In Situ Fabrication of Nano-hydroxyapatite in a Macroporous Chitosan Scaffold for Tissue Engineering

A chitosan (CS)/hydroxyapatite (HAP) nanohybrid scaffold with high porosity and homogeneous nanostructure was fabricated through a bionic treatment combined with thermally-induced phase separation. The nano-HAP particles were formed in situ in the scaffold at room temperature instead of mechanically mixing the powders with the polymer component. The scaffold was macroporous with a pore size of about 100–136 μm. The nano-sized HAP particles with diameters of 90–200 nm were scattered homogeneously in the interactively connective pores. Both the improvement of the compressive modulus and yield strength of the scaffolds showed that the in situ nano-HAP particles reinforced the microstructure of the scaffold. The in vitro bioactivity study carried out in simulated body fluid (SBF) indicated good mineralization activity. The crystallization phenomenon suggested that the nano-HAP particles have positive impacts on directing apatite crystallization in the scaffold and led to the good bioactivity of the nanohybrid scaffold.     

Thiol-Containing Degradable Poly(thiourethane-urethane)s for Tissue Engineering

Poly(thiourethane-urethane)s with varying amounts of sulphur were synthesised by a two-step polycondensation consisting of the sequential addition of 1,6-hexamethylene diisocyanate and bis(2-mercaptoethyl) ether in a poly(ε-caprolactone) diol solution. Polymers prepared had high weight-average molecular weight and typical microdomains separation, as shown by size-exclusion chromatography and thermal analysis. Polymer surfaces were characterized by X-ray photoelectron spectroscopy and atomic force microscopy. The quantification of thiol groups at the surface was assessed using a fluorescent assay. Thiol concentration ranged between 7 and 14 nmol/cm, and was directly related to the amount of sulphur introduced in the polymerization and the macromolecule chains orientation at the surfaces. A preliminary in vitro degradation study and a proliferation assay were performed. The poly(thiourethane-urethane)s may have important applications as biodegradable and biocompatible materials for cartilage and bone tissue engineering. The surface thiol groups add the prospect of further functionalization. This is an important benefit compared to biodegradable poly(urethane)s that usually present low biological activity.     

Poly(ethylene glycol) hydrogels cross-linked by hydrolyzable polyrotaxane containing hydroxyapatite particles as scaffolds for bone regeneration

Poly(ethylene glycol) (PEG) hydrogels cross-linked by a hydrolyzable polyrotaxane containing hydroxyapatite particles (PRX-HAp) were developed as scaffolds for bone regeneration. Five scaffolds with various composition of the polyrotaxane, PEG and HAp particles were prepared to examine cell adhesion in vitro using rat primary cultured osteoblast. Cells were observed to attach well on a PRX-HAp that have the same weight ratio of the polyrotaxane and HAp particles at 7 days after seeding. These results indicate that HAp particles are necessary for cell adhesion and survival, but a higher ratio of the particles is not suitable for cell adhesion. The composites of rat osteoblast and the PRX-HAp were implanted subcutaneously in syngeneic rats and harvested at 5 weeks after implantation. In histological analysis, osteoblast-like cells became arrayed along the surface of the PRX-HAp, and osteoid-like tissues were observed in the region between a queue of osteoblast-like cells and PRX-HAp. These images are similar to intramembranous ossification, and it is expected that bone regeneration occurs on the surface of the PRX-HAp. This study strongly suggests the great potential of the PRX-HAp as scaffolds for bone regeneration.     

The use of poly(lactic-co-glycolic acid) microspheres as injectable cell carriers for cartilage regeneration in rabbit knees

The use of injectable scaffolding materials for in vivo tissue regeneration has raised great interest because it allows cell implantation through minimally invasive surgical procedures. Previously, we showed that poly(lactic-co-glycolic acid) (PLGA) microspheres can be used as an injectable scaffold to engineer cartilage in the subcutaneous space of athymic mice. The purpose of this study was to determine whether PLGA microspheres can be used as an injectable scaffold to regenerate hyaline cartilage in the osteochondral defects of rabbit knees. A full-thickness wound to the patellar groove of the articular cartilage was made in the knees of rabbits. Rabbit chondrocytes were mixed with PLGA microspheres and injected immediately into these osteochondral wounds. Both chondrocyte transplantations without PLGA microspheres and culture medium injections without chondrocytes served as controls. Sixteen weeks after implantation, chondrocytes implanted using the PLGA microspheres formed white cartilaginous tissues. Histological scores indicating the extent of the cartilaginous tissue repair and the absence of degenerative changes were significantly higher in the experimental group than in the control groups (P < 0.05). Histological analysis by a hematoxylin and eosin stain of the group transplanted with microspheres showed thicker and better-formed cartilage compared to the control groups. Alcian blue staining and Masson's trichrome staining indicated a higher content of the major extracellular matrices of cartilage, sulfated glycosaminoglycans and collagen in the group transplanted with microspheres than in the control groups. In addition, immunohistochemical analysis showed a higher content of collagen type II, the major collagen type in cartilage, in the microsphere transplanted group compared to the control groups. In the group transplanted without microspheres, the wounds were repaired with fibro-cartilaginous tissues. This study demonstrates the feasibility of using PLGA microspheres as an injectable scaffold for cartilage regeneration in a rabbit model of osteochondral wound repair.     

Development of Biodegradable Polyurethane Scaffolds Using Amino Acid and Dipeptide-Based Chain Extenders for Soft Tissue Engineering

The inherent flexibility of polyurethane (PU) chemistry allows the incorporation of specific chemical moieties into the backbone structure conferring a unique biological function to these synthetic polymers. We describe here the synthesis and characterization of a PU containing a Gly–Leu linkage, the cleavage site of several matrix metalloproteinases. A Gly–Leu dipeptide was introduced into the chain extender of the polyurethane through the reaction with 1,4-cyclohexane dimethanol. PUs synthesized with the Gly–Leu-based chain extender had a high weight-average molecular weight (M _w > 125 × 10³) and were phase segregated, semi-crystalline polymers with a low soft-segment glass-transition temperature (T _g < –50°C). Uniaxial tensile testing of PU films indicated that the polymer could withstand high ultimate tensile strengths (approx. 13 MPa) and were flexible with breaking strains of approx. 900%. The Gly–Leu PU had a significantly higher initial modulus, yield stress and ultimate stress compared to a PU previously developed in our laboratory containing a phenylalanine-based chain extender (Phe PU). The Gly–Leu-based chain extender allowed for better hard segment packing and hydrogen bonding leading to enhanced mechanical properties. Electrospinning was used to form scaffolds with randomly organized fibers and an average fiber diameter of approx. 3.6 μm for both the Gly–Leu and Phe PUs. Mouse embryonic fibroblasts were successfully cultured on the PU scaffolds out to 28 days. Further investigations into cell-mediated polymer degradation will help to identify the suitability of this new biomaterial as scaffolds for soft tissue applications.     

Effects of Poly(L-lysine), Poly(acrylic acid) and Poly(ethylene glycol) on the Adhesion,Proliferation and Chondrogenic Differentiation of Human Mesenchymal Stem Cells

Microenvironments, composed of many kinds of cytokines and growth factors plus extracellular matrices with diverse electrostatic properties, play key roles in controlling cell functions in vivo. In this study, three kinds of water-soluble polymers, positively charged poly(L-lysine) (PLL), negatively charged poly(acrylic acid) (PAAc) and neutral poly(ethylene glycol) (PEG), were compared based on their effects on the adhesion, spread, proliferation and chondrogenic differentiation of human mesenchymal stem cells (MSCs). The MSCs were seeded and cultured in the presence of polymers of different concentrations applied by methods using coating, mixing or covering. The effects of the water-soluble polymers depended on their electrostatic properties and method of application. The methods were in the order of coating, mixing and covering in terms of high to low influence. A low concentration of PLL promoted MSC adhesion, spread, proliferation and chondrogenic differentiation, while a high concentration of PLL was toxic. The PEG-coated surface facilitated cell aggregation and spheroid formation by inhibiting cell adhesion. A high concentration of mixed PEG (10 μg/ml) promoted cell proliferation in serum-free medium. PAAc showed no obvious effects on MSC adhesion, spread, proliferation, or chondrogenic differentiation.     

A Porous Hydroxyapatite/Gelatin Nanocomposite Scaffold for Bone Tissue Repair: In Vitro and In Vivo Evaluation

Abstract

In this study, a nano-structured scaffold was designed for bone repair using hydroxapatite and gelatin as its main components. The scaffold was prepared via layer solvent casting combined with freeze-drying and lamination techniques and characterized by the commonly used bulk techniques. The biocompatibility and osteoconductivity of this scaffold and its capacity to promote bone healing were also evaluated. Osteoblast-like cells were seeded on these scaffolds and their proliferation rate, intracellular alkaline phosphatase (ALP) activity and ability to form mineralized bone nodules were compared with those osteoblasts grown on cell culture plastic surfaces. Also, the scaffolds were implanted in a critical bone defect created on rat calvarium. Engineering analyses show that the scaffold posses a three dimensional interconnected homogenous porous structure with a porosity of about 82% and pore sizes ranging from 300 to 500 μm. Mechanical indices are in the range of spongy bones. The results obtained from biological assessment show that this scaffold does not negatively affect osteoblasts proliferation rate and improves osteoblasts function as shown by increasing the ALP activity and calcium deposition and formation of mineralized bone nodules. In addition, the scaffold promoted healing of critical size calvarial bone defect in rats. It could be concluded that this scaffold fulfills all the main requirements to be considered as a bone substitute.     

Novel Poly(L-lactide) PLLA/SWNTs Nanocomposites for Biomedical Applications: Material Characterization and Biocompatibility Evaluation

Poly(L-lactide) (PLLA)/single-walled carbon nanotubes (SWNTs) nanocomposite films were produced using the solvent casting method, and morphological, thermal and mechanical properties were investigated. Biocompatibility was evaluated by using human bone cells, performing adhesion and proliferation studies. The role of single-walled nanotube incorporation and functionalization on PLLA bio-polymers was investigated. Pristine (SWNTs) and carboxylated (SWNTs–COOH) carbon nanotubes were considered in order to control the interaction between PLLA and nanotubes. SWNTs and SWNTs–COOH showed a good dispersion in the polymer matrix and improved the PLLA crystallinity. Thermal, morphological and dynamic-mechanical analyses revealed that carboxylic groups on the tube sidewalls increased compatibility between PLLA and nanostructures. Mechanical properties demonstrated an enhancement related to introduction and functionalization of carbon nanotubes. Biological investigations showed osteoblasts cultured on PLLA/SWNTs–COOH nanocomposites has higher cell adhesion and proliferation than osteoblasts cultured on PLLA and PLLA/SWNTs nanocomposites. These studies suggest that combination of biodegradable polymers and SWNTs opens a new perspective in the self-assembly of nanomaterials and nanodevices for biomedical applications with tunable properties.     

Novel Scaffolds Based on Poly(2-hydroxyethyl methacrylate) Superporous Hydrogels for Bone Tissue Engineering

In this study, poly(2-hydroxyethyl methacrylate) (pHEMA)-based superporous hydrogels were synthesized by radical polymerization of 2-hydroxyethyl methacrylate (HEMA) in the presence of a gas blowing agent, sodium bicarbonate. These hydrogels are: pHEMA, pHEMA–gelatin, glycerol phosphate (GP) cross-linked pHEMA–gelatin, glutaraldehyde (GA) cross-linked pHEMA–gelatin superporous hydrogels (SPHs) and pHEMA–hydroxyapatite (HA) superporous hydrogel composites (SPHCs). The hydrogels have a structure of interconnected pores with pore sizes of approx. 500 μm. Although the extent of swelling decreased when gelatin and HA were incorporated to the pHEMA structure, the time to reach the equilibrium swelling (approx. 20 s) was not affected so much. In the presence of gelatin and cross-linkers, mechanical properties significantly improved when compared with pHEMA SPH. Among all the synthesized hydrogels, pHEMA–HA SPHC showed great improvement in mechanical strength and its elastic modulus value was 0.027±0.002 N/mm². Osteogenic activities of pHEMA-based scaffolds were examined by preosteoblastic MC3T3-E1 cell-culture studies. The mitochondrial activity test (MTT) showed that gelatin-containing scaffolds stimulated cell proliferation compared with other scaffolds, while alkaline phosphatase levels (ALP) and mineralization were found highest for the GP cross-linked pHEMA–gelatin SPH. However, pHEMA SPH and pHEMA–HA SPHC did not support cell proliferation and also differentiation. In conclusion, pHEMA–gelatin SPH and GP cross-linked pHEMA–gelatin SPH can be considered as potential scaffolds for bone tissue-engineering applications.     

Fibrin promotes proliferation and matrix production of intervertebral disc cells cultured in three-dimensional poly(lactic-co-glycolic acid) scaffold

Previously, we have proven that fibrin and poly(lactic-co-glycolic acid) (PLGA) scaffolds facilitate cell proliferation, matrix production and early chondrogenesis of rabbit articular chondrocytes in in vitro and in vivo experiments. In this study, we evaluated the potential of fibrin/PLGA scaffold for intervertebral disc (IVD) tissue engineering using annulus fibrosus (AF) and nucleus pulposus (NP) cells in relation to potential clinical application. PLGA scaffolds were soaked in cells–fibrin suspension and polymerized by dropping thrombin–sodium chloride (CaCl₂) solution. A PLGA–cell complex without fibrin was used as control. Higher cellular proliferation activity was observed in fibrin/PLGA-seeded AF and NP cells at each time point of 3, 7, 14 and 7 days using the MTT assay. After 3 weeks in vitro incubation, fibrin/PLGA exhibited a firmer gross morphology than PLGA groups. A significant cartilaginous tissue formation was observed in fibrin/PLGA, as proven by the development of cells cluster of various sizes and three-dimensional (3D) cartilaginous histoarchitecture and the presence of proteoglycan-rich matrix and glycosaminoglycan (GAG). The sGAG production measured by 1,9-dimethylmethylene blue (DMMB) assay revealed greater sGAG production in fibrin/PLGA than PLGA group. Immunohistochemical analyses showed expressions of collagen type II, aggrecan core protein and collagen type I genes throughout in vitro culture in both fibrin/PLGA and PLGA. In conclusion, fibrin promotes cell proliferation, stable in vitro tissue morphology, superior cartilaginous tissue formation and sGAG production of AF and NP cells cultured in PLGA scaffold. The 3D porous PLGA scaffold–cell complexes using fibrin can provide a vehicle for delivery of cells to regenerate tissue-engineered IVD tissue.     

DL—聚乳酸微球的体外和大鼠体内的降解研究

本文采用凝胶渗透色谱(GPC),观察DL—聚乳酸(PLA)降解时的分子量及分子量分布的变化,以及测定体外降解液的酸值和降解后PLA的失重、对两种不同分子量的PLA微球的体外降解和大鼠体内的降解进行了研究。实验结果表明:①PLA降解表现两个阶段,开始降解较快,随后趋向平缓;②分子量较大的比分子量较小的降解快;③PLA体内外降解行为基本一致。作者认为PLA的降解是通过酯键的水解起主导作用,酶不参与或在降解中不起主导作用。     

Chitosan,Gelatin and Poly(L-Lysine) Polyelectrolyte-Based Scaffolds and Films for Neural Tissue Engineering

Biomaterial implants are a promising strategy to replace neural tissue that is lost after traumatic nerve damage. Chitosan (Ch) is a suitable material for nerve implantation when it is used at a minimum amount of 2% (w/v). The goal of this study was to determine the best mixture of 2% Ch with gelatin (G) and poly(L-lysine) (PLL) for use in neural tissue engineering. Using different physicochemical approaches we showed that all mixtures formed polyelectrolyte complexes with distinct electrostatic interactions between their compounds. This gave rise to different gel morphologies, among which Ch + G exhibited a significantly smaller pore size, unlike Ch + G + PLL. However, thermal resistance to degradation and the wettability of the Ch-based films were not affected. Additionally, these differences affected glial cells growth in long-term (14 days) cultures performed on Ch-based films. Astrocytes and olfactory ensheathing cells proliferated on G and Ch + G films which induced both flattened and spindle cell morphologies. Meanwhile, cortical and hippocampal neurons were similarly viable in all studied films and significantly lower than those observed in controls. Lastly, neurites from dorsal root ganglia extended the most on Ch + G films. These results show that a Ch + G mixture is a promising candidate for use in neural tissue engineering.     

生物降解材料聚乳酸－聚乙二醇共聚物体内外降解

本文对ＤＬ－聚乳酸－聚乙二醇共聚物（ＰＥＬＡ）体内外降解进行了研究。采用凝胶渗透色层法（ＧＰＣ）测定分子量，扫描电镜（ＳＥＭ）观察。ＰＥＬＡ在最初第５天出现降解，２月时分子量丧失体内外分别为９７．２８％和９２．０％，而重量仅丧失８％。对其原因进行了讨论，同时将ＰＥＬＡ与ＤＬ－聚乳酸（ＰＬＡ）进行了比较，探讨分析了二者的降解机理。结果表明ＰＥＬＡ是一种新型生物降解材料。     

Pores Created by Laser Surface Modification of Poly(vinylalcohol)-Collagen with Glycosaminoglycan Scaffold for Cell Culture in Tissue Engineering

A PVA-GAG-COL composite scaffold is fabricated by polyvinyl alcohol (PVA),glycosaminoglycan (GAG) and collagen (COL).Laser surface modification technology is used to make holes on the surface of the sc...     

Paclitaxel and etoposide-loaded Poly (lactic-co-glycolic acid) microspheres fabricated by coaxial electrospraying for dual drug delivery

Abstract

In this study, we fabricated paclitaxel (PTX) and etoposide (ETP) loaded Poly (lactic-co-glycolic acid) (PLGA) microspheres with core–shell structures and particle sizes ranging from 1 to 4?µm by coaxial electrospraying. The microspheres were analyzed by scanning electron microscopy (SEM), transmission electron microscopy (TEM). The drug loading rate and entrapment efficiency of the microspheres were detected by high performance liquid chromatograph (HPLC). Moreover, the drug release profiles and degradation of drug-loaded PLGA microspheres in vitro were investigated, respectively. The distinct layered structure that existed in the manufactured core–shell microspheres can be observed by TEM. The in vitro release profiles indicated that the PLGA/PTX?+?ETP (PLGA/PE) microspheres exhibited the controlled release of two drugs in a sequential manner. Cell Counting Kit-8 was used to detect the toxic and side effects of the microspheres on bone tumor cells. PTX and ETP for combination drug therapy loaded microspheres had more cytotoxic effect on saos-2 osteosarcoma cells than the individual drugs. In conclusion, core–shell PLGA microspheres by electrospraying for combination drug therapy is promising for medicine applications, the PLGA/PE microspheres have some potential for osteosarcoma treatment.     

Synthesis and In Vitro Biocompatibility Assessment of a Poly(amic acid) Derived from Ethylenediaminetetraacetic Dianhydride

Poly(amic acid) (PAA) derived from ethylenediaminetetracetic dianhydride shows great potential as a biomaterial suitable for biomedical applications. To evaluate this polymer class further, in vitro cell toxicity (WST-1/ECS, ELISA based) and cell compatibility (cell adhesion and cell proliferation) tests were conducted to establish structure–toxicity relationships. PAAs with a number-average molecular weight ranging between 100 to 200 kg/mol were synthesized at 37°C after 24 h. Porcine radial artery cells (RACs) and descending aorta endothelial cells (ECs) were seeded independently in a 96-well cell culture plate at a cell density of 5000 cells/cm² to observe toxic effects. Similarly, RACs and ECs were seeded independently onto PAA coated and uncoated cover slips at a cell density of 7000 cells/cm² to observe growth patterns. Our results showed no toxicity after 96 h of incubation and in addition, both RACs and ECs adhered and proliferated on the PAA films, preserving their phenotype during this time. The tested synthetic material seems promising as a future biomaterial and should elicit a desired cellular response upon implantation.     

In Vitro and In Vivo Biocompatibility of Novel Zwitterionic Poly(Beta Amino)Ester Hydrogels Based on Diacrylate and Glycine for Site‐Specific Controlled Drug Release

New (β‐aminoester) hydrogels (PBAE) based on di(ethylene glycol)diacrylate and glycine are successfully synthesized and characterized for the first time in this work. PBAE macromers are obtained using Michael addition. By changing the diacrylate/amine stoichiometric ratio, but maintaining it >1, samples with different chemical structure containing acrylate end‐groups are obtained. The hydrogels are synthesized from macromers utilizing free radical polymerization. Chemical structure of macromers and hydrogels is confirmed by proton nuclear magnetic resonance, and Fourier transform infra‐red spectroscopy. Swelling and degradation rates in physiological pH range change notably with pH and monomer molar ratio, validating pH sensitivity and zwitterionic behavior, which can be finely tuned by changing any of these parameters. In vitro cytotoxicity and in vivo acute embryotoxicity in zebrafish (Danio rerio) performed to assess the biocompatibility of the novel hydrogel materials and their degradation products reveal that materials are nontoxic and biocompatible. The Cephalexin in vitro drug release study, at pH values 2.20, 5.50, and 7.40, demonstrates pH‐sensitive delivery with the release profiles effectively controlled by pH and the hydrogel composition. PBAE hydrogels exhibit great potential for a variety of biomedical applications, including tissue regeneration and intelligent drug delivery systems.     

Caprine (Goat) Collagen: A Potential Biomaterial for Skin Tissue Engineering

Collagens presently used in tissue engineering are primarily of bovine or porcine origin. However, a risk of a spongiform encephalopathy epidemic has limited the use of collagen from these sources. Keeping the aforementioned perspective in mind, we explored the possibility of using domestic goat available in the subcontinent as a potential source of collagen for tissue-engineering application. This article delineates the isolation, physico-chemical characterization, biocompatibility study and wound healing application of acid soluble caprine (goat) tendon collagen (GTC). Physico-chemical characterization of 1% acetic acid extracted GTC was done by SDS-PAGE, amino-acid composition analysis, FT-IR and CD spectroscopy. Results revealed that GTC was comprised of type-I collagen. Biocompatibility study showed that GTC augmented cell adhesion, cell cycle progression and proliferation. Immuno-cytochemical analysis in conjugation with traction force microscopy further confirmed a superior focal adhesion complex mediated cell–substrate interaction in GTC. Finally, in vivo study in mice model revealed that GTC has low immunogenicity and it augments healing process significantly. Throughout the study, calf skin collagen (CSC) was used as standard for comparative evaluation. In conclusion, it can be said that GTC may find its application as biomaterial in skin tissue engineering.     

Design,Synthesis and Evaluation of Cationic Poly(N-substituent acrylamide)s for In Vitro Gene Delivery

A series of poly(N-substituent acrylamide)s (PAms) that differ in alkylamine side-chain was synthesized via free radical polymerization. The PAms were designed to examine the effects of the methylene numbers (from 2 to 12) in the alkylamine side-chain on cytotoxicity, plasmid DNA (pDNA) binding affinity, cellular uptake efficiency and gene expression. The cytotoxicity of PAms evaluated in HEK293 cells using the MTT assay showed a trend of decreasing toxicity with increasing side-chain length and the IC₅₀ values of all PAms were lower than that of polyethylenimine (PEI) control. The primary amine-based polymers were able to efficiently condense pDNA to form complexes with size ranging from 100 to 350 nm. The gene transfection ability of PAms is dominantly determined by the specific side-chain length that P8Am (with an octylamine side-chain) reveals higher gene expression than other PAms containing the same backbone structure. Although the gene transfection efficiency of PEI was better than all of PAms, PAms were found not to be uptake-limited. This was supported by the effect of chloroquine on transfection activity, based on the protease inhibition activity of chloroquine. Especially, complexes formed from P8Am displayed high uptake level relative to PEI, which was attributed to the proper structure of P8Am to compact pDNA to form stable nanoparticles in the heparin replacement assay. The present study offers the understanding to polymer structure that influences the transfection ability and gives useful information when designs efficient polymeric gene carrier.     

Synthesis of Poly(ethylene glycol)-g-Chitosan-g-Poly(ethylene imine) Co-polymer and In Vitro Study of Its Suitability as a Gene-Delivery Vector

There are two main hindrances for the application of chitosan (CS) as a gene-delivery vector: poor water solubility and low transfection efficiency. To address these problems, we modified chitosan with poly(ethylene glycol) (PEG) and poly(ethylene imine) (PEI). As previously described, PEG was grafted onto CS by a reaction between the activated PEG and CS amine. This increased the solubility of CS in neutral or basic solution. Then, monomers of PEI (i.e., aziridine) were polymerized on the CS chain of the PEG(40k)-CS(50k) co-polymer obtained in the previous step. The resulting PEG-CS-PEI (PCP) co-polymer was characterized by ¹H-NMR, ¹³C-NMR and gel-permeation chromatography (GPC). It was found in the preliminary experiments that, amongst the series of PEG-CS-PEI co-polymers with various PEI molecular weights, PEG(40k)-CS(50k)-PEI(20k) was the most efficient one; therefore, it was chosen for the study. The PCP co-polymer showed lower cytotoxicity compared to PEI (25k) by MTT assay. Particle size and zeta potential of PCP/DNA complexes were measured by dynamic light scattering (DLS) and were shown to be predominantly affected by N/P ratios. PCP/DNA complexes at N/P ratio 20 were observed under a transmission electron microscope (TEM) as spherical particles with a mean diameter of about 50 nm. Plasmid DNA could be efficiently protected by PCP co-polymer from DNase I. The in vitro gene-transfection efficiency of PCP/pEGFP was higher than that of PEI(25k)/pEGFP and was markedly facilitated by serum.