Biomedical subjects

Yasuhiko Tabata

Publications and source records attributed to Yasuhiko Tabata.

At least 37 records · Page 2Linked to original sources

PubMed2006

Preparation of hybrid scaffold from fibrin and biodegradable polymer fiber.

A biodegradable hybrid scaffold was prepared from fibrin and poly(glycolic acid) (PGA) fiber. Mixed fibrinogen and thrombin solution homogeneously dispersed in the presence of various amounts (0, 1.5, 3.0, and 6.0mg) of PGA fiber was freeze-dried to obtain fibrin sponges with or without PGA fiber incorporation. By scanning electron microscopy observation, the fibrin sponges had an interconnected pore structure, irrespective of the amount of PGA fiber incorporated. PGA fiber incorporation enabled the fibrin sponges to significantly enhance their compression strength. In vitro cell culture studies revealed that the number of L929 fibroblasts initially attached was significantly larger for any fibrin sponge with PGA fiber incorporation than for the fibrin sponge without PGA fiber. The shrinkage of sponges after cell seeding was suppressed by fiber incorporation. It is possible that the shrinkage suppression of sponges maintains their intraspace, resulting in the superior cell attachment of a sponge with PGA fiber incorporation. After subcutaneous implantation into the backs of mice, the residual volume of a fibrin sponge with PGA fiber incorporation was significant compared with that of a fibrin sponge without PGA fiber. Larger number of cells infiltrated deep inside the fibrin sponges with PGA fiber incorporation implanted subcutaneously. It is concluded that the fibrin sponge reinforced by fiber incorporation is a promising three-dimensional scaffold of cells for tissue engineering.

Absorbable Implants↗

PubMed2006

Periurethral injection of sustained release basic fibroblast growth factor improves sphincteric contractility of the rat urethra denervated by botulinum-a toxin.

PURPOSE: We evaluated the effects of sustained release basic fibroblast growth factor injection in rat urethra denervated by botulinum-A toxin (Wako Life Science, Osaka, Japan). MATERIALS AND METHODS: A total of 30 female Sprague-Dawley rats underwent periurethral injection of 10 U botulinum-A toxin to induce chemical denervation of the urethral sphincter. Leak point pressure in the waking state was determined and a significant decrease in leak point pressure vs that in control rats was confirmed (mean +/- SD 58.7 +/- 6.2 vs 120.7 +/- 13.0 cm H(2)O, p <0.0001). Two weeks later 0, 50 and 200 microg basic fibroblast growth factor incorporating 200 microl gelatin hydrogels in 10 rats each were injected into the urethral sphincter, enabling sustained release of basic fibroblast growth factor for 2 weeks. Four weeks later injection leak point pressure measurement and histological evaluation of the urethra were performed. RESULTS: Leak point pressure in rats with 50 and 200 microg basic fibroblast growth factor injection was significantly higher than in rats with the 0 microg injection (82.7 +/- 9.0 vs 95.1 +/- 6.2 and 119.3 +/- 8.1 cm H(2)O, p = 0.0021 and <0.0001, respectively). Maximum cross-sectional area of the urethral smooth muscle layer in the 50 and 200 microg groups significantly increased compared with that in the urethra in the 0 micro group, which was considered 100% (114.1% +/- 15.8% and 132.5% +/- 13.4%, p = 0.029 and <0.0001, respectively). Similarly the cross-sectional area of the striated sphincter in the 50 and 200 microg groups was greater than the 100% in the 0 microg group (112.3% +/- 15.6% and 124.3% +/- 14.1%, p = 0.069 and 0.0007, respectively). Vascular density in the urethral peri-atrophic zone in the 50 and 200 microg groups was significantly higher than in the 0 microg group (p = 0.027 and <0.0001, respectively). CONCLUSIONS: Sustained release basic fibroblast growth factor injection in the chemically denervated urethral sphincter facilitates regeneration of the urethral muscles and improves sphincteric contractility. Endoscopic periurethral injection of basic fibroblast growth factor incorporating gelatin hydrogels may be an attractive therapy for stress urinary incontinence.

Animals↗

PubMed2006

The wrapping method using biodegradable felt strips has a preventive effect on the thinning of the aortic wall: experimental study in the canine aorta.

OBJECTIVES: Wrapping methods have been widely used to reinforce the anastomotic site in vascular surgery; however, postoperative changes in the aortic wall wrapped by nonbiodegradable felt have not been well characterized. The purposes of this investigation are to elucidate the sequelae of wrapping with nonbiodegradable felt on the aortic wall and to modify those changes by using biodegradable felt with or without basic fibroblast growth factor (bFGF). METHODS: The descending thoracic aortas of 15 beagles were wrapped with three different materials: nonbiodegradable polytetrafluoroethylene (PTFE) felt, biodegradable polyglycol acid (PGA) material, and PGA with 100 microg bFGF (n = 5 in each group). The descending thoracic aorta was resected after 3 months. The thickness of the aortic wall, vessel density in the media and the adventitia, and the wall strength were assessed. Untreated native aortic wall served as a normal control. RESULTS: The thickness of the media of the PTFE group was lower than that of the PGA + bFGF group (66% +/- 5% vs 85% +/- 6% of control, P < .05). The adventitia-media ratio in the PTFE group decreased compared with controls (59.1% of normal, P < 0.05), whereas those in the PGA and PGA + bFGF groups increased (172.1% and 189.6% of normal, respectively, P < .01). The collagen-smooth muscle ratio in the media was higher in the PTFE group than in the controls (0.14 +/- 0.02 vs 0.07 +/- 0.01, P < .01). The number of vessels in the adventitia was higher in the PGA + bFGF group than those in PTFE or PGA groups (29.6 +/- 2.5/mm2 vs 6.4 +/- 0.8/mm2, 19.0 +/- 1.1/mm2, P < .01). The PGA + bFGF group demonstrated larger failure force than the PTFE group (4.0 +/- 0.3 kgf vs 1.6 +/- 0.3 kgf, P < .01). The failure stress in the PGA and PGA + bFGF groups was larger than that in PTFE group (PTFE:PGA + bFGF = 5.3 +/- 0.9 x10(2) kPa:11.7 +/- 1.7 x 10(2) kPa, P < .01; PTFE:PGA = 5.3 +/- 0.9 x 10(2) kPa:11.2 +/- 1.2 x 10(2) kPa, P < .05). CONCLUSION: The aortic wall wrapped with nonbiodegradable PTFE felt showed a reduced thickness and diminished vessels in the adventitia. Biodegradable felt (PGA), with or without bFGF, modified these histologic changes. The vessel-rich thickened adventitia, after wrapping by PGA with bFGF, was associated with increased aortic wall strength. CLINICAL RELEVANCE: This investigation was conducted in an attempt to elucidate mechanisms underlying the occurrence of late postoperative false aneurysm after aortic surgery. We hypothesized that sustaining compression of the aorta by the felt strip may cause structural derangement and local ischemia on the aortic wall. We used a simple wrapping of the aorta with a felt strip rather than a felt strip at anastomotic sites to simplify the experimental model and to exclude confounding factors brought by technical inconsistency on the surgical anastomosis. We further attempted to find a clue for preventing adverse effects of wrapping with a conventional felt strip. Practically, we pursued a possible application of a biodegradable felt strip to aortic wrapping in our experimental model before we proceed in a clinical application of the new material.

Aneurysm, False↗

PubMed2006

Sustained-release vancomycin sheet may help to prevent prosthetic graft methicillin-resistant Staphylococcus aureus infection.

OBJECTIVE: The purpose of this study was to evaluate the effectiveness of a sustained-release sheet with vancomycin (VCM) to prevent prosthetic graft infection. METHODS: VCM was incorporated into a poly-L-lactide-co-caprolactone (PLCA) sheet (VCM-PLCA). The release profile of VCM from the VCM-PLCA sheet and the tissue concentration of VCM released in vivo were examined. To assess the antibacterial effect of the VCM-PLCA sheet, a sterile Dacron sheet was implanted into 96 male mice (C57BL/6), who were randomly divided into four groups of 24 each and treated as follows: no treatment (group A, control group), a local bolus injection of an aqueous solution of VCM (group B), a plain PLCA sheet (group C), and a VCM-PLCA sheet (group D). After the treatment, methicillin-resistant Staphylococcus aureus (MRSA) (1 x 10(6) colony forming units) was inoculated onto the Dacron graft surface. The Dacron grafts were retrieved on days 3, 7, 10, and 14 after the implantation, and the number of MRSA in the Dacron grafts was counted. RESULTS: VCM was slowly released from the VCM-PLCA sheet over 2 weeks in vivo, and the mean in vivo concentrations of VCM in the tissue around a VCM-PLCA sheet were 7.95, 26.39, 13.87, 12.51, 8.36, and 10.33 mug/mL (the minimum inhibitory concentration of VCM against MRSA is 2.0 mug/ml), at 1, 2, 5, 7, 10, and 14 days after the implantation, respectively. MRSA colonization on the cultivated agar plates was detected in all samples from groups A, B, and C at any postoperative time points. In contrast, some samples were negative for bacterial cultures in group D (2, 3, 1, and 2 samples out of 6 samples each on days 3, 7, 10, and 14 after the implantation, respectively). At all time points, the number of MRSA bacteria in the implanted Dacron graft in group D was by far the lowest (P < .01 at each time point). CONCLUSIONS: The sustained-release sheet with VCM appears to be effective for the reduction of subcutaneous prosthetic graft infection.

Animals↗

PubMed2006

Effect of charge and molecular weight on the functionality of gelatin carriers for corneal endothelial cell therapy.

Cell transplantation strategies usually involve the use of supporting carrier materials because of the soft and fragile nature of these grafts. In this work, a cell-adhesive gelatin hydrogel carrier was fabricated to deliver cultivated human corneal endothelial cell (HCEC) sheets, which were harvested from thermo-responsive poly(N-isopropylacrylamide) (PNIPAAm)-grafted culture surfaces. The carrier disks, consisting of gelatins with a different isoelectric point (IEP = 5.0 and 9.0) and a molecular weight (MW) ranging from 3 to 100 kDa, were subjected to 16.6 kGy gamma irradiation for sterilization. The effect of IEP and MW of the raw gelatins (i.e., before irradiation) on the functionality of sterilized disks was studied by determinations of mechanical property, water content, dissolution degree, and cytocompatibility. Irrespective of the IEP of raw gelatin, hydrogel disks prepared with high MW (100 kDa) exhibited a greater tensile strength, lower water content, and slower dissolution rate than those made of low MW gelatin (8 and 3 kDa). From the investigation of cellular responses to the disks, the negatively charged gelatin (IEP = 5.0) groups were more cytocompatible when compared with their positively charged counterparts (IEP = 9.0) at the same MW (100 kDa). Additionally, in the negatively charged gelatin groups, only a slight increase in pro-inflammatory cytokine expression was observed with increasing MW of gelatin from 3 to 100 kDa. It is concluded that the gamma-sterilized hydrogel disks made from raw gelatins (IEP = 5.0, MW = 100 kDa) with appropriate dissolution degree and acceptable cytocompatibility are capable of providing stable mechanical support, making these carriers promising candidates for intraocular delivery of cultivated HCEC sheets.

Animals↗

PubMed2006

Regenerative inductive therapy based on DDS technology of protein and gene.

Recent development of biomedical engineering including biomaterials and drug delivery system (DDS) as well as basic biology and medicine has enabled cells to induce regeneration repairing of defective tissues as well as substitute the biological functions of damaged organs. For successful tissue regeneration, it is undoubtedly indispensable to give cells a local environment which allows cells to efficiently promote their proliferation and differentiation and consequently induce cell-based tissue regeneration. Tissue engineering is one of the biomedical forms to create this regeneration environment of cells. The tissue and organ repairing based on their regeneration induction has been realized by combining cells with the tissue engineering technology or methodology in a surgical or internally medical manner. This paper overviews the present status and future direction of tissue engineering for regenerative inductive therapy, briefly explaining the key technology of tissue engineering, especially DDS of growth factor and gene.

Animals↗

PubMed2006

Enhanced bone regeneration at a segmental bone defect by controlled release of bone morphogenetic protein-2 from a biodegradable hydrogel.

The objective of this study is to investigate the feasibility of a biodegradable hydrogel of gelatin as the controlled release carrier of bone morphogenetic protein-2 (BMP-2) suitable for enhancement of bone regeneration at a segmental bone defect. Hydrogels with three different water contents were prepared through glutaraldehyde crosslinking of gelatin with an isoelectric point of 9.0 under varied reaction conditions. Segmental critical-sized defects (20 mm) were created at the ulnar bone of skeletally mature New Zealand white rabbits, and gelatin hydrogels incorporating BMP-2 (17 microg/hydrogel) were implanted into the defects. When bone regeneration was evaluated by soft x-ray observation and bone mineral density (BMD) measurement, the gelatin hydrogels incorporating BMP- 2 exhibited significantly high osteoinduction activity compared with that of free BMP-2, although the activity depended on the water content of the hydrogels. Significantly higher BMD enhancement was observed in the gelatin hydrogel with a water content of 97.8 wt% than that with the lower or higher water content. We concluded that the biodegradable gelatin hydrogel is a promising controlled release carrier of BMP-2 for bone regeneration at the segmental bone defect.

Absorbable Implants↗

PubMed2006

In situ regeneration of adipose tissue in rat fat pad by combining a collagen scaffold with gelatin microspheres containing basic fibroblast growth factor.

This study is an investigation to evaluate in situ adipose tissue regeneration in fat pads. Gelatin microspheres with different water contents were prepared for the controlled release of basic fibroblast growth factor (bFGF). After a collagen sponge scaffold was incorporated by the microspheres containing 0, 0.01, 0.1, 1, and 10 microg of bFGF with or without syngeneic rat preadipocytes (1 x 10(5) cells/site) into a defect of rat fat pad, adipogenesis at the implanted site of scaffold was evaluated histologically. in situ formation of adipose tissue accompanied with angiogenesis was observed in the scaffold implanted with the microspheres containing 1.0 microg of bFGF, although the extent was less at the lower and higher bFGF doses. The in situ formation induced by the microspheres containing bFGF was significantly higher than that induced by free bFGF of the same dose. Adipogenesis was enhanced with time after implantation up to 4 weeks and thereafter leveled off. Such in situ adipogenesis was reproducibly induced by implantation of collagen scaffold incorporating gelatin microspheres containing 1 microg of bFGF, whereas addition of rat syngeneic preadipocytes did not promote the adipogenesis. The degradation of microspheres and the consequent FGF release became faster with an increase in the water content of gelatin microspheres. Less in situ formation of adipose tissue was observed at the lower water content of microspheres, which showed longer-term bFGF release. We conclude that combination of scaffold collagen with an appropriate controlled release of bFGF was essential to achieve the in situ formation of adipose tissue even without preadipocytes.

Adipose Tissue↗

PubMed2006

Cochlear protection by local insulin-like growth factor-1 application using biodegradable hydrogel.

OBJECTIVE: The aim of this experimental study was to examine the potential of local recombinant human insulin-like growth factor-1 (rhIGF-1) application through a biodegradable hydrogel for the treatment of cochleae. METHODS: A hydrogel immersed with rhIGF-1 was placed on the round window membrane of Sprague-Dawley rats while a hydrogel immersed with physiological saline was applied to control animals. On day 3 after drug application, the animals were exposed to white noise at 120 dB sound pressure level (SPL) for 2 hours. Cochlear function was monitored using measurements of auditory brain stem responses (ABRs) at frequencies of 8, 16, and 32 kHz. The temporal bones were collected 7 or 30 days after noise exposure and the loss of hair cells was quantitatively analyzed. RESULTS: Local rhIGF-1 treatment significantly reduced the elevation of ABR thresholds on days 7 and 30 after noise exposure. Histologic analysis revealed that local rhIGF-1 treatment significantly prohibited the loss of outer hair cells. CONCLUSIONS: These findings demonstrate that local IGF-1 application through the biodegradable hydrogel has the potential for protection of cochleae from noise trauma.

Administration, Topical↗

PubMed2006

Pluronic F127: application in arterial embolization.

PURPOSE: Pluronic is a substance that is widely used in medical and pharmaceutical fields. In particular, 20% Pluronic F127 solution is a unique substance that is liquid at less than 15 degrees C and gelatinous at 25 to 60 degrees C. In this study, the authors took advantage of the gelation property of Pluronic F127 at human body temperature to simulate embolization and dissolution of the embolism in the renal artery and the superior mesenteric artery (SMA) using a rabbit model. MATERIALS AND METHODS: Four female Japanese rabbits (weight, 2.5-3 kg each) were used. The renal artery was fitted with a 4-F cobra-type catheter and embolized with a 20% Pluronic F127 solution at a temperature of 20 degrees C. The embolic effect was evaluated by angiography immediately after the initial injection and every 15 minutes for 2.5 hours after embolization. After 24 hours, pathologic changes of the renal parenchyma were also evaluated. The embolic effect for SMA and ischemic changes of the intestine were evaluated in the same manner. RESULTS: Angiographic findings showed that Pluronic F127 caused embolization immediately after injection and dissolved in the renal artery and the SMA after 90 to 120 minutes. The pathologic findings showed no ischemic change in the renal parenchyma. Necrosis was not found in the intestine, but focal hemorrhagic changes were extensively present when the gel had dissolved. This suggested that Pluronic F127 dissolved before severe tissue damage could occur. CONCLUSION: Pluronic F127 can potentially be used as a temporary embolic material.

Angiography↗

PubMed2006

Collagen-poly glycolic acid hybrid matrix with basic fibroblast growth factor accelerated angiogenesis and granulation tissue formation in diabetic mice.

Because poor skin wound healing associated with diabetes is thought to be partly a result from impaired angiogenesis, treatments that improve angiogenesis could have important clinical applications. We herein report the effects of novel developed material, collagen-poly glycolic acid fiber hybrid matrix, being used together with basic fibroblast growth factor to promote wound healing of full-thickness skin defects on the back of type 2 diabetic Lepr(db) mice. Our data indicates that this therapeutic approach markedly promotes angiogenesis and granulation tissue formation in comparison with other conditions 14 days after wounding.

Animals↗

PubMed2006

Cationized gelatin delivery of a plasmid DNA expressing small interference RNA for VEGF inhibits murine squamous cell carcinoma.

Double-stranded RNA (dsRNA) plays a major role in RNA interference (RNAi), a process in which segments of dsRNA are initially cleaved by the Dicer into shorter segments (21-23 nt) called small interfering RNA (siRNA). These siRNA then specifically target homologous mRNA molecules causing them to be degraded by cellular ribonucleases. RNAi down regulates endogenous gene expression in mammalian cells. Vascular endothelial growth factor (VEGF) is a key molecule in vasculogenesis as well as in angiogenesis. Tumor growth is an angiogenesis-dependent process, and therapeutic strategies aimed at inhibiting angiogenesis are theoretically attractive. To investigate the feasibility of using siRNA for VEGF in the specific knockdown of VEGF mRNA, thereby inhibiting angiogenesis, we have performed experiments with a DNA vector based on a siRNA system that targets VEGF (siVEGF). It almost completely inhibited the expression of three different isoforms (VEGF120, VEGF164 and VEGF188) of VEGF mRNA and the secretion of VEGF protein in mouse squamous cell carcinoma NRS-1 cells. The siVEGF released from cationized gelatin microspheres suppressed tumor growth in vivo. A marked reduction in vascularity accompanied the inhibition of a siVEGF-transfected tumor. Fluorescent microscopic study showed that the complex of siVEGF with cationized gelatin microspheres was still present around the tumor 10 days after injection, while free siVEGF had vanished by that time. siVEGF gene therapy increased the fraction of vessels covered by pericytes and induced expression of angiopoietin-1 by pericytes. These data suggest that cationized-gelatin microspheres containing siVEGF can be used to normalize tumor vasculature and inhibit tumor growth in a NRS-1 squamous cell carcinoma xenograft model.

Angiopoietin-1↗

PubMed2006

Non-viral vectors for cancer therapy.

Cancers are diverse and often resistant to therapeutic strategies. Gene therapy has yet to meet the promise of a breakthrough in cancer therapy. There are several barriers to overcome in cancer gene therapy. One of the biggest challenges is the design of appropriate vectors. Numerous viral and non-viral methods for gene transfer have been developed for human gene therapy, but both viral and non-viral vectors have limitations and advantages. In this review article, recent improvements in the development of non-viral vectors for delivering gene therapy for the treatment of cancer will be discussed.

Animals↗

PubMed2006

Targeted chemotherapy against intraperitoneally disseminated colon carcinoma using a cationized gelatin-conjugated HVJ envelope vector.

The hemagglutinating virus of Japan envelope (HVJ-E; Sendai virus) vector derived from inactivated HVJ particles can be used to deliver DNA, proteins, and drugs into cells both in vitro and in vivo. HVJ-E is capable of delivering bleomycin, an anticancer drug, to various cancer cell lines, thereby producing 300-fold greater cytotoxicity than administration of bleomycin alone. In a mouse model of peritoneally disseminated colon cancer, we injected HVJ-E containing the luciferase gene into the peritoneum. Unexpectedly, luciferase gene expression was not observed within the tumor deposits or any organs. However, when combined with cationized gelatin (CG), CG-HVJ-E produced a high level of luciferase gene expression primarily within the tumor deposits. Forty-eight hours after introducing colon cancer cells into the peritoneum of experimental mice, CG-HVJ-E with or without bleomycin was injected into the abdominal cavity. Following six injections of bleomycin-incorporated CG-HVJ-E, complete responses were observed in 40% of the mice examined. All of the mice that received either empty CG-HVJ-E or bleomycin alone died within 40 days of having cancer cells introduced into the peritoneum. When the mice with complete responses were rechallenged with colon cancer cells from the same cell line, no tumors developed. Thus, CG-HVJ-E may suppress peritoneal dissemination of cancer.

Animals↗

PubMed2006

Affinity evaluation of gelatin for hepatocyte growth factor of different types to design the release carrier.

The objective of this study was to investigate the physicochemical interaction of hepatocyte growth factor (HGF) and its variant with 5 amino-acid residues deleted (dHGF) with an acidic gelatin for the design of factors release from the gelatin hydrogel. When the interaction of HGF or dHGF with gelatin-immobilized agarose beads was evaluated by Scatchard binding assay, the dissociation constant of dHGF was higher than that of HGF, although the two proteins had a similar binding ratio. dHGF was released more rapidly from the hydrogel of acidic gelatin than HGF. In vivo release study with 125I-labeled HGF or dHGF in mice subcutis showed that HGF was released from the gelatin hydrogel as a result of hydrogel degradation. In contrast, dHGF was rapidly released by a simple diffusion from the gelatin hydrogel. From electrophoresis experiments, mixing with the acidic gelatin enabled HGF to complex and suppressing the trypsin-digested molecular weight loss, in marked contrast to that of dHGF. In addition, the percentage of HGF recognized by the antibody was reduced by the gelatin complexation, but that of dHGF was not. We conclude that unlike dHGF, HGF has a strong affinity for the acidic gelatin, resulting in the controlled release of HGF accompanied with hydrogel degradation of the release carrier.

Animals↗

PubMed2006

Interaction of hepatocyte growth factor with gelatin as the carrier material.

The objective of this study was to physicochemically investigate the interaction between hepatocyte growth factor (HGF) and acidic gelatin compared with that between HGF and basic gelatin or heparin. Gelatin- or heparin-immobilized agarose beads were prepared and HGF interaction with them was evaluated by Scatchard binding assay. The dissociation constant of HGF with the acidic gelatin was about 2-3 orders of magnitude higher than that of heparin. The cell proliferation assay revealed that the proliferation promotion activity of HGF complexed with the acidic gelatin was detected, although it was lower than that of original HGF. The ability of HGF to enhance the cell proliferation was reduced by the trypsin treatment, although the extent of the reduction was significantly suppressed by HGF complexation with acidic gelatin. Electrophoresis experimentally confirmed enhanced resistance to the molecular mass loss of HGF by gelatin complexation. Moreover, the recognized level of an antibody to HGF was reduced by the complexation with the acidic gelatin, indicating that the acidic gelatin is present around HGF molecules. It is possible that the HGF molecule is covered with the acidic gelatin, resulting in protection from enzymatic digestion.

Antibodies↗

PubMed2006

In vitro transfection of plasmid DNA by cationized gelatin prepared from different amine compounds.

The objective of this paper is to compare the in vitro transfection efficiency of a luciferase plasmid DNA using cationized gelatin prepared from different amine compounds. The compounds used here were ethylenediamine, putrescine, spermidine and spermine, chemically introduced to the carboxyl group of gelatin for the cationization. Complexation of the cationized gelatin with the plasmid DNA was performed by simply mixing the two materials at various N+/P- mixing ratios (the molar number ratio of amino groups of gelatin to the phosphate groups of DNA) in aqueous solution. Gel retardation studies revealed that the formation of cationized-gelatin-plasmid DNA complexes depended on the N+/P- mixing ratio. The stronger interaction of plasmid DNA with the cationized gelatin of spermine compared to the other cationized gelatins was observed by an ethidium bromide intercalation assay and Scatchard binding analysis. When the transfection efficiency of plasmid DNA complexed with the various cationized gelatins at different N+/P- mixing ratios was evaluated for mouse L929 fibroblasts, the highest transfection efficiency was observed for the complex prepared from the cationized gelatin of spermine at a N+/P- mixing ratio of 2. The present study indicates that there is an optimal N+/P- mixing ratio and a type of amine compound or cationization extent of cationized gelatin to enhance the transfection efficiency of plasmid DNA.

Amines↗

PubMed2006

Self assembly of DNA nanoparticles with polycations for the delivery of genetic materials into cells.

Increasing attention has been paid to technology used for the delivery of genetic materials into cells for gene therapy and the generation of genetically engineered cells. So far, viral vectors have been mainly used because of their inherently high transfection efficiency of gene. However, there are some problems to be resolved for the clinical applications, such as the pathogenicity and immunogenicity of viral vectors themselves. Therefore, many research trials with non-viral vectors have been performed to enhance their efficiency to a level comparable to the viral vector. Two directions of these trials exist: Material improvement of non-viral vectors and their combination with various external physical stimuli. In this study gelatin was selected as a non-viral carrier for DNA. To give a positive charge to gelatin, different extents introduction of ethylenediamine (Ed), spermidine (Sd), and spermine (Sm) were reacted with gelatin in the presence of a water-soluble carbodiimide. When positively charged gelatin derivatives (Ed, Sd, and Sm) were mixed with negatively charged DNA, a self assembly of DNA nanoparticle (complex) was formed within few minutes through electrostatic interaction. Irrespective of the type of gelatin derivatives, the apparent molecular size of DNA was reduced by increasing the gelatin/DNA mixing ratio to attain a saturated value of about 150 nm. The condensed gelatin/DNA complexes showed the zeta potential of 10-15 mV. The amount of DNA internalized into the cells was significantly increased by the complexation with every gelatin derivative. The cells incubated with the gelatin/DNA complexes exhibited significantly stronger luciferase activities than naked plasmid DNA. This study clearly demonstrates and self-assembled DNA complexes has potential as a gene delivery vechile and are stable to transfer genetic materials to cells.

Animals↗