• Polymer(Korea)
• /
• v.36 no.6
• /
• pp.768-775
• /
• 2012

Alginate, obtained from the seaweeds, is a widely used biomaterial for cell transplantation, since its positive effect on viability of capsulized cells and its easier encapsulation capability of living cells. Demineralized bone powder (DBP), derived from the natural bone tissue, is widely applied for clinical trials for its low rate of reaction and antigenicity. A chondrocyte was seeded into an alginate with DBP of different contents, and a microcapsule was produced. The adhesion and proliferation of cells was observed through the MTT analysis, and the PCR was applied to estimate the content of the glycosaminoglycan (sGAG) and collagen, and confirm the specific genetic pattern of the chondrocytes. Also, the alginate microcapsule where the chondrocyte is seeded was extracted after transplantation under the skin of a nude mouse, and was immunochemically stained. The experimental result confirmed that the alginate microcapsule containing 1% of DBP not only showed the highest proliferation of cell but had a positive effect of chondrocytes by the interaction between the alginates and the growth factor in DBP. It can be expected that the microcapsule with application of the alginates and DBP might be an appropriate scaffold for tissue engineering.

• Journal of Chest Surgery
• /
• v.37 no.6
• /
• pp.482-491
• /
• 2004

Background: The xenogenic or allogenic valves after in Vitro repopulation with autologous cells or in vivo repo-pulation after acellularization treatment to remove the antigenicity could used as an alternative to synthetic polymer scaffold. In the present study, we evaluated the process of repopulation by recipient cell to the acellu-larized xenograft treated with NaCl-SDS solution and grafted in the right ventricular outflow tract. Material and Method: Porcine pulmonary valved conduit were treated with. NaCl-SDS solution to make the grafts acellularized and implanted in the right ventricular outflow tract of the goats under cardiopulmonary bypass. After evaluating the functions of pulmonary valves by echocardiography, goats were sacrificed at 1 week, 1 month, 3 months, 6 months, and 12 months after implantation, respectively. After retrieving the implanted valved conduits, histopathologic examination with Hematoxylin-Eosin, Masson' trichrome staining and immunohistochemical staining was performed. Result: Among the six goats, which had been implanted with acellularized pulmonary valved conduits, five survived the expected time period. Echocardiographic examinations for pulmonary valves revealed good function except mild regurgitation and stenosis. Microscopic analysis of the leaflets showed progressive cellular in-growth, composed of fibroblasts, myofibroblasts, and endothelial cells, into the acellularized leaflets over time. Severe inflammatory respon-se was detected in early phase, though it gradually decreased afterwards. The extracellular matrices were regenerated by repopulated cells on the recellularized portion of the acellularized leaflet. Conclusion: The acellularized xenogenic pulmonary valved conuits were repopulated with fibroblasts, myofibroblasts, and endothelial cells of the recipient and extracellullar matrices were regenerated by repopulted cells 12 months after the implantation. The functional integrity of pulmonary valves was well preserved. This study showed that the acellularized porcine xenogenic valved conduits could be used as an ideal valve prosthesis with long term durability.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.35 no.1
• /
• pp.1-7
• /
• 2002

To study the increased resistance in fish against Vibrio vulnificus known as an important agent of vibrio septicemia in human, we analyzed specific and nonspecific immune response in flounder after administration of V. vulnificus bacterins by oral route. It contained the comparison of antibody concentrations in the sera of flounder after oral administration by two different protocols with uncoated heat killed bacterin of V vulnificus (UHKB, 20 mg/kg body weight), i.e., 4 weeks continuously (group 4W) and taking 2 weeks resting period between the 1st and last week of administration (group 1-2-1W). Even though, 1-2-1W group showed significantly increased level of specific antibody in serum, it did not reach to that of 4W group. Certainly, flounder vaccinated twice a week for four weeks (20 mg/kg b.w.) showed increased concentration of specific antibody against V. vulnificus at week 2 after last administration by oral route and maintained throughout the experimental period. It also was confirmed by the increased numbers of specific antibody secreting cells (SASC) in the leukocytes isolated from the splenocytes of the flounder of 4W group at week 1 after last administration until the end of experimental period. However, enteric, acid-resistant film coated heat killed bacterin (ECHB) did not show both greater immune reaction for antibody production and faster elimination of a challenge dose of V. vulnificus compared with those of the UHKB. These results suggested that UHKB administered by oral route was very effective method to prevent the contamination of V vulnificus in flounder, and did not show the increased antigenicity by coating the surface with acid-resistant film.

• Journal of Life Science
• /
• v.30 no.10
• /
• pp.905-911
• /
• 2020

The adjuvant effect of PAMAM dendrimer G4 (PAMAM) on the induction of humoral and cellular immune responses against keyhole limpet hemocyanin (KLH) was examined. Mice were immunized subcutaneously twice at two-week intervals with KLH, with or without PAMAM dendrimer (100 ㎍/mouse), and the mice immunized with KLH+PAMAM showed significantly higher antibody titers against KLH than those immunized with KLH alone. The assay for determining the isotypes of the antibodies showed that PAMAM augmented the KLH-specific antibody titers of IgG1, IgG2a, IgG2b, IgG3, and IgM. In addition, mice immunized twice with KLH+PAMAM followed by a subcutaneous injection of KLH (20 ㎍/site) 7 weeks after the primary immunization exhibited a higher delayed-type hypersensitivity (DTH) reaction than those treated with KLH alone. In an in vitro analysis of T lymphocyte proliferation in response to KLH in week 8, the splenocytes of mice treated with KLH+PAMAM showed significantly higher proliferating activity than those treated with KLH alone, and the culture supernatants of cell cultures from mice immunized with added PAMAM dendrimer showed higher levels of KLH-specific cytokine (IL-4 and IFN-r) production. These results suggest that PAMAM dendrimer G4 possesses a potent immune-adjuvant activity for enhancing both humoral and cell-mediated immunity specific to foreign antigens.

• Parasites, Hosts and Diseases
• /
• v.24 no.2
• /
• pp.145-158
• /
• 1986

This study was undertaken to purify cystic fluid (CF) antigen of Taenia solium metacestodes by affinity chromatogaphy using specific monoclonal antibody(McAb) and to characterize the antigenicity of the purified antigen. The hybridoma cell lines, prepared by fusion between mouse plasmacytoma and spleen cells from BALB/c mice immunized with CF, secreted antibodies reacting to various helminthic antigens. Majority of cell lines reacted to CF only but some also reacted to parenchymal antigen of T. solium metacestodes, adult T. saginata, sparganum, hydatid cystic fluid, Paragonimus westermani and Clonorchis sinensis, either in combination with CF, other antigens or independently. Cloned cells derived from monoclonal lines also produced antibodies reacting either to CF only or to other helminthes in combination or independently. These results indicated that CF of T. solium metacestodes contained proteins which possessed antigenic determinants not only specific to CF but also cross reactive with the afore-mentioned helminthes. CF of T. solium metacestodes was purified by affinity chromatography using the McAb which reacted to CF and parenchymal antigens. The affinity-purified antigen (A-Ag) and unbound pool CF (U-Ag) were separated. A-Ag showed 2 protein bands by disc-PAGE whereas CF exhibited 6 bands and U-Ag consisted of all bands CF had. The diagnostic significance of A-Ag was evaluated by ELISA in human neurocysticercosis and other helminthic and neurologic diseases. By A-Ag, the levels of the specific IgG antibody, as shown by absorbance in sera and CSF, were lower than those of CF and U-Ag. Accordingly, the sensitivity was about 70% of CF and U-Ag. However, the nonspecific positive reactions to CF and U-Ag, observed in sparganosis, T. saginata infection and paragonimiasis did not occur when A-Ag was used. These results indicated that the affinity-purified A-Ag had the higher specificity but the lower sensitivity as a diagnostic antigen in cysticercosis, probably because it only detected a single or limited numbers of monospecific antibodies among the diverse polyclonal antibodies produced in the patients with neurocysticercosis.

• Journal of Chest Surgery
• /
• v.36 no.4
• /
• pp.229-241
• /
• 2003

Background:Autogenous vein is the preferred vascular graft for patients who require coronary artery bypass surgery or peripheral arterial bypass surgery. When an autogenous vein is not available, an allograft saphenous vein can be used as an alternative conduit. Although arterial homograft has been under investigation since the beginning of this century, the viability of endothelial cells and the optimum mode of storage for the venous and arterial allografts is controversial. In addition, with the recently gained knowledge of vascular endothelial functions, such as the production of nitric oxide or thrombomodulin, the viability and antigenicity of endothelial cells are being studied again. The purpose of this study was to evaluate the viability of endothelial cells in the preserved human saphenous veins. Material and Method: The veins were stored in a $4^{\circ}C$ RPMI (Roswell Park Memorial Institute) 1640 solution including 10% fetal calf serum, for one, three, five, seven or fourteen days. After the completion of the storage period, the veins were divided into two groups: Group I: studied immediately at $4^{\circ}C$ (cold) storage (I-1, I-3, I-5, I-7, I-14), and Group II: studied after storage at $-196^{\circ}C$ liquid nitrogen tank (cryopreservation) in an RPMI 1640 solution containing 10% DMSO for two weeks (II-1, II-3, II-5, II-7, II-14). Light microscopy and scanning electron microscopy (SEM), frypan blue exclusion testing, and thrombomodulin immunohistochemistry were performed. Result: In a morphometric study using SEM, there was statistically significant increase in Gundry Score in Groups I-7, I-14, II-5, II-7, and II-14 and showed cellular destruction (p<0.05). In the thrombomodulin immunohistochemistry study, there was reactivity in Groups I-1, I-3, and I-5, but the cryopreserved group revealed decreased reactivity (p<0.05). The trypan blue exclusion testing also showed superior viability in cold storage Group I. Conclusion: Venous allografts preserved in a $4^{\circ}C$ RPMI 1640 solution showed well preserved endothelial cellular integrity and thrombomodulin expression at up to seven days of preservation. Although cryopreservation of venous allografts stored in 10% DMSO -RPMI 1640 solution maintained the endothelial cellular structure on SEM, immunohistochemistry from the thrombomodulin and trypan blue exclusion testing showed decreased viability, It remains to be seen whether the decreased thrombomodulin reactivity could be restored, and what the nature to the relationship is between thrombomodulin and long-term patency of allografts.

• Korean Journal of Microbiology
• /
• v.44 no.3
• /
• pp.178-185
• /
• 2008

Avian influenza virus (AIV) is recognized as key to the emergence of pandemic influenza for humans; there are growing concerns that AIV H9N2 may become more efficient to transmit to humans in the near future, since the infection of poultry with AIV H9N2 has been common in recent years. In this study, we aimed to produce antisera recognizing the HA and NA proteins of AIV H9N2. Initially, coding sequences corresponding to the N-terminal regions of the HA and NA proteins of the Korean AIV H9N2 (A/Ck/Kr/MS96/96) isolated from a domestic chicken were amplified from the genomic RNA. Following cloning of the amplified cDNA fragments into pGEX4T-1 vector, two GST-fusion proteins (GST-HAln and GST-NAn) were expressed in E. coli BL21 and purified with glutathione sepharose columns; the recombinant GST-HAln and GST-NAn proteins were both used as immunogens in rabbits. The antigenicity of the rabbit antisera was analyzed by immunoblotting of the cell lysates prepared from AIV H9N2-infected MDCK cells. Overall, the recombinant HAln and NAn proteins fused to the C-terminus of GST and the rabbit antisera raised against the corresponding recombinant proteins would provide a valuable reagent for AIV diagnosis and basic research.

• Journal of Veterinary Clinics
• /
• v.27 no.5
• /
• pp.501-507
• /
• 2010

The aim of the current study was to analyze the diversity of the major surface protein (Msp) gene in Theileria buffeli, which is known as the major antigenic protein recognized by the immune system of the host. In addition, we characterized the diversification of the Msp gene and its relationship to with the pathogenicity of Theileria. Complete blood counts (CBC) and Theileria 18S rRNA PCR sequence analysis were performed for 177 Korean indigenous cattle (KIC) in Jeju Island. A total of 28 KIC (16 anemic and 12 non-anemic KIC) were then randomly selected based on 18s rRNA PCR positive samples for sequence analysis of the Theileria Msp gene, which was performed twice for each specimen. The resulting 56 Msp gene sequences were classified into five antigenicity types (type I to V), according to the variable region (517-571 bp), which exhibited high similarity (${\geq}$ 98.9%) to several available GenBank sequences (Theileria spp. from China-EU584237; T. sergenti from China-DQ078264; Theileria spp. from Thailand-AB081329; Theileria spp. from Japan-AB218442; T. sergenti from Japan-AB016280). The 56 Msp sequences consisted of 22, 15, 9, 8, and 2 cases of type I to type V Msp genes, respectively. The most prevalent type in both anemic and non-anemic KIC was type I (37.5% in anemic and 41.7% in non-anemic). Among the remaining types, type II was the most prevalent (37.5%) in anemic KIC, while type IV was the most prevalent (25%) in non-anemic KIC. The results of our study help confirm the diversity of Msp gene types and demonstrate that the gene type distribution of Msp genes varies among Theileria-infected KIC in Jeju Island.

• Journal of Chest Surgery
• /
• v.29 no.11
• /
• pp.1173-1181
• /
• 1996

The causes of degenerative changes in allograft cardiac valves are not well known to this day. Today's preserved allografts possess highly viable endothelial cells and degeneration of allografts can be facilitated by immune reaction which may be mediated by these viable cells. To test the antigenicity of endothelial cells, pieces from aortic wall were obtained from fresh and cryo-preserved rat allograft. Timings of sampling were prior to sterilization, after sterilization, after 1, 2, 7, 14 days of fresh preservation and cryopreservation. Endothelial cells were tested by immunohistochemical methods using monoclonal antibodies to MHC class I(MRC OX-18), class II(MRC OX-6) and ICAM-1 antigens. After transplantation of each group of aortic allograft at the subcutaneous layers of rats, population of CD4$^{+}$ T cell and CD8$^{+}$ T cell were analyzed with monoclonal antibodies after 1, 2, 3, 4, 6 and 8 weeks. MHC class I expression was 23.95% before preservation and increased to 35.53~48.08% after preservation(p=0.0183). MHC Class II expression was 9.72% before preservation and 10.13~13.39% after preservation(P=0.1599). ICAM-1 expression was 15.02% before preservation and increased to 19.85~35.33% after preservation(P=0.001). The proportion of CD4$^{+}$ T-cell was 42.13% before transplantation. And this was 49.23~36.8% after transplantation in No treat group (p=0.955), decreased to 29.56~32.80% in other group(p=0.0001~0.008). In all the groups, the proportion of CD8$^{+}$ T-cell increased from 25.57% before transplantation to 42.32~58.92% after transplantation(p=0.000l~0.0002). The CD4$^{+}$/CD8$^{+}$ ratio decreased from 1.22~2.28 at first week to 0.47~0.95 at eighth week(p=0.0001). The results revealed that the expression of MHC class I and ICAM-1 in aortic allograft endothelium were increased but that of MHC class II were not changed, despite the different method of preservation. During 8 weeks after transplantation of aortic allograft, the subpopulations of CD4$^{+}$ T cell were not changed or only slightly decreased but those of CD8$^{+}$ T cell were progressively increased.ely increased.