Preparation and properties of hydrogels loaded with phage endolysin Lys84 / 生物工程学报

This study investigated the application of poly(N-isopropylacrylamide)-based interpenetrating network temperature-sensitive hydrogels (notation: IPNT) as the delivery vehicle for phage endolysin Lys84 and the potential of drug-loaded hydrogels as antimicrobial materials. Interpenetrating network temperature-sensitive hydrogels were prepared by free radical polymerization of sodium alginate and N-isopropylacrylamide. Drug-loaded hydrogels (IPNT-Lys84) were obtained by dry soaking method with the endolysin Lys84 of Staphylococcus aureus phage. The physical properties of the hydrogels with and without drug loading were characterized by infrared spectroscopy, scanning electron microscopy, and differential scanning calorimetry. The swelling and deswelling of the hydrogels as well as the release of endolysin Lys84 were investigated. Moreover, the antibacterial properties of IPNT-Lys84 hydrogels at different temperatures and concentrations of the drug solution were studied. The results showed that IPNT-Lys84 hydrogel had uniform pores and a low critical solubility temperature (LCST) of 32 ℃. The equilibrium swelling of the hydrogel was 30 g/g, and the water loss rate was 88% upon deswelling. The release rate of endolysin reached more than 70% within 6 h at 37 ℃. The bactericidal rate of IPNT-Lys84 hydrogel was over 99.9%. The research results showed the feasibility of using IPNT to deliver the endolysin Lys84, and IPNT-Lys84 hydrogel might be an effective antimicrobial material against multi-drug resistant Staphylococcus aureus.

Development of thermo-sensitive poly N-isopropyl acrylamide grafted chitosan derivatives.

In this study, the thermo-sensitive copolymers were synthesized by graft copolymerization of poly N isopropyl acrylamide (PNIPAm) onto chitosan (Ch) and aminated chitosan (AmCh) derivative. Free radical polymerization using potassium persulphate (KPS) as initiator was used for performing the grafting process. FTIR, TGA and DSC analysis were used to verify the structure of the resulting graft copolymers. The thermal sensitivity of the prepared copolymers was determined by monitoring the phase transition temperature at 550nm using a UV–VIS spectrophotometer. The results show that the transition of the PNIPAm-g-Ch is sharper than that of the PNIPAm-g-AmCh and the tr-ansition, in both cases, became sharper as increase the PNIPAm grating percentage.

Microgel-Encapsulated Methylene Blue for the Treatment of Breast Cancer Cells by Photodynamic Therapy / 한국유방암학회지

PURPOSE: Photodynamic therapy (PDT) is gaining increasing recognition for breast cancer treatment because it offers local selectivity and reduced toxic side effects compared to radiotherapy and chemotherapy. In PDT, photosensitizer drugs are loaded in different nanomaterials and used in combination with light exposure. However, the most representative issue with PDT is the difficulty of nanomaterials to encapsulate anticancer drugs at high doses, which results in low efficacy of the PDT treatment. Here, we proposed the development of the poly(N-isopropylacrylamide) (PNIPAM) microgel for the encapsulation of methylene blue, an anticancer drug, for its use as breast cancer treatment in MCF-7 cell line. METHODS: We developed biocompatible microgels based on nonfunctionalized PNIPAM and its corresponding anionically functionalized PNIPAM and polyacrylic acid (PNIPAM-co-PAA) microgel. Methylene blue was used as the photosensitizer drug because of its ability to generate toxic reactive oxygen species upon exposure to light at 664 nm. Core PNIPAM and core/shell PNIPAM-co-PAA microgels were synthesized and characterized using ultraviolet-visible spectroscopy and dynamic light scattering. The effect of methylene blue was evaluated using the MCF-7 cell line. RESULTS: Loading of methylene blue in core PNIPAM microgel was higher than that in the core/shell PNIPAM-co-PAA microgel, indicating that electrostatic interactions did not play an important role in loading a cationic drug. This behavior is probably due to the skin layer inhibiting the high uptake of drugs in the PNIPAM-co-PAA microgel. Core PNIPAM microgel effectively retained the cationic drug (i.e., methylene blue) for several hours compared to core/shell PNIPAM-co-PAA and enhanced its photodynamic efficacy in vitro more than that of free methylene blue. CONCLUSION: Our results showed that the employment of core PNIPAM and core/shell PNIPAM-co-PAA microgels enhanced the encapsulation of methylene blue. Core PNIPAM microgel released the drug more slowly than did core/shell PNIPAM-co-PAA, and it effectively inhibited the growth of MCF-7 cells.

Temperature-Responsive surface for novel co-culture systems of hepatocytes with endothelial cells: 2-D patterned and double layered co-cultures

We have developed two novel cell co-culture system, without any on cell type combination limitation, utilizing a polymer surface which is temperature-sensitive with respect to its cell adhesion characteristics. One system involves a patterned co-culture of primary hepatocytes with endothelial cells utilizing patterned masked of the electron-beam cured, temperature-responsive polymer, poly (N-isopropylacrylamide) (PIPAAm) by masked electron beam irradiation. Hepatocytes were cultured to confluency at 37 degrees C on these surfaces. When the culture temperature was reduced below 32 degrees C, cells detached from the PIPAAm-grafted areas without any need for trypsin. Endothelial cells were then seeded onto the same surfaces at 37 degrees C. These subsequently seeded endothelial cells adhered only to the now-exposed PIPAAm-grafted domains and could be co-cultured with the hepatocytes initially seeded at 37 degrees C in well-ordered patterns. The other system involves a double layered co-culture obtained by overlaying endothelial cell sheets of the designed shape onto hepatocyte monolayers. The endothelial cells adhered and proliferated on the PIPAAm-grafted surface, as on polystyrene tissue culture dishes at 37 degrees C. By reducing the temperature, confluent monolayers of cells detached from the PIPAAm surfaces without trypsin. Because the recovered cells maintaed intact cell-cell junctions together with deposited extracellular matrix, the harvested endothelial cell sheets, with designed shapes, were transferable and readily adhered to hepatocyte monolayers. Stable double layered cell sheets could be co-cultivated. These two co-culture methods enabled long-term co-culture of primary hepatocytes with endothelial cells. Hepatocytes so co-cultured with endothelial cells maintained their differentiated functions, such as albumin synthesis for unexpectedly long periods. These novel two co-culture systems offer promising techniques for basic biologic researches upon intercellular communications, and for the clinical applications of tissue engineered constructs.

Preliminary study on therapeutic effect of poly(N-isopropylacrylamide)adriamycin magnetic nanoparticles via transarterial embolization on liver VX2 tumor in rabbits / 介入放射学杂志

Objective To evaluate the therapeutic effects of poly(N-isopropylacrylamide)adriamycin magnetic nanoparticles(ADM-PNIPAM-Fe_3O_4)on liver VX2 tumor in rabbits via transcatheter arterial chemoembolization.Methods VX2 tumor pieces were successfully implanted into liver lobes of rabbits with liver tumors formation.All the animals were randomly divided into 4 groups of 8 each.Group A(control group)injected with 10 ml physiologic saline,Group B(ADM group)treated with free adriamycin (1 mg/kg)via arterial infusion,Group C(ADM-PNIPAM group)ADM-PNIPAM(1.5 mg/kg)was infused through arterial route,Group D(ADM-PNIPAM-Fe_3O_4+M group)ADM-PNIPAM-Fe_3O_4(2 mg/kg)was infused through arterial route;all were in individual doses,respectively;meanwhile an persistant magnet with intensity of 0.4 T was stabilized at the tumor region.Spiral CT scans were performed to measure size of liver tumors and evaluate lung metastasis at 1 day before operation and 14 days after operation.All experimental animals were sacrficed on the 15th days after operation and followed by pathologic and histologic examination of the tumor and lung specimens including changes befor and after the operation with correlative comparisons.Results There were no significant difference in volumes of tumors among 4 groups at 1 day before operation.The average tumor volume in the group A was(23.87?7.02)cm~3 at 14 days after operation;(7.70?1.53)cm3 in group B;(4.29?0.25)cm~3 in group C;(2.05?0.18)cm~3 in group D. The average tumor volumes in the group B,group C and group D were significantly smaller than that in the control group A at same time after operation and there was significant difference among the three experimental groups.According to the order of tumor sizes from small to large was as follows:group D＜group C＜group B＜group A.It showed that the average size of group D(ADM-PNIPAM-Fe_3O_4+M group) was the smallest among the 4 groups.Lung metastatic rates were 100%,66.7%,37.5% and 12.5% in group A,group B,group C and group D at 14 days after operation,respectively.Lung metastatic rates of group D was lower than that of group A,but there were no significant differences among group A,B and C.But pathological examination showed that there were lesser number of lung metastatic tumors in group B and C than that of group A .The intratumoral necrotic degrees showed as group D＞group C＞group B＞group A. Conclusions ADM-PNIPAM-Fe_3O_4 treatment for liver tumor via vascular interventional method combined with magnetic field localized in the lesion possesses significant inhibitory effect on tumor growth of liver VX2 tumor in rabbits.ADM-PNIPAM-Fe_3O_4 is thus initially confirmed as a kind of effective praeputium in interventional chemoembolization.