血卟啉衍生物（HPD）加光对细胞损伤作用机理的研究：III.HPD加光...

The effects of HPD plus light on lysosome membrane and release of hydrolysis enzymes from lysosomes of normal rat liver and hepatoma cells were investigated. HPD was bound to lysosome either in vitro or in vivo. Lysosomes bound HPD plus light increased lipid peroxidation of unsaturated fatty acid of lysosome membrane and enhanced the activity of DNase, RNase, cathepsin and acid phosphatase. These effects were related to HPD concentration and exposure time but it was unchanged in the control. It was concluded that the enhancement of hydrolysis enzymes from lysosomes was due to the lysosome membrane damage under the action of siglet oxygen produced by HPD bound lysosomes following light activation.     

Mechanism of cell damage by hematoporphyrin derivative (HPD) plus light. II. Effect of HPD plus light on respiration and oxidative phosphorylation in hepatoma cells and normal liver mitochondria

The effects of HPD plus light on energy metabolism and membrane structure of hepatoma cell and normal liver mitochondria were investigated. HPD was bound to mitochondria either in vitro or in vivo. Mitochondria bound HPD plus light increased MDA level, decreased respiratory intensity and respiratory control, dissociated phosphorylation from oxidation and enhanced mitochondria swelling on age. These effects were related to HPD concentration and exposure time but it was unchanged in the control. These results showed that the loss of function of oxidation phosphorylation was due to the mitochondrial membrane system damage under the action of singlet oxygen produced by HPD bound mitochondria following light activation.     

Cytotoxic effect of hematoporphyrin derivative (HPD) plus light irradiation in vitro

Cytotoxic effect of HPD plus light irradiation on PTK2 and HeLa cells in vitro were studied by laser microbeam irradiation. The dynamic changes, in three different steps as observed in the cells ranged from visible damage to cell lysis, were recorded at the monocellular level. It was found that the speed of the cytotoxic process varied chiefly with the dose of light used. The cytotoxic progress of PTK2 and HeLa cells was compared and analyzed. In addition, the cytotoxic effects of three different laser wavelengths (6328A, 5145A and 4880A) plus HPD were compared. The most significant killing effect occurred at 6328A wavelength of red light.     

Mechanism of cell damage by ultrasound in combination with hematoporphyrin

The mechanism of cell damage by ultrasound in combination with hematoporphyrin was studied. Mouse sarcoma 180 cell suspensions were exposed to ultrasound for up to 60 s in the presence and absence of hematoporphyrin, with and without active oxygen scavengers. The cell damage enhancement by hematoporphyrin was suppressed by adding histidine but not by mannitol. The enhancement was doubled in rate by substitution of deuterium oxide medium for normal water. Sonoluminescence was produced in a saline solution under similar acoustic conditions and observed to have spectral components that can excite hematoporphyrin molecules. These results suggest that cell damage enhancement is probably mediated via singlet oxygen generated by ultrasonically activated hematoporphyrin.     

Gastric cancer treated by hematoporphyrin derivative(HPD)-laser--histopathological study of 10 gastrectomy specimens

10 patients with gastric carcinoma treated by HPD-laser preoperatively are reported. The gastric lesions were irradiated with laser beam delivered by a quartz fibre through the fiberoptic gastroscope 48-72 hours after intravenous injection of HPD (5.0 mg/kg) about 2 weeks before operation. In the resected specimens, the histological changes following HPD-laser therapy were studied. The cancer cells in the irradiated areas showed degeneration and necrosis in varying degrees. Because of the fact that the light spots were small and the penetration not deep enough, the cancer cells beyond the irradiated area and those infiltrating more deeply or beyond the gastric wall did not show any evident changes while metastatic cancers in the lymph nodes showed no changes at all. These facts may suggest that the HPD-laser therapy should not be used as the main therapeutic method to replace operation, radiotherapy or chemotherapy for gastric cancers in either advanced or early stage.     

抗坏血酸对血卟啉衍生物渗入肿瘤组织及其光敏作用的影响

目的了解肿瘤组织在血卟啉衍生物及抗坏血酸溶液中浸泡一定时间后,血卟啉衍生物渗入瘤组织的深度、在瘤组织中的含量及抗坏血酸对其渗入和光敏作用的影响。方法以小鼠Ｓ１８０实体瘤和光敏剂癌光啉（ＰｓＤ－００７）为材料,将肿瘤组织分别浸泡在ＰｓＤ－００７（浸Ⅰ组）及ＰｓＤ－００７－抗坏血酸（浸Ⅱ组）溶液中１小时,并以静脉注入ＰｓＤ－００７２４小时的肿瘤为对照（静注组）。通过紫外光照射,观察肿瘤组织中ＰｓＤ－００７红色荧光,测定肿瘤组织中ＰｓＤ－００７和丙二醛含量。结果（１）肿瘤横剖面及肿瘤组织冰冻切片在紫外光照射下所显示的ＰｓＤ－００７红色荧光,浸Ⅰ组和浸Ⅱ组均强,且外周强于中心部位;而静注组则弱,但分布均匀。（２）肿瘤中心部组织匀浆中,ＰｓＤ－００７和丙二醛含量均为浸Ⅱ组＞浸Ⅰ组＞静注组（Ｐ＜０．０５）。结论ＰｓＤ－００７及抗坏血酸均能通过浸泡渗入肿瘤组织,抗坏血酸有促进ＰｓＤ－００７渗入肿瘤组织和增强ＰｓＤ－００７光敏反应的作用。     

Purge of malignant cells from bone marrow by hematoporphyrin derivatives and light exposurein vitro

During autologous bone marrow graft in treatment of malignant diseases, it is critical to purge malignant cells from the marrow. In the present study, the sensitivity to photodynamic inactivation of 3 leukemic cell lines was compared with their counterpart normal hematopoietic cells. After mouse leukemic L₁₂₁₀ cells were treated with a preparation of hematoporphyrin derivatives, YHpD, 10 μg/ml for 1 hr. and irradiated with blacklight (peak wavelength 395 nm, light intensity 0.6 mW/cm²) for 5 minutes, the survival rate of clonogenic cells decreased to <10%, while that of bone marrow granulocyte-macrophage progenitor cells (CFU-GM) in DBA/2 mice remained at nearly normal level (>80%). Similar results were obtained when human leukemic HL-60 cells were compared with human CFU-GM and mouse leukemic L₆₁₅ cells with CFU-GM in 615 strain mice. It is suggested that hematoporphyrin photoradiation may be useful for selectively killing leukemic cells in bone marrow.     

Photochemotherapy of glioma cells by visible light and hematoporphyrin.

Malignant tumors take up and retain hematoporphyrin to a much greater extent than do normal tissues. Porphyrins are photodynamic agents that sensitize cells so that they are damaged by exposure to light. Treatment with hematoporphyrin followed by irradiation with light can destroy glioma cells in culture in less than 8 min and gliomas growing s.c. in rats in about 40 min. Photochemotherapy may become useful in the management of malignant tumors that are resistant to current methods of treatment.     

光动力学疗法试治鼻咽癌57例分析

Fifty-seven cases of nasopharyngeal carcinoma treated by photodynamic therapy in our hospital from 1984 to 1986 are reported. Forty-eight to 72 hours after 5 mg/kg HPD injection, 630 nm laser with over 300 mW output transmitted through quartz fiber was given. Of 57 cases, 25 (43.9%) achieved complete response, 25 (43.9%) marked response and 7 (12.2%) mild response. The indications of photodynamic therapy for nasopharyngeal carcinoma are discussed.     

Photo-chemistry therapy of gastrointestinal tumor--analysis of hematoporphyrin derivative (HpD) plus laser photodynamic therapy of 50 cases

This paper reports 50 cases of gastrointestinal tumors treated by photodynamic therapy. The dose of HpD was 5 mg/kg. Fourty-eight to 72 hours after HpD injection, laser (wavelength 630 nm, power density 100-250 J/cm2), transmitted through a quartz fiber, was given. The criteria of therapeutic effectiveness were: complete remission (CR), significant remission (SR), minor remission (MR) and no remission (NR). All patients received endoscopic examination 4 weeks after treatment. No severe complication was observed. Thirty-eight of 50 cases gave CR, SR or MR (76%). The factors effecting treatment are discussed.     

Photoradiation therapy. II. Cure of animal tumors with hematoporphyrin and light.

Exposure of mouse and rat tumors of various types to more than 600 nm light 24 or 48 hours after an injection of hematoporphyrin resulted in a substantial number of long-term cures. Since hematoporphyrin is preferentially retained in tumor tissue, selective tumor destruction could be obtained. Light penetration studies and the high efficiency of this technique indicated its applicability even to certain deep-seated human tumors.     

Inducing effect of hematoporphyrin derivative (HpD) on cell sister chromatid exchanges (SCE) in vitro

The mutagenic effect of HpD on cell SCE and the reactions of cell SCE to different sources of light combined with HpD were studied using V79 cells. There were 6 doses of HpD: 1 microgram/ml, 3 micrograms/ml, 5 micrograms/ml, 10 micrograms/ml, 50 micrograms/ml and 100 micrograms/ml. The dose of 5 micrograms/ml is equal to the maximum dose of HpD used in the clinic (HpD per milliliter of patient's blood). Our experiments demonstrated that when the cells were cultured in the dark and HpD was added to the medium no more than 5 micrograms/ml, the SCE frequencies were not increased. The cells were irradiated with different sources of light without HpD, both the fluorescence and ultraviolet light could promote SCE but the light of daylight lamp and red light did not increase it. But when HpD was added into culture medium at the dose of less than 5 micrograms/ml, every light could increase the cell SCE intensively except the daylight lamp light. The red light was more notable than the others by relation analysis.     

Time dependence of hematoporphyrin distribution in selected tissues of normal rats and in ascites hepatoma.

The distribution of hematoporphyrin was determined in normal rats and in rats bearing ascites hepatoma as a function of time after i.p. injection of 10-20 mg/kg of dye. In both cases, hematoporphyrin displayed a high affinity for the tumor cells. At 20 mg/kg, the maximum difference between the amount of hematoporphyrin accumulated in the tumor and in the liver was obtained at 12 h after injection (tumor/liver ratio = 28). Our results suggest that hematoporphyrin is almost exclusively metabolized in the liver and excreted via the biliary tract, whereas only minor amounts are metabolized in the tumor cells. Moreover, the binding between the porphyrin and tumor cells is competitive with serum protein binding.     

Candida albicans inactivation and cell membrane integrity damage by microwave irradiation

In indicating the microwave irradiation for disinfecting dentures it is necessary to see how this procedure influences Candida albicans integrity and viability. The aim of this study was to evaluate the ability of microwaves to inactivate C. albicans and damage cell membrane integrity. Two 200-ml C. albicans (ATCC 10231) suspensions were obtained. A sterile denture was placed in a beaker containing the Experimental (ES) or the Control suspension (CS). ES was microwaved at 650 W for 6 min. Suspensions were optically counted using methylene blue dye uptake as indicative of membrane-damaged cells; spread on Agar Sabouraud dextrose (ASD) for viability assay; or spectrophotometrically measured at 550 nm. Cell-free solutions were submitted to content analyses of protein (Bradford and Pyrogallol red methods); Ca++ (Cresolftaleine complexone method); DNA (spectrophotometer measurements at 260 nm) and K+ (selective electrode technique). Data were analysed by Student's t- or Wilcoxon z-tests (alpha = 0.05). All ES cells demonstrated cell membrane damage. Viable cells were non-existent in the ES ASD plates. No significant difference in optical density between ES and CS was observed (P=0.272). ES cells released significantly high protein (P<0.001, Bradford; P=0.005, Pyrogallol red), K+ (P<0.001), Ca++ (P=0.012) and DNA (P=0.046) contents. Microwaves inactivated C. albicans and damaged cell membrane integrity.     

Comparison of photodynamic inactivation of experimental stomach tumors sensitized by acridine orange or hematoporphyrin derivatives

The photodynamic inactivations of Walker carcinosarcoma 256 stomach tumors by the concomitant use of acridine orange (AO) and argon laser, and by the combined use of hematoporphyrin derivatives (HPD) and dye laser were compared. Wistar rats bearing stomach tumors of 4-6 mm in diameter 5-10 days after their implantation were injected intraperitoneally with 40 mg/kg body weight of AO or HPD, and 24 h later their stomach tumors were exposed to argon laser at 488 nm or dye laser at 630 nm, respectively, at an intensity of 15 mW/cm2 for 20 min. Temperature rise was less than 3 degrees C during irradiation. Seven days after irradiation, complete or partial necrosis with sparing of the surrounding mucosa was seen histologically in all rats treated by the two therapy methods. Phase contrast and electron microscopy showed nuclear pyknosis and damage to the inner layer of the nuclear envelope in tumor cells treated with AO and argon laser, while cytotoxicity involved damage to the outer layer of the nuclear envelope and intracytoplasmic organelles in tumor cells treated with HPD and dye laser. This difference between AO and HPD is considered to be a difference in their intracellular localization in tumor cells.