Versatile Polymerization-Induced Emission Polymers from Barbier Polymerization of Cinnamic Esters with Tunable Emission.

Cinnamic ester is a common and abundant chemical substance, which can be extracted from natural plants. Compared with traditional esters, cinnamic ester contains α,ß-unsaturated carbonyl structure with multiple reactive sites, resulting in more abundant reactivities and chemical structures. Here, a versatile polymerization-induced emission (PIE) is successfully demonstrated through Barbier polymerization of cinnamic ester. Attributed to its abundant reactivities of α,ß-unsaturated carbonyl structure, Barbier polymerization of cinnamic esters with different organodihalides gives polyalcohol and polyketone via 1,2-addition and 1,4-addition, respectively, which is also confirmed by small molecular model reactions. Meanwhile, these organodihalides dependant polyalcohol and polyketone exhibit different non-traditional intrinsic luminescence (NTIL) from aggregation-induced emission (AIE) type to aggregation-caused quenching (ACQ) type, where novel PIE luminogens (PIEgens) are revealed. Further potential applications in explosive detection are carried out, where it achieves TNT detection sensitivity at ppm level in solution and ng level on the test paper. This work therefore expands the structure and functionality libraries of monomer, polymer and NTIL, which might cause inspirations to different fields including polymer chemistry, NTIL, AIE and PIE.

Clickable Polymerization-Induced Emission Luminogens Toward Color-Tunable Modification of Non-Traditional Intrinsic Luminescent Polymers.

Non-traditional intrinsic luminescent (NTIL) polymer is an emerging field, and its color-tunable modification is highly desirable but still rarely investigated. Here, a click chemistry approach for the color-tunable modifications of NTIL polymers by introducing clickable polymerization-induced emission luminogen (PIEgen), is demonstrated. Through Cu-catalyzed azide-alkyne cycloaddition click chemistry, a series of PIEgens is successful prepared, which is further polymerized via reversible addition-fragmentation chain transfer (RAFT) polymerization. Interestingly, after clickable modification, these monomers are nonemissive in both solution and aggregation states; while, the corresponding polymers exhibit intriguing aggregation-induced emission (AIE) characteristics, confirming their PIEgen characteristics. By varying alkynyl substitutions, color-tunable NTIL polymers are achieved with emission wavelength varying from 448 to 498 nm, revealing a series of PIEgens and verifying the importance of modification of NTIL polymers. Further luminescence energy transfer application is carried out as well. This work therefore designs a series of clickable PIEgens and opens a new avenue for the modification of NTIL polymers via click chemistry, which may cause inspirations to the research fields including luminescent polymer, NTIL, click chemistry, AIE and modification.

Fibroblast growth factor 21 ameliorates pancreatic fibrogenesis via regulating polarization of macrophages.

Chronic pancreatitis (CP) is a progressive, irreversible inflammatory and fibrotic disease. The characteristics of this disease are progressive inflammation, acinar atrophy and fibrosis. Numerous factors are involved in CP such as inflammation, and oxidative stress. Recently, it has been noted that fibroblast growth factor 21 (FGF-21) reduced the severity of acute pancreatitis in mice. However, whether FGF-21 has effects on CP remains unclear. Thus, the present study was undertaken to detect the effects of FGF-21 on l-arginine induced chronic pancreatitis/islet fibrosis in mice. We used l-arginine to create a CP model in C57BL/6 mice and treated these mice with FGF-21. Compared to normal mice, blood glucose and intra-peritoneal glucose tolerance test (IPGTT) revealed significant impairment in CP animal model. CP mice also had acinar atrophy, loss of pancreas morphology, inflammatory cells infiltration, extensive deposition of collagen, elevated -SMA expression, collagen I expression, serum amylase activity, MPO activity and MDA level. All these pathological changes were significantly improved by FGF-21 treatment. Moreover, FGF-21 ameliorated inflammatory state in the serum, pancreas and peritoneal macrophages of CP mice. Furthermore, we also found that FGF-21 could regulate differentiation of macrophages so as to improve pancreatic fibrogenesis in CP mice. Taken together, our study identifies the beneficial role of FGF-21 in CP and suggests that FGF-21 improves pancreatic fibrogenesis in CP via the mTOR pathway.

[Expression of recombinant h-FGF21 in periplasmic space of Escherichia coli].

Fibroblast growth factor 21 (FGF21) is a novel metabolic regulator of glucose and lipid, which is safe, effective and independent on insulin. FGF21 is considered as a prospective anti-diabetic drug. The aim of this study was to express recombinant h-FGF21 in periplasmic space of Escherichia coli. The pET27b plasmid was used to create the expression vectors of h-FGF21 with a PelB secretion signal. The ph-FGF21 (periplasmic expression of h-FGF21) was successfully expressed in the periplasm of E. coli BL21 (DE3), and soluble ph-FGF21 was isolated by disruption of the outer membrane. After twice of ion exchange chromatography, the purity of ph-FGF21 was above 95% in an analysis with a gray analysis software. The molecular weight of ph-FGF21 was about 20 kDa in SDS-PAGE and Western blotting analysis. The activity of ph-FGF21 and ih-FGF21 (intracellular expression of h-FGF21) was observed in vitro in the glucose uptake assay in HepG2 cells. The activity was observed in type 2 diabetic db/db mice after short or long-term treatments. The results suggest that the ph-FGF21 has a consistent activity with ih-FGF21 in vitro and in vivo.

[Mechanisms of the role of fibroblast growth factor 21 in attenuating insulin resistance].

This study is to evaluate the therapeutic effect of fibroblast growth factor 21 (FGF21) on type 2 diabetic mice model and to provide mechanistic insights into its therapeutic effect. Type 2 diabetic animal model was established with high calorie fat diet and low dose streptozotocin (STZ) injection. Mice were then randomized into 5 groups: model control, FGF21 0.25 and 0.05 µmol x kg(-1) x d(-1) groups, insulin treatment group. Ten age-matched normal KM mouse administered with saline were used as normal controls. Serum glucose, insulin, lipid products and the change of serum and liver tissue inflammation factor levels between five groups of mouse were determined. The results showed that blood glucose, insulin, free fatty acids (FFAs), triglycerides, and inflammatory factor average FGF-21 of type 2 diabetes model group and normal control group were significantly higher (P < 0.01), while compared with insulin group, no difference was significant. Average blood glucose, insulin, blood lipid and inflammatory factor of FGF-21 treatment group compared with type 2 diabetes group was significantly lower (P < 0.01) and insulin group has no difference with the model control group. The results of OGTT and HOMA-IR showed that insulin resistance state was significantly relieved in a dose-dependent manner. Thus, this study demonstrates that FGF-21 significantly remits type 2 diabetic mice model's insulin resistance state and participates in the regulation of inflammatory factor levels and type 2 diabetes metabolic disorders.

Removing residual DNA from Vero-cell culture-derived human rabies vaccine by using nuclease.

The clearance of host cell DNA is a critical indicator for Vero-cell culture-derived rabies vaccine. In this study, we evaluated the clearance of DNA in Vero-cell culture-derived rabies vaccine by purification process utilizing ultrafiltration, nuclease digestion, and gel filtration chromatography. The results showed that the bioprocess of using nuclease decreased residual DNA. Dot-blot hybridization analysis showed that the residual host cell DNA was <100 pg/ml in the final product. The residual nuclease in rabies vaccine was less than 0.1 ng/ml protein. The residual nuclease could not paly the biologically active role of digestion of DNA. Experiments of stability showed that the freeze-drying rabies virus vaccine was stable and titers were >5.0 IU/ml. Immunogenicity test and protection experiments indicated mice were greatly induced generation of neutralizing antibodies and invoked protective effects immunized with intraperitoneal injections of the rabies vaccine. These results demonstrated that the residual DNA was removed from virus particles and nuclease was removed by gel filtration chromatography. The date indicated that technology was an efficient method to produce rabies vaccine for human use by using nuclease.

Anchored periplasmic expression (APEx)-based bacterial display for rapid and high-throughput screening of B cell epitopes.

We truncated the VP2 protein of infectious bursal disease virus into five fragments: V1-5. All fragments were displayed on the inner membrane of the Escherichia coli periplasm. After disruption of the outer membrane, spheroplasts that had anchored with the VP2 fragment were incubated with an anti-VP2 polyclonal antibody (pAb). Prey pairs were detected and quantitated by flow cytometry with V1, V3, V4 and V5 fragments reacting with the pAb. The antigenicity of all five fragments was analyzed, and our results indicated that epitopes were localized in V1, V3, V4 and V5, consistent with our flow cytometry analysis. Antigenicity analysis of purified VP2 fusion proteins using Western blots confirmed this. Our method provides a rapid, quantitative and simple strategy for identifying linear B cell epitopes.

scFv antibodies against infectious bursal disease virus isolated from a combinatorial antibody library by flow cytometry.

Infectious bursal disease is an economically important disease that affects chickens worldwide. Here, a recombinant single chain variable fragment (scFv) antibody library derived from chickens immunized with VP2 protein of infectious bursal disease virus (IBDV) was constructed. The library was subjected to three rounds of screening by flow cytometry against VP2 protein through a bacteria display technology, resulting in the enrichment of scFv. Three scFv clones with different fluorescence intensity were obtained by random colony pick up. The isolated scFv antibodies were expressed and purified. Relative affinity assay showed the three clones had different sensitivity to VP2, in accordance with fluorescence activity cell sorting analysis. The potential use of the selected IBDV-specific scFv antibodies was demonstrated by the successful application of the isolated antibodies in western blotting assay and ELISA.

[FGF-21 protects H9c2 cardiomyoblasts against hydrogen peroxide-induced oxidative stress injury].

Fibroblast growth factor-21 (FGF-21) is an important metabolism regulator, however, whether FGF-21 has effects on cardiovascular remains unclear. In this study, H2O2-induced injury in H9c2 cells was used as a cell model, the anti-apoptosis potential and mechanism of FGF-21 against oxidative injury were evaluated by MTT assay, flow cytometry assay and real-time PCR. The results showed that FGF-21 could increase the cell survival of H2O2-induced injury in H9c2 cells and prevent H9c2 cells from oxidative stress-induced apoptosis. Furthermore, FGF-21 can elevate SOD activity and regulate Bcl-2/Bax expression in H9c2 cells. The results suggest that FGF-21 have protective effect against the H2O2-induced apoptosis in H9c2 cells.

[The synergistic effect of FGF-21 and insulin on regulating glucose metabolism and its mechanism].

Previous studies proposed that the synergistic effect of fibroblast growth factor-21 (FGF-21) and insulin may be due to the improvement of insulin sensitivity by FGF-21. However, there is no experimental evidence to support this. This study was designed to elucidate the mechanism of synergistic effect of FGF-21 and insulin in the regulation of glucose metabolism. The synergistic effect of FGF-21 and insulin on regulating glucose metabolism was demonstrated by investigating the glucose absorption rate by insulin resistance HepG2 cell model and the blood glucose chances in type 2 diabetic db/db mice after treatments with different concentrations of FGF-21 or/and insulin; The synergistic metabolism was revealed through detecting GLUT1 and GLUT4 transcription levels in the liver by real-time PCR method. The experimental results showed that FGF-21 and insulin have a synergistic effect on the regulation of glucose metabolism. The results of real-time PCR showed that the effective dose of FGF-21 could up-regulate the transcription level of GLUT1 in a dose-dependent manner, but had no effect on the transcription level of GLUT4. Insulin (4 u) alone could up-regulate the transcription level of GLUT4, yet had no effect on that of GLUT1. Ineffective dose 0.1 mg kg(-1) FGF-21 alone could not change the transcription level of GLUT1 or GLUT4. However, when the ineffective dose 0.1 mg x kg(-1) FGF-21 was used in combination with insulin (4 u) significantly increased the transcription levels of both GLUT1 and GLUT4, the transcription level of GLUT1 was similar to that treated with 5 time concentration of FGF-21 alone; the transcription level of GLUT4 is higher than that treated with insulin (4 u) alone. In summary, in the presence of FGF-21, insulin increases the sensitivity of FGF-21 through enhancing GLUT1 transcription. Vice versa, FGF-21 increases the sensitivity of insulin by stimulating GLUT4 transcription in the presence of insulin. FGF-21 and insulin exert a synergistic effect on glucose metabolism through mutual sensitization.

[The synergism and mechanism of action of rClone30-hDR5 in combination with TRAIL on HCC].

To investigate the cell-killing effect and its possible mechanism of rClone30-hDR5 in combination with TRAIL on human hepatic carcinoma (HCC) cell line, first of all, recombinant plasmid pee12.4-hDR5 was introduced into HepG2 cells by liposome transfection. After five rounds of screening by flow cytometry, HepG2 cells expressing high levels of DR5 on cell surface were isolated. The cytotoxicity of TRAIL to selected cells was higher than that of TRAIL to HepG2 cells by MTT method (P < 0.01). The result suggested that the cloned hDR5 gene had biological activity. MTT assay showed that, rClone30- hDR5 in combination with TRAIL more efficiently inhibited the tumor growth of HepG2 cells compared to rClone30-hDR5 or TRAIL in vitro. The results of Annexin V-FITC/PI staining and Quantitative Real-time PCR indicated that rClone30-hDR5 in combination with TRAIL significantly increased the mRNA levels of caspase 3 and caspase 8, and induced the apoptosis of tumor cells. HepG2 cells were infected with rClone30-hDR5 or rClone30 at MOI of 1. The expression of hDR5 on tumor surface increased significantly by rClone30-hDR5 compared to that by rClone30, which contributed to the sensitivity to TRAIL. In conclusion, rClone30-hDR5 in combination with TRAIL has potential application value in cancer treatment.

[Effect of FGF-21 on learning and memory ability and antioxidant capacity in brain tissue of D-galactose-induced aging mice].

This study aims to investigate the effects of fibroblast growth factor 21 (FGF-21) on learning and memory abilities and antioxidant capacity of D-galactose-induced aging mice. Kunming mice (37.1 +/- 0.62) g were randomly divided into normal control group, model group and FGF-21 high, medium and low dose groups (n = 8). Each group was injected in cervical part subcutaneously with D-galactose 180 mg x kg(-1) x d(-1) once a day for 8 weeks. At the same time, FGF-21-treated mice were administered with FGF-21 by giving subcutaneous injection in cervical part at the daily doses of 5, 2 and 1 mg x kg(-1) x d(-1). The normal control group was given with normal saline by subcutaneous injection in cervical part. At seventh week of the experiment, the learning and memory abilities of mice were determined by water maze and jumping stand tests. At the end of the experiment, the mice were sacrificed and the cells damage of hippocampus was observed by HE staining in each group. Reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) and total antioxidant capacity (T-AOC) in the brain of mice were determined. The results showed that different doses of FGF-21 could reduce the time reaching the end (P < 0.01 or P < 0.05) and the number of touching blind side (P < 0.01 or P < 0.05) in the water maze comparing with the model group. It could also prolong the latency time (P < 0.05) and decrease the number of errors (P < 0.01 or P < 0.05) in the step down test. The result of HE staining showed that FGF-21 could significantly reduce brain cell damage in the hippocampus. The ROS and MDA levels of three different doses FGF-21 treatment group reduced significantly than that of the model group [(5.58 +/- 1.07), (7.78 +/- 1.92), (9.03 +/- 1.77) vs (12.75 +/- 2.02) pmol (DCF) x min(-1) x mg(-1), P < 0.01 or P < 0.05], [(2.92 +/- 0.71), (4.21 +/- 0.81), (4.41 +/- 0.97) vs (5.62 +/- 0.63) nmol x mg(-1) (protein), P < 0.01]. Comparing with the model group, the activities of SOD, GPx, CAT and T-AOC of the three different doses FGF-21 treatment groups were also improved in a dose-dependent manner. This study demonstrates that FGF-21 can ameliorate learning and memory abilities of D-galactose induced aging mice, improve the antioxidant abilities in brain tissue and delay brain aging. This finding provides a theoretical support for clinical application of FGF-21 as a novel therapeutics for preventing aging.

[Antitumor efficacy of the recombinant Newcastle disease virus rNDV-IL15 on melanoma models].

In order to enhance the antitumor efficacy of recombinant Newcastle disease virus, rNDV-IL15 was rescued in this study. Recombinant plasmid prNDV-IL15 was constructed, and BHK21 cells were transfected with the recombinant plasmid. Finally, the recombinant Newcastle disease virus rNDV-IL15 was successfully rescued. The growth curves of these two recombinant viruses were determined. Murine melanoma B16F10 cells were infected with rNDV-IL15 at MOI of 0.1, and the expression level of IL15 in the supernatant was detected by ELISA. The antitumor efficacy of rNDV-IL15 and rNDV was compared in vitro and in vivo. Results showed that prNDV-IL15 was constructed and recombinant virus rNDV-IL15 was successfully rescued. The growth curve of rNDV-IL15 showed that the growth of rNDV-IL15 had not been changed after insertion of IL15 gene. Results showed that there was high level of IL15 expression in the supernatant of rNDV-IL5-infected B16F10 cells (1 044.3 +/- 27.7 ng x mL(-1)). rNDV-IL15 and rNDV significantly inhibited the growth of B16F10 cells in vitro in a time-dependent manner. However, there was no significant difference between them. In animal experiments, rNDV-IL15 efficiently suppressed tumor growth in vivo when compared with rNDV, and the difference was statistically significant. The results suggested that rNDV-IL15 is a more effective antitumor agent.

[Therapeutic efficacy of three bispecific antibodies on rheumatoid arthritis mice models].

In order to obtain the lead compound for treatment of rheumatoid arthritis (RA), in this study, therapeutic efficacy of three bispecific antibodies (BsAB-1, BsAB-2 and BsAB-3) against both hIL-1beta and hIL-17 were compared on CIA model mice. First, by ELISA method we compared the binding capacity of the three bispecific antibodies to the two antigens. The results showed that all three antibodies could simultaneously bind both antigens, among these antibodies, BsAB-1 was superior over BsAB-2 and BsAB-3. CIA model was established with chicken type II collagen (CII) and developed RA-like symptoms such as ankle swelling, skin tight, hind foot skin hyperemia. The CIA mice were treated with three antibodies once every two days for total of 29 days. Compared with the CIA model mice, the RA-like symptoms of the antibody treated-mice significantly relieved, while the BsAB-1 treated-mice were almost recovered. CII antibody level in the serum and cytokines (IL-2, IL-1beta, IL-17A and TNF-alpha) expression in the spleen were examined. Compared with the CIA model mice, all three antibodies could significantly reduce CII antibody and cytokine expression levels. BsAB-1 antibody was more potent than BsAB-2 and BsAB-3. In summary, BsAB-1 is superior over BsAB-2 and BsAB-3 in amelioration of RA symptoms and regulation of CII antibody production and pro-inflammatory cytokine expression, therefore, BsAB-1 can be chosen as a lead compound for further development of drug candidate for treatment of RA.

[Antitumor activity of the recombinant rClone30-CD/5-FC system].

5-Flucytosine (5-FC) could be changed to 5-fluorouracil (5-FU) by cytosine deaminase (CD), the latter is able to kill cancer cells. However, there is no efficient method to deliver the CD gene into the tumor cells, which hampers the application of the suicide gene system. In this experiment, for the first time, the NDV has been utilized as a vector to deliver the CD gene into the cancer cells, the virus can infect the cancer cells specifically, replicate and assemble, while the cytosine deaminase is expressed. Then the CD converts the prodrug 5-FC into 5-FU to achieve the purpose of inhibiting tumor. Firstly, the whole genome of E. coli JM109 was extracted, and the CD gene was obtained by cloning method. Then the CD and IRES-EGFP were ligated into the pEE12.4 expression vector to become a recombinant pEE12.4IE-CD eukaryotic expression plasmid. The human liver cancer cells were transfected with the plasmid. The cells were treated with different concentrations of 5-FC, MTT method was used to determine the killing effect of CD/5-FC system on the human liver cancer cells. The cell deaths were 18.07%, 42.98% and 62.20% respectively when the concentrations of prodrug were at 10, 20 and 30 mmol x L(-1). In 5-FC acute toxicity experiment, Kunming mice were injected with different concentrations of 5-FC at intervals of 1:0.5. The LD50 of 5-FC through iv injection was determined by improved Karber's method, the LD50 was 507 mg x kg(-1) and the 95% confidence limit was 374-695 mg x kg(-1). According to the maximum LD0 dose of the LD50, the maximum safe dose was 200 mg x kg(-1). Body weight and clinic symptoms of the experimental animals were observed. These results laid the foundation to verify the antitumor effect and safety of CD/5-FC system in animal models. The CD gene was ligated into the NDV (rClone30) carrier, then the tumor-bearing animal was established to perform the tumor inhibiting experiment. The result showed that the recombinant rClone30-CD/5-FC system has a high antitumor activity in vivo. To summarize, CD gene has been cloned and its bioactivity has been confirmed in the mammalian cells. It is the first time in this study to utilize the recombinant NDV to deliver the CD gene into the tumor cells; our result proves the rClone30-CD/5-FC system is a potential method for cancer therapy.

[The long lasting effect of the murine fibroblast growth factor-21 on blood glucose control of diabetic animals].

Insulin is the most common medicine used for diabetic patients, unfortunately, its effective time is short, even the long-acting insulin cannot obtain a satisfactory effect. Fibroblast growth factor (FGF)-21 is a recently discovered glucose mediator and expected to be a potential anti-diabetic drug that does not rely on insulin. In this study, db/db mice were used as the type 2 diabetic model to examine whether mFGF-21 has the long-term blood lowering effect on the animal model. The results showed that mFGF-21 could stably maintain the blood glucose at normal level for a long-term in a dose-dependent manner. Administration of mFGF-21 once a day with three doses (0.125, 0.25 and 0.5 mg x kg(-1)) could maintain blood glucose of the model animals at normal level for at least 24 h. Administration of mFGF-21 every two days with the same doses could maintain blood glucose of the model animals at normal level for at least 48 h, although it took longer time for blood glucose to reach to normal level depending on doses used (twenty injections for 0.125 mg x kg(-1) and 0.25 mg x kg(-1) doses, ten injections for 0.5 mg x kg(-1) dose). Surprisingly, the blood glucose of the treated model animals still maintained at normal level for 24 h after the experiment terminated. Glycosylated hemoglobin level of the animals treated with mFGF-21, which represented long-term glucose status, decreased significantly compared to the control group and the insulin group. The results suggest that FGF-21 has potential to become a long-acting and potent anti-diabetic drug.

[Therapeutic effect of fibroblast growth factor 21 on hypertension induced by insulin resistance].

This study is to evaluate the therapeutic effect of fibroblast growth factor 21 (FGF21) on hypertension induced by insulin resistance in rats and to provide mechanistic insights into its therapeutic effect. Male Sprague-Dawley (SD) rats were fed with high-fructose (10%) water to develop mild hypertensive models within 4 weeks, then randomized into 4 groups: model control, FGF21 0.25, 0.1 and 0.05 micromol x kg(-1) x d(-1) groups. Five age-matched normal SD rats administrated with saline were used as normal controls. The rats in each group were treated once a day for 4 weeks. Body weight was measured weekly, systolic blood pressure (SBP) was measured noninvasively using a tail-cuff method, insulin sensitivity was assessed using oral glucose tolerance test (OGTT) and HOMA-IR assay. At the end of the treatment, blood samples were collected, and blood glucose, serum cholesterol, serum triglyceride and serum insulin were measured. The results showed that blood pressure of the rats treated with different doses of FGF21 returned to normal levels [(122.2 +/- 3.5) mmHg, P < 0.01] after 4-week treatment, whereas, SBP of untreated (model control) rats maintained a high level [(142.5 +/- 4.5) mmHg] throughout the treatment. The observation of blood pressure in 24 h revealed that SBP of FGF21 treated-rats maintained at (130 +/- 4.5) mmHg vs. (143 +/- 5.5) mmHg for model control (P < 0.01). FGF21 treatment groups improved serum lipids obviously, total cholesterol (TC) and triglyceride (TG) levels decreased significantly to normal levels. The serum NO levels of three different doses FGF21 treatment group were significantly higher than that of the model control group [(7.32 +/- 0.11), (7.24 +/- 0.13), (6.94 +/- 0.08) vs. (6.56 +/- 0.19) micromol x L(-1), P < 0.01], and the degree of improvement showed obvious dose-dependent manner, indicating that FGF21 can significant increase serum NO in fructose-induced hypertension rat model and improve endothelial NO release function. The results of OGTT and HOMA-IR showed that insulin resistance state was significantly relieved in a dose-dependent manner. Thus, this study demonstrates that FGF21 significantly ameliorates blood pressure in fructose-induced hypertension model by relieving insulin resistance. This finding provides a theoretical support for clinical application of FGF21 as a novel therapeutics for treatment of essential hypertension.

[Therapeutic effect of fibroblast growth factor 21 on NAFLD in MSG-iR mice and its mechanism].

This study is to evaluate the therapeutic effect of fibroblast growth factor 21 (FGF21) on NAFLD in MSG-IR mice and to provide mechanism insights into its therapeutic effect. The MSG-IR mice with insulin resistance were treated with high dose (0.1 micromol.kg-1d-1) and low dose (0.025 micromol.kg-1d-1) of FGF21 once a day for 5 weeks. Body weight was measured weekly. At the end of the experiment, serum lipids, insulin and aminotransferases were measured. Hepatic steatosis was observed. The expression of key genes regulating energy metabolism were detected by real-time PCR. The results showed that after 5 weeks treatment, both doses of FGF21 reduced body weight (P<0.01), corrected dyslipidemia (P<0.01), reversed steatosis and restored the liver morphology in the MSG model mice and significantly ameliorated insulin resistance. Additionally, real-time PCR showed that FGF21 significantly reduced transcription levels of fat synthetic genes, decreased fat synthesis and promoted lipolysis and energy metabolism by up-regulating key genes of lipolysis, thereby liver fat accumulation was reduced and liver function was restored to normal levels. In conclusion, FGF21 significantly reduces body weight of the MSG-IR mice, ameliorates insulin resistance, reverses hepatic steatosis. These findings provide a theoretical support for clinical application of FGF21 as a novel therapeutics for treatment of NAFLD.

Generation and characterization of a recombinant Newcastle disease virus expressing the red fluorescent protein for use in co-infection studies.

BACKGROUND: Many viruses have evolved multiple strategies to prevent super infection of host cells by more than one virion. This phenomenon, known as super infection exclusion, may play an important role on virus evolution because it can affect the frequency of reassortment and/or recombination. Newcastle disease virus (NDV), a negative sense single-stranded RNA virus, is characterized by its continuous evolutionary dynamics and by a low frequency of recombination events. However, the mechanisms that contribute to the low recombination rates on NDV are still not completely understood. METHODS: In this study we assessed the ability of two NDV strains (LaSota and B1) to super infect host cells in vitro. We generated a recombinant NDV strain LaSota expressing the red fluorescent protein (RFP) and used it in co-infection assays with a related NDV strain B1 expressing the green fluorescent protein (GFP). DF-1 cells were inoculated with both viruses at the same time or at different intervals between primary infection and super infection. RESULTS: When both viruses were inoculated at the same time point, a 27% co-infection rate was observed, whereas when they were inoculated at different time points the super infection rates decreased to levels as low as 1.4%. CONCLUSIONS: These results indicate that although different NDV strains can co-infect host cells in vitro, the super infection rates are low, specially as the time between the primary infection and super infection increases. These results confirm the occurrence of super infection exclusion between different strains of NDV.

[Optimization and characterization of a novel FGF21 mutant].

Fibroblast growth factor 21 (FGF21) is a member of FGF family. It has been demonstrated that FGF21 is an independent, safe and effective regulator of blood glucose levels in vivo. In order to improve the activity of FGF21, we exchanged the beta10-beta12 domain of the human FGF21 with that of the mouse FGF21 to construct a novel FGF21 gene (named hmFGF21), and then subcloned hmFGF21 gene into the SUMO expression vector to create pSUMO-hmFGF21 and transformed it into E. coli Rosetta for expression of the fusion protein SUMO-hmFGF21. Both in vitro and in vivo glucose regulation activity of hmFGF21 was evaluated. The SDS-PAGE result showed that compared with wild-type hFGF21, the soluble expression of hmFGF21 increased about 2-fold. HmFGF21 was more potent in stimulation of glucose uptake in HepG2 cells in vitro. The results of anti-diabetic effect on db/db mice demonstrated that hmFGF21 had better efficacy on controlling the blood glucose of the db/db diabetic animals than wild-type hFGF21. These results suggest that the biological properties of FGF21 are significantly improved by optimization.