ABSTRACT
<p><b>OBJECTIVE</b>To evaluate the method of adenovirus vector-mediated herpes simplex virus-thymidine kinase gene (ADV-TK)/ganciclovir(GCV) system and photodynamic therapy(PDT) for treating the oral malignant tumor.</p><p><b>METHODS</b>Ten patients who were suffering from oral malignant tumor of the different positions were selected, and injected with the hematoporphyrin derivative (HPD), irradiated by picking the corresponding laser frequency, and injected the ADV-TK gene into the tumor body and periphery. By the imaging and the hemodynamics analysis, the clinical efficacy after infusing vein with GCV was assessed.</p><p><b>RESULTS</b>After the combination method treatments, patients' tumors appeared clear shrinkage or complete extinction. There was obvious fall of the blood flow volume in the tumor body. Imaging results showed significantly differences. So it was a perfect and effective treatment.</p><p><b>CONCLUSION</b>There are many advantages to apply the ADV-TK/GCV system and PDT treatment on the oral malignant tumor, minimal side effects and greater clinical security. It is a safe and credible therapy which can be offered for curing the oral malignant tumor systematically.</p>
Subject(s)
Humans , Adenoviridae , Ganciclovir , Genetic Vectors , Neoplasms , Photochemotherapy , Simplexvirus , Thymidine KinaseABSTRACT
BACKGROUND:Because human cells for culturing alveolar bone cell line are from alveolar bone, which is in oral cavity,and easily polluted, so laboratory study is often unsuccessful. Because the samples are from adults, so cell division index and the successful rate of culture are low.OBJECTIVE: To compare the biological characteristics of survived cell line established through passage,cryopreservation and revitalization following in vitro culturing the alveolar bone tissue obtained from normal persons and patients with chronic periodontitis accompanied with osteoporosis in aseptic operation; To compare the biological characteristics of two kinds of cells so as to provide theoretical and related experimental evidence for defect, repair and treatment of alveolar bone.DESIGN: Controlled observation.SETTING: Department of Stomatology, Beijing Chaoyang Hospital, Capital Medical University; Institute of Orthopaedics,General Hospital of Chinese PLA.MATERIALS: Alveolar bone tissue obtained from normal persons and patients with chronic periodontitis confirmed in clinic was used in aseptic operation.METHODS: Alveolar bone tissue from normal persons and chronic periodontitis accompanied with osteoporosis were cultured in vitro. In the four cell lines (H-171, H-258, 261, 262) cultured primarily, cell lines H-171 and H-258 were chosen from periodonitis patients group and normal group respectively, and stained with histochemical and immunohistochemical methods. Cell morphology was observed. Doubling time and division index of two kinds of cells were calculated with cytometry. After several circles of passage, cryopreservation and revitalization, growth and aging rule of cells were compared.MAIN OUTCOME MEASURES: Passage and biological characteristics of two groups of cell lines.RESULTS: ①In the abnormal alveolar bone group, there was one successful primary culture and cells presented short-spindle shape. There were 3 times of cryopreservation and 3 times of revitalization. Its doubling time was 53.4 hours. The average division index was about 4‰. Cells well grew after 20 times of passages. ②In the normal alveolar bone group, there were 26 cases of cell lines cultured primarily, but passage was found in only 3 cases of cell lines due to various causes. There were 10 passages and the cells presented long-spindle shape. After two circles of cryopreservation and revitalization, the survival and growth rate of cells were inferior as compared with cell line H-171.Doubling time was 65.9 hours and the average division index was 3.5‰. ③Both two kinds of cells adhered the wall, with the characteristics of osteoblasts: AKP, toluidine blue, PAS, tetracycline-labeled mineralized nodus, type Ⅰ collagen and BMP-2 immunohistochemical staining all presented positive.CONCLUSION: Both two kinds of cultured cells have the characteristics of osteoblasts. The growth speed of cell line H-171 is faster than that of cell line H-258. No obvious mutation is found in 20 passages. In the 8th generation of H-258,aging appears and growth speed becomes slow.
ABSTRACT
BACKGROUND: The transcription factor, hypoxia-inducible factor-1al pha (HIF-1α), is the key regulator that controls the hypoxic response of mammalian cells. However, the role of HIF-1α in the therapeutic efficacy of chemo-radiotherapy in oral squamous cell carcinomas (OSC) is poorly understood. OBJECTIVE: To investigate the effect of HIF-α on the susceptibility of OSC cell lines against chemotherapeutic drugs and radiation. DESIGN: AN observational comparative experiment. SETTING: Beijing Chaoyang Hospital Affiliated to Capital Medical University. MATERIALS: OSC-2, OSC-4, OSC-5, and OSC-6 cell lines were estab lished from oral squamous cell carcinoma (The cell lines were from the De partment of Oral Oncology, Kochi Medical School, Japan). METHODS: The experiments were completed in Beijing Chaoyang Hospi tal affiliated to Capital Medical University from September 2004 to August 2006. The cells were cultured in Dulbecco's modified Eagle's medium (DMEM) containing fetal bovine serum (FBS), L-glutamine, penicillin and streptomycin. ① Total cell lysates were extracted from untreated OSC cells and those treated with 30 Gy γ-rays, 100 μmol/L cis-dichlorodiamine plat inum (CDDP), or 100 μmol/L 5-fluorouracil (5-FU). The expressions of HIF-1α protein and mRNA were determined with western blotting analysis and real-time polymerase chain reaction (PCR). ② Inhibition of cell prolif eration after different interventions: OSC cells were seeded in 96-well mi croplates (1.0×104) and cultured for 48 hours after the irradiation with 30 Gy γ-rays or in the presence of 100 μmol/L CDDP or 100 mmol/L 5-FU, and then the inhibition of cell proliferation was detected with 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. ③ Apoptosis after different interventions: OSC cell lines were cultured for 24 hours after the irradiation with 30 Gy γ-rays or in the presence of 100 μmol/L CDDP or 100 μmol/L 5-FU. The cells were then stained with annexin V-FITC and propidium iodide, and the numbers of apoptotic cells were analyzed by a FACScan cytometer (only for the OSC cells treated with γ-rays and CDDP). ④ Plasmid construction and transfection: Human HIF 1α cDNA was cloned into the pcDNA3.1/V5-His TOPO expression vector. OSC-2 cells were temporarily transfected with HIF-1α cDNA. OSC-5 cells were temporarily transfected with HIF-1α siRNA (Sense sequence is 5' CUGAUGACCAGCAACUUGAtt 3'). The blank control group was also set, the inhibition of cell proliferation was observed, and the apoptosis was ana lyzed (only in the CDDP-treated group) by using the methods mentioned above. MAIN OUTCOME MEASURES: ① Expressions of HIF-1α protein and mRNA in different OSC cell lines; ② Inhibition of proliferation and the apoptosis of OSC cell lines after different interventions; ③ Overexpression and knockdown of HIF-1α by expression vector and siRNA; ④ Influence of overexpression and knockdown of HIF-1α on the susceptibility of OSC cell lines against γ-rays and chemotherapeutic drugs. RESULTS: ① Expressions of HIF-1α protein and mRNA in different OSC cell lines: The relative expression levels of HIF-1α mRNA were 1.0, 2.2, 4.3 and 4.0 in OSC-2, OSC-4, OSC-5 and OSC-6 cells, respectively. Simi larly, the expression of HIF-1α total cell lysates proteins and nuclear proteins were much lower in OSC-2 and OSC-4 cells than in OSC-5 and OSC6 cells. ② Inhibition of proliferation and the apoptosis in OSC cell lines after different interventions: After the treatment with g-rays, CDDP, and 5-FU, the cell numbers decreased obviously to (39.5±3.2)%, (39.2±1.2)%,and(47.9±3.6)% in OSC-2 cells, (53.9±6.6)%, (54.3±1.4)%, (54.8±3.8)%in OSC-4 cells. Those decreased to (74.1±3.8)%, (76.5±9.1)%, (69.6±7.7)%in OSC-5 cells and (71.4±7.4)%, (84.4±8.8)%, (82.0±4.5)% in OSC-6 cells.After the OSC cells were treated of with CDDP, the numbers of apoptotic cells were (50.9±1.3)%, (67.3±2.2)%, (12.2±0.8)% and (38.6±0.9)% in OSC-2, OSC-4, OSC-5 and OSC-6 cells. G-rays induced the apoptosis were (21.2±1.1)%, (14.6±0.9)%, (9.7±1.0)% and (10.4±0.8)% in OSC-2, OSC-4,OSC-5 and OSC-6 cells. ③ Inhibition of proliferation and the apoptosis in the transfected cell after different interventions: The cell numbers of HIF-1 α-transfected OSC-2 cells treated with γ-rays, CDDP and 5-FU were much more than those in the control group (t=-4.693 ,-8.617,-6.721, P ＜ 0.01),whereas the cell numbers of OSC-5 cells transfected with siRNA were obviously fewer than those in the control group (t=5.800, 5.595, 4.253, P ＜ 0.05-0.01). After the incubation of HIF-1α-transfected OSC-2 cells with CDDP for 24 hours, apoptosis was detected in (34.0±1.9)% of the cells, which was lower than that in the control group [(49.6±3.4)%, t=6.937, P ＜ 0.01]. After the incubation of HIF-1α siRNA transfected OSC-5 cells with CDDP for 24hours, apoptosis was detected in (27.7±2.3)% of the HIF-1α siRNA transfected OSC-5 cells, which was obviously higher than that in the blank control group [(11.4±2.1)%, t=-8.941, P ＜ 0.01].CONCLUSION: The expression level of HIF-1α correlates negatively with the susceptibility of OSC cells against chemotherapeutic drugs and radiation. The down-regulation of HIF-1α expression enhances the susceptibility of OSC cells against chemotherapeutic drugs and radiation. Therefore, the down-regulation of HIF-1α may be a potentially effective strategy for cancer treatment.
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the biological characteristics of cell lines of healthy and diseased human dental alveoli.</p><p><b>METHODS</b>Primary cell lines from either healthy or diseased human dental alveoli were obtained. Two cell lines, H-258 and H-171 derived from healthy and diseased human tissues respectively, were selected for morphological study and research on their growth and aging, using cell counting, and histochemical and immunohistochemical staining.</p><p><b>RESULTS</b>Primary cell lines were successfully established from innormal dental alveoli. After freezing and thawing for three times, cell growth was continued and no morphological alterations were observed. The doubling time was 53.4 hours and mean division index (MDI) was 4 per thousand. Cells were kept normal after twenty generations with no obvious reduction of doubling time and MDI. Of twenty-six primary cell lines derived from healthy human dental alveoli, only three cell lines achieved generation. After freezing and thawing for twice, cultured cells were still alive at a decreased growth speed, with doubling time of 85.9 hours and MDI of 3 per thousand. Both cell lines, H-171 and H-258, shared the characteristics of osteoblast.</p><p><b>CONCLUSIONS</b>Primary cell lines of diseased human dental alveoli show greater growth potential. All cell lines of dental alveoli share characteristics of osteoblast. The technique we developed may be put into practice for the treatment of abnormal dental alveoli.</p>