Non-integrating lentiviral vectors based on the minimal S/MAR sequence retain transgene expression in dividing cells

Safe and efficient gene transfer systems are the basis of gene therapy applications. Non-integrating lentiviral(NIL) vectors are among the most promising candidates for gene transfer tools, because they exhibit high transfer efficiency in both dividing and non-dividing cells and do not present a risk of insertional mutagenesis. However, non-integrating lentiviral vectors cannot introduce stable exogenous gene expression to dividing cells, thereby limiting their application. Here, we report the design of a non-integrating lentiviral vector that contains the minimal scaffold/matrix attachment region(S/MAR) sequence(SNIL), and this SNIL vector is able to retain episomal transgene expression in dividing cells. Using SNIL vectors, we detected the expression of the eGFP gene for 61 days in SNIL-transduced stable CHO cells, either with selection or not. In the NIL group without the S/MAR sequence, however, the transduced cells died under selection for the transient expression of NIL vectors. Furthermore,Southern blot assays demonstrated that the SNIL vectors were retained extrachromosomally in the CHO cells. In conclusion,the minimal S/MAR sequence retained the non-integrating lentiviral vectors in dividing cells, which indicates that SNIL vectors have the potential for use as a gene transfer tool.

Insulin producing cells established using non-integrated lentiviral vector harboring PDX1 gene

AIM: To investigate reprogramming of human adipose tissue derived stem cells into insulin producing cells using non-integrated lentivirus harboring PDX1 gene.METHODS: In this study, human adipose tissue derived stem cells(hADSCs) were obtained from abdominal adipose tissues by liposuction, selected by plastic adhesion, and characterized by flow cytometric analysis.Human ADSCs were differentiated into adipocytes and osteocytes using differentiating medium to confirm their multipotency. Non-integrated lentiviruses harboring PDX1(Non-integrated LV-PDX1) were constructed using specific plasmids(pLV-HELP, pMD2G, LV-105-PDX1-1).Then, hADSCs were transduced with non-integrated LVPDX1. After transduction, ADSCsPDX1+were cultured in high glucose DMEM medium supplement by B27, nicotinamide and βFGF for 21 d. Expressions of PDX1 andinsulin were detected at protein level by immunofluorescence analysis. Expressions of PDX1, neurogenin3(Ngn3), glucagon, glucose transporter2(Glut2) and somatostatin as specific marker genes were investigated at mRNA level by quantitative RT-PCR. Insulin secretion of hADSCsPDX1+in the high-glucose medium was detected by electrochemiluminescence test. Human ADSCsPDX1+were implanted into hyperglycemic rats.RESULTS: Human ADSCs exhibited their fibroblast-like morphology and made colonies after 7-10 d of culture.Determination of hADSCs identified by FACS analysis showed that hADSCs were positive for mesenchymal cell markers and negative for hematopoietic cell markers that guaranteed the lack of hematopoietic contamination. In vitro differentiation of hADSCs into osteocytes and adipocytes were detected by Alizarin red and Oil red O staining and confirmed their multilineage differentiation ability. Transduced hADSCs+PDX1became round and clusters in the differentiation medium. The appropriate expression of PDX1 and insulin proteins was confirmed using immunocytochemistry analysis.Significant expressions of PDX1, Ngn3, glucagon, Glut2and somatostatin were detected by quantitative RTPCR. hADSCsPDX1+revealed the glucose sensing ability by expressing Glut2 when they were cultured in the medium containing high glucose concentration. The insulin secretion of hADSCsPDX1+in the high glucose medium was 2.32 μU/mL. hADSCsPDX1+implantation into hyperglycemic rats cured it two days after injection by reducing blood glucose levels from 485 mg/dL to the normal level.CONCLUSION: Human ADSCs can differentiate into IPCs by non-integrated LV-PDX1 transduction and have the potential to be used as a resource in type 1 diabetes cell therapy.

FIRST-PASSAGE TIME OF QUASI-NON-INTEGRABLE-HAMILTONIAN SYSTEM

Studies on first-passage failure are extended to the multi-degree-of-freedom quasi-non-integrable-Hamiltonian systems under parametric excitations of Gaussian white noises in this paper. By the stochastic averaging method of energy envelope, the system's energy can be modeled as a one-dimensional approximate diffusion process by which the classical Pontryagin equation with suitable boundary conditions is applicable to analyzing the statistical moments of the first-passage time of an arbitrary order. An example is studied in detail and some numerical results are given to illustrate the above procedure.

STOCHASTIC OPTIMAL CONTROL FOR THE RESPONSE OF QUASI NON-INTEGRABLE HAMILTONIAN SYSTEMS

A strategy is proposed based on the stochastic averaging method for quasi non- integrable Hamiltonian systems and the stochastic dynamical programming principle.The pro- posed strategy can be used to design nonlinear stochastic optimal control to minimize the response of quasi non-integrable Hamiltonian systems subject to Gaussian white noise excitation.By using the stochastic averaging method for quasi non-integrable Hamiltonian systems the equations of motion of a controlled quasi non-integrable Hamiltonian system is reduced to a one-dimensional av- eraged It stochastic differential equation.By using the stochastic dynamical programming princi- ple the dynamical programming equation for minimizing the response of the system is formulated. The optimal control law is derived from the dynamical programming equation and the bounded control constraints.The response of optimally controlled systems is predicted through solving the FPK equation associated with It stochastic differential equation.An example is worked out in detail to illustrate the application of the control strategy proposed.

Non-integrable Movement Equations in Landsliding System

The answer of the question was considered as non-integrable basically. The integrable system was thought in the exceptional.Since Poincare had presented in a theorem and proved that non-integrable system is normal, and yet integrable system is abuormal. In the landsliding system, discussed question was regarded to be true.Sortie fundamentals were considered; other systems and example with engineering control are concerned and discussed in details.

On the Differential Formulas for Singular Integrals of High Non-Integral Order

When kernel density is in the class of continuous function to possessing sufficient derivative of high order(and needn't in the class of corresponding Holder function),in this paper it is given the continuity and the differential formulas for singular integrals of high non--integral order. The above results themselves and in order to prove in future the formulas to changing order of integration for singular integrals of high non-integral order(another paper) will have important significance. The method to prove in this paper is more different from the method in the corresponding cass of singular integrals of high integral order.

Six types of spin solitons in three-component Bose-Einstein condensates

Exact analytical solutions are good candidates for studying and explaining the dynamics of solitons in nonlinear systems.We further extend the region of existence of spin solitons in the nonlinearity coefficient space for the spin-1 Bose-Einstein condensate.Six types of spin soliton solutions can be obtained,and they exist in different regions.Stability analysis and numerical simulation results indicate that three types of spin solitons are stable against weak noise.The non-integrable properties of the model can induce shape oscillation and increase in speed after the collision between two spin solitons.These results further enrich the soliton family for non-integrable models and can provide theoretical references for experimental studies.

Exciting rogue waves, breathers, and solitons in coherent atomic media

We propose a scheme that excites rogue waves via electromagnetically induced transparency(EIT),which can also excite breathers and solitons.The system is a resonantΛ-type atomic ensemble.Under EIT conditions,the envelope equation of the probe field can be reduced to several different models,such as the saturable nonlinear Schr?dinger equation(SNLSE),and SNLSE with the trapping potential provided by a far-detuned laser field or a magnetic field.In this scheme,rogue waves can be generated by different initial pulses,such as the Gaussian wave with(or without)the uniform background.The scheme can be used to obtain rogue waves,breathers and solitons.We show the existence regions of rogue waves,breathers,and solitons as the function of the amplitude and width of the initial pulse.The novelty of our paper is that,we not only show rogue waves in the integrable system numerically,but also present the method to generate and control rogue waves in the non-integrable system.