In vitro targeting efficiency evaluation of reduction-responsive co-loaded doxorubicin/siRNA nanoparticles / 药学学报

In this study a reduction-responsive nanoparticles (NPs) modified with hyaluronic acid (HA) was prepared for the co-delivery of doxorubicin (DOX) and siRNA and then evaluated as a lung cancer targeting delivery system in vitro. The amphiphilic polymer of poly-L-lysine-lipoic acid (PLA) based on poly-L-lysine (PLL) with lipoic acid (LA) was synthesized via amidation reaction and characterized by 1H NMR. The DOX loaded PLA NPs were prepared via dialysis method, and siRNA was loaded via electrostatic attraction to prepare the co-delivery NPs system (PLA/DOX-siRNA-NPs). Then PLA/DOX-siRNA-NPs were coated with HA to obtain HA-PLA/DOX-siRNA-NPs. The tumor microenvironment-responsive properties under different pH or reduction condition of HA-PLA/DOX-siRNA-NPs were evaluated by investigating the particle size and zeta potential. Cellular uptake of HA-PLA/DOX-siRNAFAM-NPs by A549 cells and endosomal escape of siRNA were studied using confocal laser scanning microscope (CLSM). 1H NMR spectrum demonstrated that PLA was successfully synthesized with LA grafting rate of 25.1%. The encapsulation efficiency (EE) and drug loading (DL) of HA-PLA/DOX-NPs was (86.93±8.91)% and (4.17±0.68)%, respectively, and siRNA was loaded at an N/P of 6:1 in carrier. HA-PLA/DOX-siRNA-NPs exhibited a suitable size of (167.3±9.9) nm and negative charge of (-15.5±1.4) mV with the optimal ratio of PLA and HA of 1:3. Additionally, the zeta potential of HA-PLA/DOX-siRNA-NPs significantly increased with charge reversal from negative to positive after the treatment with HAase, and the particle size of HA-PLA/DOX-siRNA-NPs changed significantly under the condition of 10 mmol·L-1 glutathione (GSH). The release profiles in vitro demonstrated that HA-PLA/DOX-NPs exhibited a maintained release behavior at pH 7.4 and the adding of GSH (10 mmol·L-1) led to rapid release of DOX from NPs. In vitro cellular uptake and subcellular distribution study demonstrated that themodification of HA enhanced the affinity of NPs to A549 cells and targeting ability, and the cellular uptake of HA-PLA/DOX-siRNAFAM-NPs significantly increased after the treatment with HAase. It was observed that HA-PLA/DOX-siRNAFAM-NPs could escape from endo-lysosomes followed by sharp payloads release to their relative targets. All these results demonstrated that the co-loaded NPs have a high entrapment efficiency of DOX and siRNA. And they also exhibited an active tumor targeting efficiency and tumor microenvironment-responsive properties, which were beneficial to cellular uptake and intracellular release of DOX and siRNA. In conclusion, these reduction-responsive NPs modified with HA have great potential as co-delivery systems for antitumor agents and siRNA.

Effect of poly (L-lysine) modified silk fibroin film on the growth and differentiation of neural stem cells / 生物工程学报

In order to provide a basic theory for the materials of repairing central nervous system injury, we have studied the growth and differentiation of neural stem cells (NSCs) on poly (L-lysine) modified silk fibroin film. First, we used poly (L-lysine) to modify silk fibroin film and confirmed by UV-vis and 1H NMR spectra. Then NSCs were isolated and seeded on the silk fibroin film (Silk group), poly (L-lysine) (PLL group) and poly (L-lysine) modified Silk fibroin film (Silk-PIL group). The proliferation of NSCs was evaluated by Cell Counting Kit-8 (CCK-8) assay on days 1, 3, 5 and 7 after seeding. Immunofluorescence was used to analyze the differentiation of NSCs at the 7th day. The levels of apoptosis were detected by Western blotting and TUNEL. The mRNA level of brain derived neurotrophic factor (BDNF) was identified by real-time PCR. UV-vis and 1H NMR spectra confirmed that poly (L-lysine) was successfully grafted onto the silk fibroin film. From the 3rd day after seeding to the 7th day, the CCK-8 test showed that proliferation rate of NSCs in the Silk-PIL was significantly higher than Silk group (P<0.05) but had no significant difference compared with PLL group (P>0.05). Immunofluorescence staining displayed that more NSCs in Silk-PIL group were differentiated into neuron compared with Silk group (P<0.05), however, there was no significant difference compared with PLL group (P>0.05). The number of NSCs differentiated into astrocytes was not significantly different between the three groups. Western blotting and TUNEL test presented that the degree of apoptosis of NSCs in the Silk-PIL group was significantly lower than Silk group (P<0.05). RT-PCR exhibited that mRNA level of brain derived neurotrophic factor (BDNF) of NSCs was higher in Silk-PIL group compared with Silk group (P<0.05) but had no significant difference compared with PLL group (P>0.05). Thus, poly (L-lysine) modified silk fibroin film could promote the proliferation of NSCs and reduce NSCs apoptosis. Furthermore, it also can enhance the differentiation of NSCs into neurons. It is expected to become a new type of tissue engineering scaffold carrying NSCs to repair central nervous system injury.

Synthesis of barbell-like peptide dendrimer / 国际药学研究杂志

Objective To synthesize barbell-like peptide dendrimer. Methods Through the introduction of small molecule Fmoc-Gly-OH as the linker, NH2-CH2-COO-PEG2000-OOC-CH2-NH2 was obtained efficiently. And then N-α-N-ε-di-Fmoc-L-lysine and N-α-Fmoc-N-ε-Boc-L-lysine were used as branching agents, and barbell-like poly(ethylene glycol)-block-poly(L-lysine)dendrimer with a large number of surface amino groups was synthesized by the liquid-phase peptide synthesis method and divergent approach. Results and Conclusion The structure and relative molecular mass of the final products and intermediates were characterized and confirmed by 1H NMR and MS. The results revealed that barbell-like peptide dendrimer can be obtained by this method, which lays the foundation of its application in biological areas.

Synthesis of barbell-like peptide dendrimer / 国际药学研究杂志

Objective To synthesize barbell-like peptide dendrimer. Methods Through the introduction of small molecule Fmoc-Gly-OH as the linker,NH2-CH2-COO-PEG2000-OOC-CH2-NH2 was obtained efficiently. And then N-α-N-ε-di-Fmoc-L-lysine and N-α-Fmoc-N-ε-Boc-L-lysine were used as branching agents,and barbell-like poly(ethylene glycol)-block-poly(L-lysine)dendrimer with a large number of surface amino groups was synthesized by the liquid-phase peptide synthesis method and divergent approach. Re?sults and Conclusion The structure and relative molecular mass of the final products and intermediates were characterized and con?firmed by 1H NMR and MS. The results revealed that barbell-like peptide dendrimer can be obtained by this method,which lays the foundation of its application in biological areas.

Study on the preparation of ligand of hepatic asialoglycoprotein receptor / 国际生物医学工程杂志

Objective To optimize the preparation of high-efficiency galactocylated poly-L-lysine (Gal-PLL) ligand of the asialoglycoprotein receptor in liver, providing premise and foundation for upper preparation of ultrasound contrast agent of liver targeted nanoscale perfluorocarbon microballoon and the liver targeted molecular imaging. Methods Chemical reactions of reductive amination were carried out on group A and group B according to different proportions of reaction component. Each group was subdivided into three subgroups. In group A, three different molar ratios of D-galactose and poly-L-lysine (PLL) were compounded respectively with equivalent and sufficient reductant borohydride. In group B, identical molar ratios of D-galactose and PLL were compounded respectively with three unequal reductants borohydride. Products of each group were separated and purified by sephadex column to acquire different molecular weight distributions and the results were analyzed. Results In the condition of identical reductant, the peak curve of compound's molecular weight appeared earlier when D-galactose decreased properly. In the condition of identical molar ratio of D-galactose and PLL,the peak curve of compound's molecular weight appeared also earlier when reductant decreased properly. When the molar ratio of D-galactose and reductant was 1∶1, the peak curve of compound Gal-PLL and free components was more obvious, and the quantity of compound Gal-PLL increased to maximum. Conclusions In the condition of identical reductant, coupling effect of D-galactose and PLL increased when D-galactose decreased properly. In the condition of identical molar ratio of D-galactose and PLL, coupling effect was better when reductant decreased properly. When the molar ratio of D-galactose and reductant was 1∶1, coupling effect of them was the best. The coupling of D-galactose and PLL was related to not only the proportion of D-galactose and PLL, but also the proportion of D-galactose and reductant.

Biocompatibility of olfactory ensheathing cells and poly-L-lactic acid reinforced by chitosan in vitro / 中国组织工程研究

BACKGROUND:Previous studies have shown that composite scaffold of chitosan and poly-L-lactic acid has good biocompatibility with some cells. OBJECTIVE:To study the biocompatibility of poly-L-lactic acid reinforced by chitosan and olfactory ensheathing cells. METHODS:In experimental group, olfactory ensheathing cells from Sprague-Dawley rats aged 1-3 days were incubated onto chitosan-reinforced poly-L-lactic acid film. And in control group, olfactory ensheathing cells were co-cultured with poly-L-lysine. The proliferative ability of olfactory ensheathing cells was detected and the cells were observed with immunofluorescence histochemical staining at 1, 3, 5, 7 days after culture. RESULTS AND CONCLUSION:Olfactory ensheathing cells could survive on the chitosan-reinforced poly-L-lactic acid film, and the cytotoxic grade wasⅠ. Morphology of the cells in the experimental group was round or oval, with little processes and the cells aggregated into groups. One day after implantation, the periphery cells of the mass extended short projections and gradual y spread outward;3 days after implantation, the cells spread and most of the cells generated projections, most of which were bipolar or tri-polar;5 days after implantation, cel processes significantly extended, most cells were bipolar and tri-polar cells, while some were oval cells and irregular triangular cells;7 days after implantation, the cel density increased, and cel processes extended. Cel morphology of the control group had similar characteristics as the experimental group. There was no obvious difference between the control and the experimental group in number, perimeter or area of the cells (P>0.05). It showed that chitosan-reinforced poly-L-lactic acid had good biocompatibility with olfactory ensheathing cells.

Effects of poly-L-lysine of different molecular weights on growth of primary hippocampal neurons / 中华神经医学杂志

Objective To investigate the effects of poly-L-lysine of different molecular weights on the growth of primary hippocampal neurons in neonatal rats.Methods The hippocampal neurons from neonatal rats were prepared with the method of extrusion by needle core; and then,the hippocampal neurons were divided into three groups and respectively planted in culture plates where coated poly-L-lysine of different molecular weights (no poly-L-lysine,70 000-150 000 and 150 000-300 000).The neurons were maintained in Neurobasal-A medium without fetal bovine serum.The neurons were viewed at different time points and indentified by neuron specificity enolization enzyme immunofluorescence staining; the purity of the cells was calculated and the effects of poly-L-Lysine of different molecular weights on the growth of cells were observed.Results The neurons attached to the culture plates 1 h after the plantation.Four d after the plantation,the neurons had shining body,integrity stucture,having 2-3 synapses.Eight d after the plantation,the neurons became mature; the axons of neurons interweaved into the net; the cells were identified as neurons with an average purity of (92.6± 4.62)％.All neurons without poly-L-lysine almost died; the cells in the plates of poly-L-lysine of 70 000-150 000 distributed uniformly.Conclusion Different molecular weights of poly-L-lysine can affect the neurons adhesion and behavior; poly-L-lysine with large molecular weight (150 000-300 000) is most suitable for neurons.

Evaluation of umbilical cord mesenchymal stem cells labeling with superparamagnetic iron oxide nanoparticles coated with dextran and complexed with Poly-L-Lysine / Avaliação da marcação de células-tronco mesenquimais de cordão umbilical com nanopartículas superparamagnéticas de óxido de ferro recobertas com Dextran e complexadas a Poli-L-Lisina

Objetivo: O objetivo deste estudo foi avaliar o efeito da marcação de células-tronco mesenquimais obtidas da parede da veia do cordão umbilical com nanopartículas de óxido de ferro superparamagnéticas recobertas com dextran e complexadas a um agente transfector não viral denominado de Poli-L-Lisina. Métodos: A marcação das células-tronco mesenquimais foi realizada utilizando as nanopartículas de óxido de ferro superparamagnéticas recobertas com dextran complexadas e não complexadas a Poli-L-Lisina. As nanopartículas de óxido de ferro superparamagnéticas recobertas com dextran foram incubadas com o Poli-L-Lisina em um sonicador ultrassonico a 37ºC por 10 minutos, para a formação do complexo através de interação eletrostática. Em seguida, as células-tronco mesenquimais foram incubadas overnight com as nanopartículas de óxido de ferro superparamagnéticas complexadas e não com Poli-L-Lisina. Após o período de incubação as células-tronco mesenquimais foram avaliadas quanto à internalização do complexo nanopartícula de óxido de ferro superparamagnéticas /dextran/Poli-L-Lisina e nanopartícula de óxido de ferro superparamagnéticas /dextran através de ensaio citoquímico com azul de prússia. A viabilidade celular das célulastronco mesenquimais marcadas foi avaliada através do ensaio de proliferação celular utilizando o método de 5,6-carboxy-fluoresceinsuccinimidyl-ester e de morte celular através do método de anexinaiodeto de propídeo, ambos utilizando o recurso de citometria de fluxo. Resultados: Observamos nos ensaios citoquímicos que as célulastronco mesenquimais que foram marcadas com as nanopartícula de óxido de ferro superparamagnéticas /dextran sem a Poli-L-Lisina, não internalizaram com eficiência as nanopartículas devido pouca detecção de sua presença no interior das células. As células-tronco mesenquimais marcadas com o complexo nanopartícula de óxido de ferro superparamagnéticas /dextran/Poli-L-Lisina internalizaram com eficiência as nanopartículas devido à maior presença destas no interior das células. Os ensaios de viabilidade e morte celular demonstraram respectivamente que as células-tronco mesenquimais marcadas com as nanopartícula de óxido de ferro superparamagnéticas /dextran/Poli-L-Lisina continuam proliferando ao longo de sete dias e a porcentagem de células em apoptose inicial e tardia é baixa em relação à porcentagem de células vivas ao longo de três dias. Conclusão: Evidenciamos através de nossos resultados a necessidade da utilização da Poli-L-Lisina complexada com a nanopartícula de óxido de ferro superparamagnéticas /dextran para melhor internalização nas célulastronco mesenquimais. Paralelamente, demonstramos que este tipo de marcação não é citotóxico para as células-tronco mesenquimais já que os testes de morte e viabilidade celular mostraram que as células continuam vivas e proliferando.

Inhibition of polylysine-EDTA on rabbit lens epithelial cell growth in vitro / 中华实验眼科杂志

BackgroundPosterior capsule opacification(PCO) is the main cause inducing low vision after extacapsular cataract extraction. Our previous study determined that polylysine-ethylene diamine tetraacetic acid (EDTA) (PLE) can suppress the incidence of PCO. ObjectiveThe goal of this experiment was to investigate the inhibition of polylysine-EDTA on rabbit lens epithelial cells (LECs)proliferation in vitro and the effective concentrations of polylysine-EDTA. MethodsThe anterior capsular membranes from 10 3-month-old clean New Zealand white rabbits were digested and then cultured to obtain the LECs. The second and third generation of LECs were inoculated on the 96-hole culture plate with the cell density of the 1 × 105/ml. 12.5,25.0,50. 0,100. 0 μmol/Lof PLE were added into the culture medium for 48 hours respectively,and the DMSO medium was used at the same way as the control group. The proliferation of the LECs was then detected by MTT method and the inhibitory rate of PLE on LECs growth was calculated. ResultsLECs grew at a near normal state in ≤25.0 μmol/L PLE groups,however,cultured LECs were out of shape and the numbers decreased with the weakened adhesion ability in ≥50.0 μ mol/L PLE groups. The A490 values of LECs were 0. 278±0. 013,0. 266±0. 028,0. 260±0. 022 and 0. 247±0. 012 in 12. 5,25.0,50. 0, 100. 0 μmol/L polylysine-EDTA groups respectively and were lower than 0. 311 ±0. 038 of DMSO control group( P=0. 035,0. 011,0. 009,0.013 ). The inhibitory rates of 12. 5,25.0,50. 0, 100.0 μmoL/L PLE on LECs proliferation were 10.61％ , 14.47％ , 16.40％ and 20. 58％ respectively. ConclusionsPolylysine-EDTA can inhibit the growth and proliferation of LECs in vitro at a dose-dependent manner.

Physical Properties and Biocompatibility of Ba-Alginate-Poly-L-Ornithine-Alginate microcapsules / 中国康复理论与实践

@#ObjectiveTo test and verify whether Ba-alginate-Poly-L-Ornithine-Alginate microcapsules(B-PLO-A) can improve the physical properties and biocompatibility of the traditional BPA microcapsules.MethodsThe B-PLO-A and Ba-alginate-Poly-L-lysine-alginate（B-PLL-A） microcapsules were made by the static generator. The physical property of the microcapsules was evaluated by observing the morphological changes of the microcapsules in the hypotonic environment, changes in diameter of microcapsules in vitro culture and calculating broken microcapsules ratio by shaking method. The biocompatibility was observed by transplanting into peritoneal cavity of rat.ResultsB-PLO-A microcapsules are stronger and more stable in a hypotonic environment than B-PLL-A microcapsules. After 96 h mechanism shaking, the unbroken microcapsules ratio of B-PLO-A and B-PLL-A microcapsules were (99.3±1.0)% and (96.2±1.5)% respectively. The microcapsules were retrieved from peritoneal cavity of rat at 2, 4 and 8 weeks after transplantation, most of the microcapsules were of integrity, rotundity, and surface smooth without obviously bundled by connective tissue. 8 weeks after transplantation the intact microcapsules ratio of B-PLO-A and B-PLL-A microcapsules were (97.3±2.1)% and (95.4±2.4)% respectively.ConclusionB-PLO-A microcapsules as a whole have bettermechanical strength compared with B-PLL-A microcapsules, while maintaining a good biocompatibility.

Superparamagnetic iron oxide labeled mesenchymal stem cells and their magnetic resonance imaging in vitro / 第三军医大学学报

Objective To explore the optimal situation of labeling bone mesenchymal stem cells (BMSCs) with superparamagnetic iron oxides (SPIO) mediated by poly-L-lysine (PLL), and determine the most optimal protocol of magnetic resonance imaging according to the patterns of MR in vitro. Methods BMSCs were isolated from white rat and purified, incubated with SPIO-PLL complexes at the range of concentrations (0, 4.2, 8.4, 21, 42, 84 ?g Fe per ml medium). The labeling ratio and distribution of SPIO particles in BMSCs, and the morphological evidence of abnormal visualization were evaluated by Prussian blue staining, fluorescent microscope and electron microscopy. MTT growth curves and magnetic resonance imagings were obtained at the range of concentrations. Trypan blue exclusion test was performed to elevate the viability of BMSCs labeled with PLL at the range of concentrations (0, 0.05, 0.25, 0.5, 1.0, 5.0 ?g PLL per ml medium). Results The cellular labeling ratio was strongly correlated to the concentrations of SPIO (P

The Effect of Poly-L-lysine on Proliferation and Differentiation of Preadipocyte in the Alginate Gels

Alginate is widely used for scaffold in tissue engineering. However, it has a limitation of cell proliferation due to the lack of cell-to-matrix adhesion. Authors were trying to find out that the alginate gel become an efficient three-dimensional biomatrix in case of mixing with poly-L-lysine (PLL). After harvesting preadipocyte from rat epididymal fat, the proper concentration of PLL for an efficient cell culture was examined in the alginate gel and the level of proliferation of cells were measured in order to find out the efficacy of PLL for the experimental group(alginate/PLL mixed gel) compared to the control group(alginate gel without PLL). In addition, the number of surviving cell was counted and the fat cell stained with oil-red O was observed on the 21st day of the culture. The preadipocytes in the alginate gel were most viable in the PLL concentration of 50 microgram/ml. After 4 days in culture, the level of cell proliferation and the number of preadipocytes were significantly higher in the experimental group than those in the control group. A small number of fat cells stained with oil-red O were starting to be appeared on the 14th day and the larger number of cells on the 21st day of the culture in two groups. These results suggest that PLL increased the proliferation of preadipocyte in the alginate gel through the enhancement of cell-to-matrix adhesion. It also shows that alginate has the advantage of inducing the differentiation of preadipocyte in case of alginate/PLL mixed gel. In conclusion, alginate/PLL mixed gel is turned out to be effective for making a three-dimensional biomatrix.

The Cytotoxicity of Poly - L - lysine in Different Concentration to Preadipocytes Harvested from Living Rats

Authors are trying to prove the fact that Poly-L-lysine (PLL) might be mixed with alginate to enhance cell-to- matrix adhesion. Before that experiment, the proportion of preadipocytes in cells obtained from rat epididymal fat was detected, and PLL cytotoxicity on preadipocytes was measured. All cells harvested after third passage of culture were differentiated into adipocytes, and there was no decrease in the proliferation of preadipocytes in the culture media under the PLL concentration of 5 microgram/m4. These results suggest that all cells harvested from rat epididymal fat after 3rd passage of culture were preadipocytes and PLL has no cytotoxicity to preadipocytes of rats under the concentration of 5 microgram/mP.

THE EXPERIMENTAL STUDY ON CULTURE OF CHONDROCYTES SEEDED ONTO POLYGLYCOLIC ACID SCAFFOLDINGS COATING WITH DIFFERENT MATERIALS / 中华显微外科杂志

Objective:This experiment aims to find a appropriate scaffold for Tissue Engineer- ing.Methods:The chondrocytes were cultured when they were seeded onto PGA+PLA scaffolds coat- ing with Lecithin(LEC)and Poly-l-lysine(PLYS)together and respectively.With light microscope and scanning electron microscope,the observation of the scaffolds in hydrophilia and adsorptivity to chondrocytes and the function of the cells was made.Results:The PGA scaffolds coating with LEC and PLYS have better hydrophilia and adsorptivity to the cells;on which the chodrocytes produce more ma- trices.Conclusion:LEC can chang the hydrophilia of the scaffolds;while PLYS can strengthen the ad- sorptivity of the scaffolds;the PGA coating with LEC and PLYS is an ideal scaffold in Tissue Engineer- ing.

Lactoferrin as a gene delivery vehicle to hepatocytes

Using lactoferrin as the specific ligand, we developed a simplified method for preparation of molecular conjugate for gene delivery. Replacement of column chromatography and dialysis by one step centrifugal filtration (Centricon, cut off size : 30,000), resulted in the rapid purification of bovine lactoferrin/polylysine (bLf/pL) and human lactoferrin/polylysine (hLf/pL) conjugates and easy separation of unconjugated polylysine. The Lf/pL conjugates prepared by this method efficiently transferred the reporter genes, CAT and LacZ gene, to HeLa and hepatic cells. The bLf/pL and hLf/pL conjugates could transfer the reporter genes to various hepatocytes including primary mouse hepatocyte, Hepa 1-6, SK-Hep1 and Chang liver, but not to NIH 3T3 mouse fibroblast cells, indicating that the Lf/pL conjugates conferred hepatocyte-specific gene transfer. The bLf/pL and hLf/pL conjugates prepared in the present study exhibited higher transfection efficiencies for mouse and human hepatocytes than the commercially available transferrin/polylysine (Tf/pL) conjugate.

Distributional Characteristics of Sulphated Glycosaminoglycans in Normal, Edematous Nasal Mucosa and Nasal Polyp

Sulphated glycosaminoglycans (GAGs) are found on the vascular endothelial surface and in the extracellular matrix in various tissue and organs, suggesting that these materials constitute a negatively charged screen restricting the movement of circulating plasma molecules. The present study was designed to elucidate the ditributional characteristics of sulphated GAGs in normal mucosa, edematous nasal mucosa and nasal polyp in order to understand their roles in the formation of the nasal polyp. Their presence in nasal mucosa was lightmicroscopically detected with the histochemical method using poly-L-lysine conjugated colloidal gold followed by silver enhancement. Sulphated GAGs in normal and edematous inferior turbinate mucosa were distributed only on the vascular endothelial surface in the superficial layer, while in the deeper layer they were found on the extracellular matrix as well as vascular endothelial surface. Their expression in normal and edematous ethmoid sinus mucosa was restricted to the glandular secretory product, but not found on the endothelial surface and subepithelial extracellular matrix. Sulphated GAGs in nasal polyp tissue are quite variable in staining intensity or distributional pattern. These results suggest that the distributional pattern of sulphated GAGs in the nasal mucosa may be regionally different and play an important roles in the regulation of the vascular permeability of nasal mucosa.

The Influence of Polycationic Polypetride on the Transfective Efficacy of NonViral Vector / 中国肿瘤生物治疗杂志

Objective: Purpose to investigate the different in vitro function of targetable non-viral vector containing poly-L-lysine or protamine. Methods: Using GV1 and GV2 targetable non-viral vectors, the influences of the poly-L-lysine and protamine on in vitro gene transfer efficiency and the course of gene expression were observed. Results: ?-galactosidase was expressed at intermediate level (50% ) in A375 cells using a complex containing either protamine or poly-L-lysine. Howerver, in case of ABAE cells, ?－galactosidase expression level was low (20% ) transferred with a comPlex containing protamine. On the contrary, ?－ galactosidase expression was at high level (70% ) provided that protamine was replaced with poly-L-lysine. In addition, ?-galac- tosidase activity reached the peak at the 6th day after transfection with the complex containing protamine. The expression was not altered with subsequent subcultures, at least for 3 passages. Using poly-L-lysine, the expression peak in A375 reached the peak at the 7th day after transfection, but the level declined along with subsequent passages of cells. Conclusion: The apllication of protamine in VEGF receptor mediated gene delivery system was limited.

Effect of different substrates coated on spinal motor neurons from embryonic rat cultured in vitro / 中国免疫学杂志

Objective:To study the effect of different substrates coated on the cell survival and neurite outgrowth of spinal motor neurons (SMN) from embryonic rat cultured in vitro.Methods:The ventral spinal tissue was isolated from embryonic rats and digested into dissociated cell suspention for culture,then the cells were identified as SMN by immunohistochemistry stain.Poly-L-lysine(PLL) was dissolved into distilled water,phosphate-buffered saline solution,boric acid at 0.1 and 0.01 mol/L concentration respectively.The different substrates include PLL,collagen Ⅰ,laminin and PLL combined with laminin.Distilled water was used as control.The neuron survival numbers and the mean length of the neurites were measured and compared.Results:The cells on the PLL dissolved into boric acid at 0.01 mol/L concentration survived well.The SMNs grown on the PLL combined with laminin were in dispersed disitribution with high survival rate.Conclusion:PLL combined with laminin is the best for the study of the motor neuron including both soma and neurite.