up-regulating osteoprotegrin (OPG), as well as down-regulating receptor activator of NF-κB ligand (RANKL) expression. In conclusion, this study indicated that DRTF treatment effectively suppressed bone mass loss in an ovariectomized rat model, and in vitro evidence suggested that the effects were exerted through actions on both osteoblasts and osteoclasts.

The complete chloroplast genome sequence of medicinal plant Pinellia ternata.

Pinellia ternata is an important medicinal plant used in the treatment of cough, to dispel phlegm, to calm vomiting and to terminate early pregnancy, as an anti-ulcer and anti-tumor medicine. In this study, we found that the complete chloroplast genome of Pinellia ternata was 164 013 bp in length, containing a pair of inverted repeats of 25 625 bp separated by a large single-copy region and a small single-copy region of 89 783 bp and 22 980 bp, respectively. The chloroplast genome encodes 132 predicted functional genes, including 87 protein-coding genes, eight ribosomal RNA genes, and 37 transfer RNA genes. The chloroplast DNA is GC-rich (36.7%). The phylogenetic analysis showed a strong sister relationship with Colocasia esculenta, which also strongly supports the position of Pinellia ternata. The complete chloroplast genome sequence of Pinellia ternata reported here has the potential to advance population and phylogenetic studies of this medicinal plant.

Transcriptome and proteome of the highly neurotoxic venom of Gloydius intermedius.

The venomics of Gloydius intermedius were investigated using expressed sequence tags (ESTs) analyses, 2D gel-electrophoresis combined with MALDI-TOF/TOF, and LC-MS/MS. A total of 1920 ESTs from the venom gland cDNA library were sequenced; 74% of them belonged to toxin-families. The four most abundant families among the toxin transcripts were: serine protease (SP, 36.2%), bradykinin potentiating peptide (25.3%), l-amino acid oxidase (LAAO, 13.1%), and phospholipase A2 (PLA2, 9.9%). Moreover, the full sequences of four PLA2s, eight SPs, cysteine-rich secretory protein (CRISP), C-type-lectin-like-protein (CTLP), hyaluronidase, metalloproteinase, and nerve growth factor were deduced from the cDNA sequences. Excluding the CRISP and hyaluronidase, most of the G. intermedius venom proteins bear 92-99% sequence identities to those of other pitviper venoms. The most abundant components are PLA2s (37%), SPs (20%) and LAAO (6%), while metalloproteinase, CTLP, and other components each account for <3% of the total venom proteins. The abundance of Gintexin (a crotoxin-like neurotoxin) and low levels of hemorrhagic metalloproteases, disintegrins and CTLPs highlight the great venom differences between G. intermedius and other hemorrhagic pitvipers. The bimorphism of hemorrhagic and neurotoxic venoms among Gloydius is confirmed; our results shed more lights on the co-evolution of both neurotoxicity and hypotension in some viperid venoms.

Reversal effect of rosmarinic acid on multidrug resistance in SGC7901/Adr cell.

Multidrug resistance (MDR) has been a major problem in cancer chemotherapy. In this study, the aim was to explore the reversal effect and its potential mechanism of rosmarinic acid (RA) on SGC7901/Adr cells. 3-(4,5-Dimethylthiazol)-2,5-diphenyl tetrazolium bromide (MTT) assay was used to investigate the reversal index of RA in SGC7901/Adr cell line. The intracellular accumulation of adriamycin, rhodamine123 (Rh123), and the expression of P-glycoprotein (P-gp) were assayed by flow cytometry. The influence of RA on the transcription of MDR1 gene was determined by reverse transcription-polymerase chain reaction. The results showed that RA could reverse the MDR of SGC7901/Adr cells, increase the intracellular accumulation of Adr and Rh123, and decrease the transcription of MDR1 gene and the expression of P-gp in SGC7901/Adr cells. These results indicated that RA was a potential multidrug resistance-reversing agent and warranted further investigations.

[Analysis of essential oil from different organs of Caryopteris tangutica].

OBJECTIVE: To study the main chemical components of essential oils from Caryopteris tangutica. METHODS: The chemical compositions of essential oils obtained by hydrodistillation from different organs of Caryopteris tangutica were separately analyzed and identified by GC-MS and Kovats retention index. The relative contents of the components by the peak-area normalization method adopted in gas chromatography. RESULTS: 13.48 and 42 components were separately identified from stems, leaves and flowers. The components and relative contents of the leaves were similar with the flowers, and the main constituents were Myrtenal (2.73%, 1.69%), trans-Pinocarvyl acetate (46.69%, 55.48%), Myrtenyl acetate (1.17%, 1.42%), beta-Cedrene (1.62%, 3.21%), Caryophyllene oxide (1.67%, 2.73%) and so on. The components of the stems were different from others, and the main constituents were hexadecanoic acid (47.32%), trans-Pinocarvyl acetate (24.19%), phytol (0.77%). CONCLUSION: The components and relative contents of the essential oils among stems, leaves and flowers were distinctly different, and each of them had a great range of potential utilities and a prospect of development.

Comparative analysis of essential oil components of three Phlomis species in Qinling Mountains of China.

The essential oils of three wild-growing Phlomis species (Phlomis umbrosa Turcz., Phlomis megalantha Diels and Phlomis szechuanensis C.Y. Wu), collected from Qinling Mountains of China during the bloom stage, were obtained by hydrodistillation and analyzed by gas chromatography (GC) and gas chromatography/mass spectrometry (GC-MS). Under the optimum extraction and analysis conditions, 22, 26 and 19 constituents (mainly aliphatic compounds) were identified in P. umbrosa, P. megalantha and P. szechuanensis which represented 92.5%, 96.3% and 93.1% of the oils, respectively. The main constituents were hexadecanoic acid (7.1-52.1%), trans-phytol (5.7-50.8%) and 9,12,15-octadecatrien-1-ol (2.2-24.8%). Fatty acids and aliphatic esters were the major groups of P. umbrosa and P. megalantha, but P. szechuanensis showed higher content of alcohols. P. megalantha has relatively higher amounts of oxygenated monoterpenes and oxygenated sesquiterpenes than the others. The comparison of essential oil components of Phlomis species between the present and previous work indicated that the composition of oils vary greatly with respect to the geographical environment, mainly for the proportion of aliphatic compounds and terpenoids. This study is the first report on the chemical composition of essential oils of the three wild-growing herbs mentioned above.

Expression of human papillomavirus type 16 L1 protein in transgenic tobacco plants.

To develop a plant expression system for the production of the human papillomavirus type 16 (HPV16) vaccine, we investigated whether the HPV16 L1 protein can be expressed in tobacco plants and whether it can be used as the cheapest form of edible vaccine. The HPV16 L1 coding sequence was amplified by PCR using specific primers from the plasmid pGEM-T-HPV16 containing the template sequence, and subcloned into the intermediate vector pUCmT and binary vector pBI121 consecutively to obtain the plant expression plasmid pBI-L1. The T-DNA regions of the pBI-L1 binary vector contained the constitutive Cauliflower mosaic virus (CaMV) 35S promoter and the neomycin phosphotransferase npt II gene, which allowed the selection of transformed plants using kanamycin. The tobacco plants were transformed by co-cultivating them, using the leaf disc method, with Agrobacterium tumefaciens LBA4404, which harbored the plant expression plasmid. The regenerated transgenic tobacco plants were selected using kanamycin, and confirmed by PCR. The results of the Southern blot assay also showed that the HPV16 L1 gene was integrated stably into the genome of the transformed tobacco plants. The Western blot analysis showed that the transformed tobacco leaves could express the HPV16 L1 protein. Furthermore, it was demonstrated by ELISA assay that the expressed protein accounted for 0.034%-0.076% of the total soluble leaf protein, was able to form 55 nm virus-like particles compatible with HPV virus-like particle (VLP), and induced mouse erythrocyte hemagglutination in vitro. The present results indicate that the HPV16 L1 protein can be expressed in transgenic tobacco plants and the expressed protein possesses the natural features of the HPV16 L1 protein, implying that the HPV16 L1 transgenic plants can be potentially used as an edible vaccine.