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Somatic cell nuclear transfer (SCNT) is used widely in cloning, stem cell research, and regenerative medicine. The type of donor cells is a key factor affecting the SCNT efficiency.This study examined whether urine-derived somatic cells could be used as donors for SCNT in pigs.The viability of cells isolated from urine was assessed using trypan blue and propidium iodide staining. The H3K9me3/H3K27me3 level of the cells was analyzed by immunofluorescence. TheMost sow urine-derived cells were viable and could be cultured and propagated easily. On the other hand, most of the somatic cells isolated from the boar urine exhibited poor cellular activity. TheSow urine-derived cells could be used to produce SCNT embryos.

Shenlian extract (SL), extracted from Salvia miltiorrhiza Bunge and Andrographis paniculata (Burm. f.) Nees, has been proved to be effective in the prevention and treatment of atherosclerosis. Recently, we have partially elucidated the mechanisms involved in the therapeutic effects of SL on myocardial ischemia (MI). However, the underlying mechanisms remain largely unclear.This study aims to explore the potential molecular mechanism of SL on MI on the basis of network pharmacology.First, the main active ingredients of SL were screened in the Traditional Chinese Medicine Integrated Database, and the MI-associated targets were collected from the DisGeNET database. Then, we used compound-target and target-pathway networks to uncover the therapeutic mechanisms of SL. On the basis of network pharmacology analysis results, we assessed the effects of SL in MI rat model and oxygen glucose deprivation model of H9c2 cells and validated the possible molecular mechanisms of SL on myocardial injury in vivo and in vitro.The network pharmacology results showed that 37 potential targets were recognized, including TNF-α, Bcl-2, STAT3, PI3K and MMP2. These results revealed that the possible targets of SL were involved in the regulation of inflammation and apoptosis signaling pathway. Then, in vivo experiments indicated that SL significantly reduced the myocardial infarction size of MI rats. Serum CK-MB, cTnT, CK, LDH, and AST levels were significantly decreased by SL (P 0.05 or P 0.01). In vitro, SL significantly increased H9c2 cell viability. The levels of inflammation factors including TNF-α and MMP2 were significantly decreased by SL (P 0.05 or P 0.01). TUNEL and Annexin V/propidium iodide assays indicated that SL could significantly decrease the cell apoptotic rate in vivo and in vitro (P 0.05 or P 0.01). The remarkable upregulation of anti-apoptotic Bcl-2 and downregulation of pro-apoptotic Bax protein level further confirmed this result. Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the PI3K-AKT and JAK2-STAT3 pathways were significantly enriched in SL. Compared with the model group, SL treatment significantly activated the PI3K-AKT and JAK2-STAT3 pathways in vivo and in vitro according to Western blot analyses.SL could protect the myocardium from MI injury. The underlying mechanism may be related to the reduction of inflammation and apoptosis by activating the PI3K/AKT and JAK2/STAT3 pathways.

It is important to utilize an antioxidant to ameliorate oxidative damage during boar sperm cryopreservation and thawing. Some studies have shown that Astragalus polysaccharide (APS) has antioxidant capabilities in sperm storage at low temperatures. However, the effects of APS on thawed sperm are unclear. In this study, the effect of supplementation of thawing boar semen extender with APS (0.5, 1, 5, 10 mg/mL) on sperm quality parameters (viability, motility, acrosome integrity and mitochondrial activity) was evaluated. Next, we investigated the effect of APS (0.5 mg/mL) supplementation on antioxidant parameters. Semen from two straws was thawed and diluted with three volumes of Beltsville Thawing Solution (BTS) and immediately divided into a control group without addition of antioxidants (CTR) and the APS group. Sperm and antioxidant parameters were evaluated in the CTR and APS groups after 1 h of incubation at 37 °C. Finally, we studied the effect of APS (0.5 mg/mL) supplementation on in vitro fertilization (IVF) and embryo development. The addition of different doses of APS to thawed sperm did not induce any significant effects on the sperm viability or motility compared to the sperm without APS treatment. However, the addition of 0.5 mg/mL APS to thawed sperm showed improved mitochondrial activity, higher penetration rate and increased total IVF efficiency compared with those of the control group. Moreover, our results indicate that the supplementation of APS in thawed sperm decreased the concentration of reactive oxygen species (ROS) and improved the activity of superoxide dismutase (SOD) and catalase (CAT). Finally, the addition of APS significantly increased the cleavage rate and blastocyst rate compared to those of the control group. In conclusion, the addition of APS to thawed boar sperm can enhance the antioxidant ability of sperm and improve in vitro fertilization (IVF) parameters and the outcomes of embryonic development. These results imply that APS has practical potential to enhance boar sperm reproductive performance.

Ya Jie Wang,1 Qi Li,1 Hong Bin Xiao,1 Yu Jie Li,1 Qing Yang,1 Xiao Xi Kan,1 Ying Chen,1 Xiao Ni Liu,2 Xiao Gang Weng,1 Xi Chen,1 Wei Yan Cai,1 Yan Guo,1 He Fei Huang,1 Xiao Xin Zhu11Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, 2Beijing Institute of Hepatology and Beijing Youan Hospital, Capital Medical University, Beijing, People’s Republic of ChinaAbstract: Multidrug resistance (MDR) is the main obstacle limiting the efficacy of cancer chemotherapy. Looking for novel anti-MDR agents is an important way to conquer cancer drug resistance. We recently established that chamaejasmin B (CHB), a natural biflavone from Stellera chamaejasme L., is the major active component. However, its anti-MDR activity is still unknown. This study investigated the anti-MDR effect of CHB and the underlying mechanisms. First, it was found that CHB inhibited the growth of both sensitive and resistant cell lines in vitro, and the average resistant factor (RF) of CHB was only 1.26. Furthermore, CHB also displayed favorable anti-MDR activity in KB and KBV200 cancer cells xenograft mice. Subsequent study showed that CHB induced G0/G1 cell cycle arrest as well as apoptosis both in KB and in resistant KBV200 cancer cells. Further studies showed that CHB had no influence on the level of Fas/FasL and activation of procaspase 8. However, CHB-induced apoptosis was dependent on the activation of caspase 9 and caspase 3. Moreover, CHB treatment resulted in the elevation of the Bax/Bcl-2 ratio, attenuation of mitochondrial membrane potential (ΔΨm), and release of cytochrome c and apoptosis-inducing factor from mitochondria into cytoplasm both in KB and KBV200 cells. In conclusion, CHB exhibited good anti-MDR activity in vitro and in vivo, and the underlying mechanisms may be related to the activation of mitochondrial-dependant intrinsic apoptosis pathway. These findings provide a new leading compound for MDR therapy and supply a new evidence for the potential of CHB to be employed in clinical trial of MDR therapy in cancers.Keywords: drug resistance, chamaejasmin B, apoptosis, anticancer, Bax/Bcl-2

A L9 (3(4)) orthogonal design table to be used to get nine combinations of extraction of three herbs of Wuji pill: Coptis chinensis, Tetradium ruticarpum and Paeonia lactiflora Pall., and nine extraction of single herbs correspondingly, altogether eighteen combinations. Quantification of five representative bioactive ingredients: berberine, palmatine, evodiamine, rutaecarpine, paeoniflorin in rat liver by ultra high liquid chromatography-tandem mass spectrometry after oral administration at 2 h time point of eighteen combinations. The result shows the bioactive ingredients have different concentrations betweem different combinations and the single herb with the same dosage significantly as well as the same dose combinations. C. chinensis with evodiamine concentration of low and high dose T. ruticarpum was positively correlated. T. ruticarpum with berberine concentration of low dose C. chinensis was negatively correlated and of meddle dose C. chinensis was correlated positively. T. ruticarpum with paeoniflorin concentration of middle dose P. lactiflora was correlated positively. P. lactiflora with palmatine concentration of middle dose C. chinensis was negatively correlated and with evodiamine and rutaecarpine concentration of middle dose T. ruticarpum was negatively correlated. These shows the three single herbs interactions resulted in the differences of each ingredients concentration in rat liver. The orthogonal analysis indicates the combination 12: 6: 6 make the maximum concentration in rat liver.

In this study, we aimed to investigate the therapeutic effects and involved mechanisms of resveratrol on an established non-alcoholic fatty liver disease (NAFLD) murine model. Wild-type and autophagic mediator ULK1 heterozygous knockout mice were induced to have NAFLD by high-fat diet for 8weeks. After that, resveratrol treatment was applied with the high-fat diet feeding for another 4weeks. Typical features of NAFLD, including histological changes, fibrosis, insulin resistance, oxidative status, and inflammation were characterized. After-treatment with resveratrol showed ameliorative effects on all measured features of NAFLD, from histology, insulin resistance, glucose tolerance to oxidative stress and inflammation. resveratrol treatment also reduced the activity of nuclear factor-κB (NF-κB) through the restoration of its inhibitor IκBα. Partial inhibition of ULK1 expression impaired the ameliorative effects of resveratrol on hepatic histology, fibrosis, oxidative status, inflammation, and NF-κB activity. In conclusion, resveratrol improved NAFLD-caused hepatic injury partially through regulating autophagic and IκBα-NF-κB pathways.

Probiotic could be a promising alternative to antibiotics for the prevention of enteric infections; however, further information on the dose effects is required. In this study, weanling piglets were orally administered low- or high-dose Lactobacillus rhamnosus ACTT 7469 (10(10) CFU/d or 10(12) CFU/d) for 1 week before F4 (K88)-positive Escherichia coli challenge. The compositions of faecal and gastrointestinal microbiota were recorded; gene expression in the intestines was assessed by real-time PCR; serum tumour necrosis factor-α (TNF-α) concentrations and intestinal Toll-like receptor 4 (TLR4) were detected by ELISA and immunohistochemistry, respectively. Unexpectedly, high-dose administration increased the incidence of diarrhoea before F4(+)ETEC challenge, despite the fact that both doses ameliorated F4(+)ETEC-induced diarrhoea with increased Lactobacillus and Bifidobacterium counts accompanied by reduced coliform shedding in faeces. Interestingly, L. rhamnosus administration reduced Lactobacillus and Bifidobacterium counts in the colonic contents, and the high-dose piglets also had lower Lactobacillius and Bacteroides counts in the ileal contents. An increase in the concentration of serum TNF-α induced by F4(+)ETEC was observed, but the increase was delayed by L. rhamnosus. In piglets exposed to F4(+)ETEC, jejunal TLR4 expression increased at the mRNA and protein levels, while jejunal interleukin (IL)-8 and ileal porcine β-defensins 2 (pBD2) mRNA expression increased; however, these increases were attenuated by administration of L. rhamnosus. Notably, expression of jejunal TLR2, ileal TLR9, Nod-like receptor NOD1 and TNF-α mRNA was upregulated in the low-dose piglets after F4(+)ETEC challenge, but not in the high-dose piglets. These findings indicate that pretreatment with a low dose of L. rhamnosus might be more effective than a high dose at ameliorating diarrhoea. There is a risk that high-dose L. rhamnosus pretreatment may negate the preventative effects, thus decreasing the prophylactic benefits against potential enteric pathogens. Our data suggest a safe threshold for preventative use of probiotics in clinical practice.

Using in vitro everted gut seas to research the intestinal absorption of the extractive Rhizoma Coptidis at the different intestinal section and the different density.Berberine (BER) and palmatine (PAL) which are representative compositions of the extractive Rhizoma Coptidis in everted gut seas are detected by HPLC, and calculated the absorption parameter to describe the character of absorption.The absorption of BER and PAL is linearity in different intestine and different dose, and the square of coefficient correlation exceed 0.9, which consistent with zero order rate process. The K(a) of BER and PAL increases along with the raised dosage of the extractive Rhizoma Coptidis (P0.05), indicated it is the passive absorption. The absorption of BER and PAL in the jejunum is the most quick, the ileum and colon are slower.In the different dosage of extractive Rhizoma Coptidis, the absorption of BER and PAL Conforms to the zero order rate process at the different intestine, and is the passive absorption.