• Maxillofacial Plastic and Reconstructive Surgery
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• v.32 no.4
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• pp.299-305
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• 2010

Purpose: Adipose tissue is located beneath the skin, around internal organs, and in the bone marrow in humans. Its main role is to store energy in the form of fat, although it also cushions and insulates the body. Adipose tissue also has the ability to dynamically expand and shrink throughout the life of an adult. Recently, it has been shown that adipose tissue contains a population of adult multipotent mesenchymal stem cells and endothelial progenitor cells that, in cell culture conditions, have extensive proliferative capacity and are able to differentiate into several lineages, including, osteogenic, chondrogenic, endothelial cells, and myogenic lineages. Materials and Methods: This study focused on endothelial cell culture from the adipose tissue. Adipose tissues were harvested from buccal fat pad during bilateral sagittal split ramus osteotomy for surgical correction of mandibular prognathism. The tissues were treated with 0.075% type I collagenase. The samples were neutralized with DMEM/and centrifuged for 10 min at 2,400 rpm. The pellet was treated with 3 volume of RBC lysis buffer and filtered through a 100 ${\mu}m$ nylon cell strainer. The filtered cells were centrifuged for 10 min at 2,400 rpm. The cells were further cultured in the endothelial cell culture medium (EGM-2, Cambrex, Walkersville, Md., USA) supplemented with 10% fetal bovine serum, human EGF, human VEGF, human insulin-like growth factor-1, human FGF-$\beta$, heparin, ascorbic acid and hydrocortisone at a density of $1{\times}10^5$ cells/well in a 24-well plate. Low positivity of endothelial cell markers, such as CD31 and CD146, was observed during early passage of cells. Results: Increase of CD146 positivity was observed in passage 5 to 7 adipose tissue-derived cells. However, CD44, representative mesenchymal stem cell marker, was also strongly expressed. CD146 sorted adipose tissue-derived cells was cultured using immuno-magnetic beads. Magnetic labeling with 100 ${\mu}l$ microbeads per 108 cells was performed for 30 minutes at $4^{\circ}C$ a using CD146 direct cell isolation kit. Magnetic separation was carried out and a separator under a biological hood. Aliquous of CD146+ sorted cells were evaluated for purity by flow cytometry. Sorted cells were 96.04% positivity for CD146. And then tube formation was examined. These CD146 sorted adipose tissue-derived cells formed tube-like structures on Matrigel. Conclusion: These results suggest that adipose tissue-derived cells are endothelial cells. With the fabrication of the vascularized scaffold construct, novel approaches could be developed to enhance the engineered scaffold by the addition of adipose tissue-derived endothelial cells and periosteal-derived osteoblastic cells to promote bone growth.

• Journal of Chest Surgery
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• v.36 no.10
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• pp.711-720
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• 2003

Bakground : Complete resection by the surgery has been selected as the treatment of choice in lung cancer patients, but in cases of recurrence after excision or inoperable cases, the importance of anticancer chemotherapy has been emphasized. If one can select a set of the sensitive chemotherapeutic agents before anticancer chemotherapy, it will give more favourable results. Subrenal capsular assay has been recognized as a useful in-vivo chemosensitivity test of thoracic and abdominal tumors and it can be done in a short time for a rapid interpretation of tumor responsiveness to anticancer chemotherapeutic drugs. It has been reported that various kinds of cancer cells can be implantable to the kidney, but so far there is no comparative study of xenogeneic cell implantation on liver, spleen and kidney. The author implanted the human lung cancer cells under the capsule of S.D rat's liver, spleen and kidney respectively and compared the pattern of growth and histology. Material and Method: After incubation of human lung cancer cell line (SW-900 G IV) in RPMI 1640 (Leibovitz L-15 medium) culture media, 3${\times}$3${\times}$3 mm size fibrin clots which contain 108 cancer cells were made. Thereafter the fibrin clots were implanted at subcapsule area of liver, spleen and kidney of S.D. female rat. For immune suppression, cyclosporin-A (80 mg/Kg) was injected subcutaneously daily from post-implantation first day to sixth day. The body weight was measured at pre and post implantation periods. The growth pattern and the size of tumor mass were observed and the pathologic examination and serum tumor marker tests were performed. Result: Body weight increased in both of control and experimental groups. Serum Cyfra 21-1 was not detected. Serum levels of CEA and NSE revealed no significant change. The SCC-Ag increased significantly in implanted group. The growth rate of human lung cancer cells which was implanted on spleen was higher than on liver or kidney. The surface area, thickness, and volume of tumor mass were predominant at spleen. The success rates of implantation were 80% on kidney, 76.7% on spleen and 43.3% on liver. Pathologic examination of implanted tumors showed characteristic findings according to different organs. Tumors that were implanted on kidney grew in a round shape, small and regular pattern. In the spleen, tumors grew well and microscopic neovascularization and tumor thrombi were also found, but the growth pattern was irregular representing frequent daughter mass. Human lung cancer cells that were implanted in the liver, invaded to the liver parenchyme, and had low success rate of implantation. Microscopically, coagulation necrosis and myxoid fibrous lesion were observed. Conclusion: The success rate of implantation was highest in the kidney. And the mass revealed regular growth that could be measured easily. The SCC-Ag was presented earlier than CEA or Cyfra21-1. The Cyfra21-1 was not detected at early time after implantation. The best model for tumor implantation experiment for chemosensitivity test was subrenal capsular analysis than liver and spleen and the useful serum tumor marker in early period of implantation was the SCC-Ag.

• Journal of Korean Neurosurgical Society
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• v.30 no.5
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• pp.553-560
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• 2001

Objective : The objective of this study was to determine the photodynamic therapeutic response of U-87 human glioma cell in vitro as well as in the nude rat xenograft model using photofrin as photosensitizer. Material and Method : U-87 cells were cultured on 96-well culture plates, photofrin(Quadralogic Technologies Inc., Vancouver, Canada) was added into the cell culture medium at concentration of $1{\mu}g/ml$, $2.5{\mu}g/ml$, $5{\mu}g/ml$, $10{\mu}g/ml$ and $20{\mu}g/ml$. 24 hour after drug treatment, cells were treated with optical(632nm) irradiation of $100mJ/cm^2$, $200mJ/cm^2$ and $400mJ/cm^2$. Photofrin(12.5mg/kg, i.p.) was administered to 28 nude rats containing intracerebral U-87 human glioma as well as 26 normal nude rats. 48 hours after administration, animals were treated with optical irradiation(632nm) of $35J/cm^2$, $140J/cm^2$ and $280J/cm^2$ to exposed tumor and normal brain. The photofrin concentration was measured in tumor and normal brain in a separate population of animals. Results : By MTT assay, there was 100% cytotoxicity at any dose of photofrin with optical irradiation of $200mJ/cm^2$ and $400mJ/cm^2$. But at the optical irradiation of $100mJ/cm^2$ cells were killed in dose dependent manner 28.5%, 49.1%, 54.4%, 78.2%, and 84.6% at concentration of $1{\mu}g/ml$, $2.5{\mu}g/ml$, $5{\mu}g/ml$, $10{\mu}g/ml$ and $20{\mu}g/ml$, respectively. Dose dependent PDT lesions in both tumor and normal brain were observed. In the tumor lesion, only superficial tissue damage was found with optical irradiation of $35J/cm^2$. However, in the optical irradiation group of $140J/cm^2$ and $280J/cm^2$ the volume of lesions was measured of $7.2mm^3$ and $14.0mm^3$ for treatment at $140J/cm^2$ and $280J/cm^2$, respectively. The U-87 bearing rats showed a photofrin concentration in tumor tissue of $6.53{\pm}2.16{\mu}g/g$, 23 times higher than that found in the contralateral hemisphere of $0.28{\pm}0.15{\mu}g/g$. Conclusion : Our data indicate that the U-87 human glioma in vitro and in the xenografted rats is responsive to PDT. At these doses, a reproducible injury can be delivered to human glioma in this model. Strategies to spare the normal brain collateral damage are being studied.

• Tuberculosis and Respiratory Diseases
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• v.44 no.5
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• pp.1114-1124
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• 1997

Background : Endothelin(ET) is a very potent vasoconstrictive peptide produced by endothelial cells of pulmonary artery. The endothelin level was increased in plasma of primary pulmonary hypertension and acute pulmonary thromboembolism and it was suggested that the endothelin might do a critical role in the cardiopulmonary dysfunction in these two conditions. But the exact mechanism of increase of ET has not been known. In these two conditions, platelet activation and thrombosis are the main pathophysiologic findings. So there is a possibility that the platelet might stimulate endothelin secretion from endothelial cells. Therefore, we performed this study to evaluate the role of platelet and its mediators on endothelin production in bovine pulmonary artery endothelial(BPAE) cells. Method : Bovine pulmonary artery endothelial cells, ATCC certified cell line 209, were cultured and treated with human platelets($10^6{\sim}10^8/ml$), thrombin (0.1~10u/ml), TGF-${\beta}1$(1~100uM), serotonin(1~100uM), and endotoxin(1ug/ml) in a final volume of 500ul for 18 hours. Levels of ir(immunoreactive)-ET in each conditioned medium were measured by a radioimmunoassay specific for ET. Result : The increase of ir-ET levels was platelet number and time dependent over 18 hours. When washed human platelets were added($10^8/ml$), the ir-ET levels were significantly higher than that of control(p<0.05) at 8 and 18 hours after culture. Subthreshold concentration of platelets($10^7/ml$) coincubated with endotoxin(1ug/ml) or subthreshold dose of thrombin(0.1u/ml) stimulated ir-ET secretion from BPAE cells significantly(p<0.05) compared with control. Thrombin(1ug/ml, 10ug/ml) and TGF-${\beta}1$(100pM, 1000pM) significantly increased ir-ET secretion from BP AE cells(p<0.05) compared with control, but serotoin(1~100uM) and endotoxin(1ug/ml) did not stimulate the ir-ET secretion. Conclusions : Platelets stimulate endothelin secretion from bovine pulmonary artery endothelial cells. The mechanism of increase of endothelin secretion seems to be a stimulation by platelet itself or by mediators, such as TGF-${\beta}1$, secreted from activated platelets. And, in this study, the priming effect of platelets on endothelin secretion from BPAE cells could be another possibility.