Co-expression of Human Proteins (IL-10, TPO and/or Lactoferrin) into Milk of Cross-Breed Transgenic Mouse

Zheng, Zhen-Yu,Lee, Hyo-Sang,Oh, Keon-Bong,Koo, Deog-Bon,Han, Yong-Mahn,Lee, Kyung-Kwang The Korean Society of Animal Reproduction 2008 Reproductive & developmental biology Vol.32 No.1

We have previously produced transgenic (TG) mice expressing the human lactoferrin (hLF), interleukin-10 (hIL-10), and thrombopoietin (hTPO) proteins in the milk. In this study, we examined whether simple crossbreeding between two kids of a single transgenic mouse can produce double transgenics co-expressing two human proteins.. The hLF male, and the hIL-10 male were crossbred with the hIL-10 and hTPO females, and the hTPO female, respectively. PCR analysis for genotyping showed 32%, 23% and 24% double transgenic rates for hLF/hIL-10, hLF/hTPO, and hIL-10/hTPO transgenes, respectively. We analyzed the expression levels of the human proteins from double transgenic mice and compared those with their single transgenic siblings. All double transgenic co-expressed two human proteins at comparable levels to singles', unless hTPO was not co-expressed: for hLF, 1.1 mg/ml in hLF/hIL-10, whereas 0.5 mg/ml in hLF/hTPO; for hIL-10, 4.1 mg/ml in hIL-10/hLF, whereas 1.4 mg/ml in hIL-10/hTPO. Ihe downregulation of hTPO to half level of singles' was observed in double transgenic mice. The possible reason why hTPO co-expressed might lead to down-regulation of another human protein was discussed. These results suggested that double transgenic generated by crossbreeding between two singles' could be useful system for bioreactor.

Expression of the Transgene is Consistently Inherited to High Numbers of Generations and Independent on Its Source

Zhen-Yu Zheng,Keon Bong Oh,Deog-Bon Koo,Yong-Mahn Han,Kyung-Kwang Lee 한국동물생명공학회(구 한국동물번식학회) 2008 Reproductive & developmental biology Vol.32 No.1

Most studies on transgenic bioreactors have focused on expression levels of interest genes. In this study we examined whether transgenic bioreactors would inherit expression level of the Oansgene to long-term generations independently of transgene sources. We employed three transgenic mice, which were separately reported, carrying different transgenes and copy numbers, 27 kb of hLF and 22 kb of hIL-10 genomic sequences, and 1.3 kb of hTPO cDNA, respectively. Three females of the transgenic lineages crossbred with a wild-type male up to 20 generations to test transgenic frequencies of their progenies and to determine expression levels of the transgenes. Ultimately, transmission rates of kLF, hIL-10, and hTPO were 64.3+-7.0, 59.3+-9.8, and 56.1+-9.7, respectively, appeared following Mendelian pattern of inheritance. Notably, we found that levels of expressions of hLF, hIL-10, and hTPO in milk were sustained to high numbers of generations. No transgene silencing of expression was observed in every generations of all transgenic mice. In conclusion, we suggest that once established animal bioreactors could consistently transmit the transgene to continual generations, without loss of expressional activity, independently of transgene sources.

Co-expression of Human Proteins (IL-10, TPO and/or Lactoferrin) into Milk of Cross-Breed Transgenic Mouse

Zhen-Yu Zheng,Hyo-Sang Lee,Keon Bong Oh,Deog-Bon Koo,Yong-Mahn Han 한국동물생명공학회(구 한국동물번식학회) 2008 Reproductive & developmental biology Vol.32 No.1

We have previously produced transgenic (TG) mice expressing the human lactoferrin (hLF), interleukin-10 (hIL-10), and thrombopoietin (hTPO) proteins in the milk. In this study, we examined whether simple crossbreeding between two kids of a single transgenic mouse can produce double transgenics co-expressing two human proteins.. The hLF male, and the hIL-10 male were crossbred with the hIL-10 and hTPO females, and the hTPO female, respectively. PCR analysis for genotyping showed 32%, 23% and 24% double transgenic rates for hLF/hIL-10, hLF/hTPO, and hIL-10/hTPO transgenes, respectively. We analyzed the expression levels of the human proteins from double transgenic mice and compared those with their single transgenic siblings. All double transgenic co-expressed two human proteins at comparable levels to singles', unless hTPO was not co-expressed: for hLF, 1.1 mg/ml in hLF/hIL-10, whereas 0.5 mg/ml in hLF/hTPO; for hIL-10, 4.1 mg/ml in hIL-10/hLF, whereas 1.4 mg/ml in hIL-10/hTPO. Ihe downregulation of hTPO to half level of singles' was observed in double transgenic mice. The possible reason why hTPO co-expressed might lead to down-regulation of another human protein was discussed. These results suggested that double transgenic generated by crossbreeding between two singles' could be useful system for bioreactor.

Expression of the Transgene is Consistently Inherited to High Numbers of Generations and Independent on Its Source

Zheng, Zhen-Yu,Oh, Keon-Bong,Koo, Deog-Bon,Han, Yong-Mahn,Lee, Kyung-Kwang The Korean Society of Animal Reproduction 2008 Reproductive & developmental biology Vol.32 No.1

Most studies on transgenic bioreactors have focused on expression levels of interest genes. In this study we examined whether transgenic bioreactors would inherit expression level of the Oansgene to long-term generations independently of transgene sources. We employed three transgenic mice, which were separately reported, carrying different transgenes and copy numbers, 27 kb of hLF and 22 kb of hIL-10 genomic sequences, and 1.3 kb of hTPO cDNA, respectively. Three females of the transgenic lineages crossbred with a wild-type male up to 20 generations to test transgenic frequencies of their progenies and to determine expression levels of the transgenes. Ultimately, transmission rates of kLF, hIL-10, and hTPO were $64.3{\pm}7.0$, $59.3{\pm}9.8$, and $56.1{\pm}9.7$, respectively, appeared following Mendelian pattern of inheritance. Notably, we found that levels of expressions of hLF, hIL-10, and hTPO in milk were sustained to high numbers of generations. No transgene silencing of expression was observed in every generations of all transgenic mice. In conclusion, we suggest that once established animal bioreactors could consistently transmit the transgene to continual generations, without loss of expressional activity, independently of transgene sources.

Protective Effects of Polysaccharide Extracted from Portulacae oleracea L. on Colitis Induced by Dextran Sulfate Sodium

Zheng Wang,Yanni Liang,Dongbo Zhang,Xue Wu,Jingao Yu,Zhen Zhang,Yi Li,Chen Sun,Zhishu Tang,Li Liu 한국식품영양과학회 2020 Journal of medicinal food Vol.23 No.2

Polysaccharide from Ma-chi-xian (Portulacae oleracea L., POLP) was prepared and the therapeutic effect on dextran sodium sulfate-induced colitis mice was investigated in this study. The results of clinical activity score and H&E staining confirmed the therapeutic effect of POLP. POLP could diminished the symptoms of colitis and improve colon histopathological structure of the colitis mice. The expression levels of four cytokines were determined. The concentrations of PGE2 and IL-6 were downregulated by POLP treatment. The COX-2 protein expression levels and the STAT3 phosphorylation levels were detected. The results showed that these two protein levels were all increased in colitis and decreased after POLP treatment, indicating that these two proteins were closely related with the protective effect of POLP. Because the synthesis of PGE2 is catalyzed by COX-2 and phosphorylation of STAT3 can induce the expression of COX-2, it was concluded that STAT3 was a key protein related to the POLP exerting its activity in colitis.

Construction of Processing Maps based on Expanded Data by BP-ANN and Identification of Optimal Deforming Parameters for Ti-6Al-4V Alloy

Guo-zheng Quan,Hai-rong Wen,Jia-Pan,Zhen-yu Zou 한국정밀공학회 2016 International Journal of Precision Engineering and Vol.17 No.2

The intrinsic relationships between deforming parameters and microstructural mechanisms for Ti-6Al-4V alloy were analyzed by processing maps. A series of thermal compression tests were carried out in the temperatures range of 1023~1323 K (across β-transus) and strain rates range of 0.01~10 s-1 on a Gleeble-3500 thermo-mechanical simulator. Based on the stress-strain data collected from compression tests, a back-propagation artificial neural network (BP-ANN) model was developed, which presents reliable performance in tracking and predicting strain-stress data. By utilizing this model, the volume of stress-strain data was expanded. According to the intensive stress-strain data, the apparent activation energy was calculated to be 564.05 kJ mol-1 and 300.20 kJ mol-1 for α+β-phase field and single β-phase field, respectively. Moreover, the processing maps were constructed at finer intervals of temperature, from which, the stable regions with higher power dissipation efficiency (η > 0.3) and unstable regions with negative instability parameter (ξ < 0) were clarified clearly. By combining processing map with microstructure observations, two main stable softening mechanisms, i.e., globularization and dynamic recovery (DRV) were identified, and globularization-predominant (0.3 < η < 0.55) parameter domain ( < 0.1 s-1) in α+β-phase field and DRV-predominant (0.25 < η < 0.41) parameter domain (0.032 s-1< <1 s-1) in β- phase field were recommended. Manuscript

Modelling of the Hot Flow Behaviors for Ti-13Nb-13Zr Alloy by BP-ANN Model and Its Application

Guo-zheng Quan,Shi-ao Pu,Zong-yang Zhan,Zhen-yu Zou,Ying-ying Liu,Yu-feng Xia 한국정밀공학회 2015 International Journal of Precision Engineering and Vol. No.

The plastic deformation mechanisms and the constitutive model of flow behaviors at different deformation conditions in biomedical titanium alloy are an essential step to optimize the design of any forging process for implant productions. A series of isothermal compressions tests on Ti-13Nb-13Zr alloy in a wide range of true strain, temperature and strain rate were conducted on a thermomechanical simulator. The hot flow behaviors with different softening mechanisms, including dynamic recrystallization and dynamic recovery, were characterized based on true strain-stress curves. A back-propagational artificial neural network (BP-ANN) method was conducted to evaluate and predict this non-linear problem by self-training to be adaptable to the material characteristics. The flow stress of this material a wide deformation condition range can be predicted accurately by the BP-ANN model obtained in this study. The prediction ability of this BP-ANN Model was evaluated by three accuracy indexes, Absolute error, Relative error and Average absolute relative error. Sequently, the developed BP-ANN model was programed and implanted into the finite element (FE) analysis platform, Msc.Marc software. The results have sufficiently articulated that the well-trained ANN model has excellent capability to deal with the complex flow behaviors and has great application potentiality in hot deformation processes.

Multi-Variable and Bi-Objective Optimization of Electric Upsetting Process for Grain Refinement and Its Uniform Distribution

Guo-zheng Quan,Le Zhang,Chao An,Zhen-yu Zou 한국정밀공학회 2018 International Journal of Precision Engineering and Vol.19 No.6

It is significant to adjust the microstructures of preforms in pursuit of high-quality exhaust valves. This work is a novel attempt to identify the optimum process parameters in electric upsetting of 3Cr20Ni10W2 high-alloy to achieve grain refinement and uniform distribution by multi-objective genetic algorithm (MOGA) optimization. A finite element (FE) model on basis of electric-thermalmechanical and macro-micro sequential multi-physics analysis methods was developed in software MSC. Mar. And different schedules of four independent process variables (heating current (I), clamping length (L), upsetting pressure (Pset) and velocity of the anvil cylinder (v)) were performed aiming to achieve two objective indicators (average grain size (dav) and inhomogeneous degree of grain distribution (σd)). Then, two objective response surfaces were constructed as the functions between the two indicators and the four independent process variables. As per the criterion that simultaneously minimize dav and σd, the processing parameters (Pset, L, v, I) were optimized by MOGA, and corresponding numerical simulation were performed. The results show that both dav and σd are improved significantly at the optimal process condition as verified by the trial-manufacture experiments, which validated the optimal design and corresponding simulation based on grain refinement and uniform distribution by MOGA was credible and effective.

Effect of Cornus Officinalis Extract on the Differentiation of MC3T3-E1 Osteoblast-like Cells

( Ming Zhen Zheng ),( Sook Young Lee ),( Sang Joun Yu ),( Byung Ock Kim ) 한국조직공학·재생의학회 2015 조직공학과 재생의학 Vol.12 No.2s

Cornus officinalis has been widely used as herbal medicine exerting various pharmacological actions such as diuretic, anti-allergic, hypotensive and antibiotic activities. Thus we studied the effect of Cornus officinalis on the proliferation, differentiation and matrix mineralization of MC3T3-E1 osteoblast-like cell line. To do the experiments, Cornus officinalis extract (COE) was prepared using ethanol at different concentrations. The cytotoxic effect and the osteogenic differentiation were examined using MTT assay, alkaline phosphatase (ALP) activity, ALP staining, Alizarin red S staining and reverse transcriptase-polymerase chain reaction for osteogenic differentiation markers such as collagen type I (Col-I), ostoecalcin (OCN). The results indicated that there was no effect on cell proliferation on both 1st and 3rd days after treatment in MC3T-E1 cells. The level of ALP, Col-I mRNA, OCN mRNA and mineralized nodule were significantly higher in COE treated group than no treated group. Our results suggest that the COE promotes the osteoblast differentiation and mineralization without observable cytotoxicity. Therefore, it may play an important role in osteoblastic bone formation, and may possibly to be an effective measure for bone regeneration. Tissue Eng Regen Med 2015;12(Suppl 2):113-121

Tumor-Derived Transforming Growth Factor-β is Critical for Tumor Progression and Evasion from Immune Surveillance

Li, Zheng,Zhang, Li-Juan,Zhang, Hong-Ru,Tian, Gao-Fei,Tian, Jun,Mao, Xiao-Li,Jia, Zheng-Hu,Meng, Zi-Yu,Zhao, Li-Qing,Yin, Zhi-Nan,Wu, Zhen-Zhou Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.13

Tumors have evolved numerous mechanisms by which they can escape from immune surveillance. One of these is to produce immunosuppressive cytokines. Transforming growth factor-${\beta}$(TGF-${\beta}$) is a pleiotropic cytokine with a crucial function in mediating immune suppression, especially in the tumor microenvironment. TGF-${\beta}$ produced by T cells has been demonstrated as an important factor for suppressing antitumor immune responses, but the role of tumor-derived TGF-${\beta}$ in this process is poorly understood. In this study, we demonstrated that knockdown of tumor-derived TGF-${\beta}$ using shRNA resulted in dramatically reduced tumor size, slowing tumor formation, prolonging survival rate of tumor-bearing mice and inhibiting metastasis. We revealed possible underlying mechanisms as reducing the number of myeloid-derived suppressor cells (MDSC) and $CD4^+Foxp3^+$ Treg cells, and consequently enhanced IFN-${\gamma}$ production by CTLs. Knockdown of tumor-derived TGF-${\beta}$ also significantly reduced the conversion of na$\ddot{i}$ve $CD4^+$ T cells into Treg cells in vitro. Finally, we found that knockdown of TGF-${\beta}$ suppressed cell migration, but did not change the proliferation and apoptosis of tumor cells in vitro. In summary, our study provided evidence that tumor-derived TGF-${\beta}$ is a critical factor for tumor progression and evasion of immune surveillance, and blocking tumor-derived TGF-${\beta}$ may serve as a potential therapeutic approach for cancer.