5-脂氧合酶抑制剂对黑色素瘤细胞a375转移的抑制作用
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摘要
黑色素瘤是起源于皮肤色素细胞的一种恶性肿瘤，易发生转移，目前尚无特异有效药物治疗其扩散性病变。流行病学研究表明，饮食中多不饱和脂肪酸含量与黑色素瘤的发生密切相关。花生四烯酸是多不饱和脂肪酸的主要成分，其脂氧合酶代谢产物在黑色素瘤的转移过程中起重要作用。本研究通过考察5-脂氧合酶抑制剂对黑色素瘤细胞a375转移过程中相关因素的影响，阐明5-脂氧合酶抑制剂影响黑色素瘤转移的作用机制，为研制开发抗黑色素瘤转移药物提供思路。
实验采用光密度法测定黑色素瘤细胞a375与细胞外基质蛋白的粘附率：以流式细胞仪测定a375细胞表面整合素的表达；以酶联免疫吸附法、流式细胞仪、蛋白质印迹分析测定a375细胞ICAM-1的表达，以Northern blot测定a375细胞ICAM-1的mRNA的表达水平；以双抗体夹心法检测a375细胞NF-κB的活化水平。
以滤膜涂布Matrigel基质的Transwell Chamber测定a375细胞的侵袭能力；以明胶酶谱法测定a375细胞分泌MMP-2和MMP-9的酶活性；以蛋白质印迹分析测定a375细胞产生MMP-2、MMP-9和TIMP-1的蛋白量；以Northern blot测定a375细胞中MMP-2、MMP-9和TIMP-1的mRNA表达水平。
实验结果表明：作为对照的环氧合酶抑制剂吲哚美辛作用后，对黑色素瘤细胞a375与细胞外基质的粘附率没有影响。在5-20μM浓度范围内，非选择性脂氧合酶抑制剂NDGA、5-脂氧合酶抑制剂AA861和5-脂氧合酶激动蛋白抑制剂MK886剂量依赖性地抑制a375细胞对Ⅳ型胶原的粘附，对细胞粘附至纤维粘连蛋白及层粘连蛋白没有影响。抑制剂对a375细胞粘附的抑制作用能被5-脂氧合酶代谢产物5-HETE所逆转，白三烯类对a375细胞粘附性改变无明显作用。
作用浓度为20μM时，NDGA、AA861及MK886能明显抑制细胞内[Ca~(2+)]升高导致的a375细胞对Ⅳ型胶原粘附率的增加，对a375细胞PKC活化导致对Ⅳ型胶原粘附增加只有部分抑制作用。
相同浓度的NDGA、AA861及MK886作用后，明显降低a375细胞表面的整合素β_1表达，对细胞整合素α_(Ⅱb)β_3的表达基本没有影响。
吲哚美辛对a375细胞ICAM-1表达没有明显影响，在5～20μM浓度范围内，NDGA、AA861及MK886剂量依赖性地降低a375细胞ICAM-1蛋白表达量及mRNA
表达水平。NDGA、AA861及MK886作用后，剂量依赖性地抑制a375细胞NF a
B活化。
叼噪美辛对a375细胞侵袭重组基底膜能力没有明显影响，NDGA、AA861 和
MK886剂量依赖性地抑制a375细胞对Matytael膜的侵袭。这种抑制作用能被5－HETE
逆转，白三烯类对0乃细胞的侵袭性改变没有明显影响。
作用浓度为20 p M时，NDGA、AA861及MK886显著抑制a375细胞产生MMP－2
和 MMP－9蛋白量及其InRNA表达水平，对a375细胞产生 TIMPI的蛋白量及mRNA
表达水平没有明显影响。
这些结果表明，5．脂氧合酶代谢产物的其中一种，5－HETE，在黑色素瘤转移过
程中起重要的作用。证实5－脂氧合酶抑制剂通过抑制a375细胞核转录因子活化，在
基因转录水平抑制a3 75细胞粘附分子的表达，进而抑制a375细胞对细胞外基质的
粘附。5－脂氧合酶抑制剂通过抑制 a375细胞产生、分泌IV型胶原酶，降低 a375细
胞对重组基底膜的侵袭性。
本研究结果提示5．脂氧合酶抑制剂作为可能的抗黑色素瘤转移药物，值得深入
研究。
Melanoma is a kind of malignant tumor originating from pigment cells of skin and
prone to metastasis. So far no specific drug has been aPplied to control the diffused
pathological process. Epidemiological studies show that the content of polyunsaturated
fatty acids in diet were closely associated with incidence of malignant melanoma.
Arachidonic acid is the main ingredients of polyunsaturated fatty acids and its
lipoxygenase metabolites play a key role in the metastasis process of melanoma cancer
cells. We conducted experiments trying to stlldy the effects and mechanism of 5-
lipoxygenase inhibitors on metastasis of human melanoma cell a375 and to obtain the
clue of 5-lipoxygenase inhibitor as poteniial anti-metastatic agents for melanoma.
Adhesion of a375 cell to extracellular matrix was determined by optical density
assay. The expression of integrins on cancer cell surface was determined by fiow
cytometry ELlSA, flow cytometry and Western blot was used to detect the production of
lCAM-l in a375 cell. Northern blot was used to measure the expressing level of ICAM-1
mRNA in a375 cell. The 1evels of NF- K B in a375 cell was measured by sandwich
ELISA.
Invasiveness of a375 cells was assayed by using Transwell chamber wth filter
membrane coated with Matrigel. The secretion of active matrix metalloproteinases
(MMP-2 and MMP-9) was measured by gelatin zymogram. Western blot was used to
detect the production of MMP-2, MMP-9 and tissues inhibitors of metalloproteinase
(TIMP-l) and the expressing leveI of MMP-2, MMP-9 and TIMP-l mKNA was
measured by Northern blot.
The results showed that in control grouP treated with indomethacin, a
cyclooxygenase inhibitors, adhesion of a375 cells to extracellular matrix was not altered.
In the range from 5~20 ll M, lipoxygenase inhibitors nordihydroguaiaretic acid (NDGA),
5-lipoxygenase inhibitors AA861 and specific 5-lipoxygenase-activating-prote inhibitor
MK886 could all inhibit adhesion of a375 cell to collagen IVin a dose-dependent manner
and had no effect on adhesion to fibronectin or vitronectin. 5-lipoxygenase metabolites
(5-HETE) reversed the adhesion behavior of inhibitor-treat cancer cell and leukotrienes
had no effect.
Pretreated with 20 ll M of NDGA, AA861 and MK886 remarkably inhibited the
A23l87-induced enhancement of a375 cell adhesion to collagen IV respectively and
only partly inhibited the TPA-induced enhancement.
A375 cells tfeated with NDGA, AA861 and MK886 in the same concentfation
exhibited reduction of the expression of integrin 6 l on the surface of melanoma cell, but
not of integrin Q lIb fl 3'
Indomechacin had no effect on the production of ICAM-l protein in a375 cell.
NDGA, AA861 and MK886 could reduce the production of ICAM-l protein and its
mRNA of a375 cell and suPpress NF- K B activation in a dose-dependeni manner in the
range from 5~20 ll M.
No significant inhibition of invasiveness to recombinant basement membrane was
seen when a375 ceIls were treated with indomethacin. In the range from 5~20ll M
NDGA, AA861 and MK886 significantly inhibited invasiveness of a375 cell through
Matrigel membrane. 5-HETE could reverse this inhibitory effect and leukotrienes had no
effect.
NDGA, AA86l and MK886 could significantly inhibit production of MMP-2 and
MMP-9 proteins as well as their mRNA expression level. The inhibition of 5-
lipoxygenase activity with the shese inhibitors had no significant effect on production of
TIMP-l protein and mRNA level in a375 ceII.
These results suggest that one of the 5-lipoxygenase metabolites, 5-HETE, plays an
important role in the metastatic progress of melanoma. 5-lipoxygenase inhibitors can
inhibit the process by suPpressing the expression of adhesion molecule through inhibiting
the nuclear factor activation of a375 cell so as to inhibit a375 cell adhesion to
extracel1u1ar matrix and by suPPressing the production of MMP-2 and MMP-9 in a375
cell so as to decrease the invasiveness to through recombinant basement membrane.
That
实验采用光密度法测定黑色素瘤细胞a375与细胞外基质蛋白的粘附率：以流式细胞仪测定a375细胞表面整合素的表达；以酶联免疫吸附法、流式细胞仪、蛋白质印迹分析测定a375细胞ICAM-1的表达，以Northern blot测定a375细胞ICAM-1的mRNA的表达水平；以双抗体夹心法检测a375细胞NF-κB的活化水平。
以滤膜涂布Matrigel基质的Transwell Chamber测定a375细胞的侵袭能力；以明胶酶谱法测定a375细胞分泌MMP-2和MMP-9的酶活性；以蛋白质印迹分析测定a375细胞产生MMP-2、MMP-9和TIMP-1的蛋白量；以Northern blot测定a375细胞中MMP-2、MMP-9和TIMP-1的mRNA表达水平。
实验结果表明：作为对照的环氧合酶抑制剂吲哚美辛作用后，对黑色素瘤细胞a375与细胞外基质的粘附率没有影响。在5-20μM浓度范围内，非选择性脂氧合酶抑制剂NDGA、5-脂氧合酶抑制剂AA861和5-脂氧合酶激动蛋白抑制剂MK886剂量依赖性地抑制a375细胞对Ⅳ型胶原的粘附，对细胞粘附至纤维粘连蛋白及层粘连蛋白没有影响。抑制剂对a375细胞粘附的抑制作用能被5-脂氧合酶代谢产物5-HETE所逆转，白三烯类对a375细胞粘附性改变无明显作用。
作用浓度为20μM时，NDGA、AA861及MK886能明显抑制细胞内[Ca~(2+)]升高导致的a375细胞对Ⅳ型胶原粘附率的增加，对a375细胞PKC活化导致对Ⅳ型胶原粘附增加只有部分抑制作用。
相同浓度的NDGA、AA861及MK886作用后，明显降低a375细胞表面的整合素β_1表达，对细胞整合素α_(Ⅱb)β_3的表达基本没有影响。
吲哚美辛对a375细胞ICAM-1表达没有明显影响，在5～20μM浓度范围内，NDGA、AA861及MK886剂量依赖性地降低a375细胞ICAM-1蛋白表达量及mRNA
表达水平。NDGA、AA861及MK886作用后，剂量依赖性地抑制a375细胞NF a
B活化。
叼噪美辛对a375细胞侵袭重组基底膜能力没有明显影响，NDGA、AA861 和
MK886剂量依赖性地抑制a375细胞对Matytael膜的侵袭。这种抑制作用能被5－HETE
逆转，白三烯类对0乃细胞的侵袭性改变没有明显影响。
作用浓度为20 p M时，NDGA、AA861及MK886显著抑制a375细胞产生MMP－2
和 MMP－9蛋白量及其InRNA表达水平，对a375细胞产生 TIMPI的蛋白量及mRNA
表达水平没有明显影响。
这些结果表明，5．脂氧合酶代谢产物的其中一种，5－HETE，在黑色素瘤转移过
程中起重要的作用。证实5－脂氧合酶抑制剂通过抑制a375细胞核转录因子活化，在
基因转录水平抑制a3 75细胞粘附分子的表达，进而抑制a375细胞对细胞外基质的
粘附。5－脂氧合酶抑制剂通过抑制 a375细胞产生、分泌IV型胶原酶，降低 a375细
胞对重组基底膜的侵袭性。
本研究结果提示5．脂氧合酶抑制剂作为可能的抗黑色素瘤转移药物，值得深入
研究。
Melanoma is a kind of malignant tumor originating from pigment cells of skin and
prone to metastasis. So far no specific drug has been aPplied to control the diffused
pathological process. Epidemiological studies show that the content of polyunsaturated
fatty acids in diet were closely associated with incidence of malignant melanoma.
Arachidonic acid is the main ingredients of polyunsaturated fatty acids and its
lipoxygenase metabolites play a key role in the metastasis process of melanoma cancer
cells. We conducted experiments trying to stlldy the effects and mechanism of 5-
lipoxygenase inhibitors on metastasis of human melanoma cell a375 and to obtain the
clue of 5-lipoxygenase inhibitor as poteniial anti-metastatic agents for melanoma.
Adhesion of a375 cell to extracellular matrix was determined by optical density
assay. The expression of integrins on cancer cell surface was determined by fiow
cytometry ELlSA, flow cytometry and Western blot was used to detect the production of
lCAM-l in a375 cell. Northern blot was used to measure the expressing level of ICAM-1
mRNA in a375 cell. The 1evels of NF- K B in a375 cell was measured by sandwich
ELISA.
Invasiveness of a375 cells was assayed by using Transwell chamber wth filter
membrane coated with Matrigel. The secretion of active matrix metalloproteinases
(MMP-2 and MMP-9) was measured by gelatin zymogram. Western blot was used to
detect the production of MMP-2, MMP-9 and tissues inhibitors of metalloproteinase
(TIMP-l) and the expressing leveI of MMP-2, MMP-9 and TIMP-l mKNA was
measured by Northern blot.
The results showed that in control grouP treated with indomethacin, a
cyclooxygenase inhibitors, adhesion of a375 cells to extracellular matrix was not altered.
In the range from 5~20 ll M, lipoxygenase inhibitors nordihydroguaiaretic acid (NDGA),
5-lipoxygenase inhibitors AA861 and specific 5-lipoxygenase-activating-prote inhibitor
MK886 could all inhibit adhesion of a375 cell to collagen IVin a dose-dependent manner
and had no effect on adhesion to fibronectin or vitronectin. 5-lipoxygenase metabolites
(5-HETE) reversed the adhesion behavior of inhibitor-treat cancer cell and leukotrienes
had no effect.
Pretreated with 20 ll M of NDGA, AA861 and MK886 remarkably inhibited the
A23l87-induced enhancement of a375 cell adhesion to collagen IV respectively and
only partly inhibited the TPA-induced enhancement.
A375 cells tfeated with NDGA, AA861 and MK886 in the same concentfation
exhibited reduction of the expression of integrin 6 l on the surface of melanoma cell, but
not of integrin Q lIb fl 3'
Indomechacin had no effect on the production of ICAM-l protein in a375 cell.
NDGA, AA861 and MK886 could reduce the production of ICAM-l protein and its
mRNA of a375 cell and suPpress NF- K B activation in a dose-dependeni manner in the
range from 5~20 ll M.
No significant inhibition of invasiveness to recombinant basement membrane was
seen when a375 ceIls were treated with indomethacin. In the range from 5~20ll M
NDGA, AA861 and MK886 significantly inhibited invasiveness of a375 cell through
Matrigel membrane. 5-HETE could reverse this inhibitory effect and leukotrienes had no
effect.
NDGA, AA86l and MK886 could significantly inhibit production of MMP-2 and
MMP-9 proteins as well as their mRNA expression level. The inhibition of 5-
lipoxygenase activity with the shese inhibitors had no significant effect on production of
TIMP-l protein and mRNA level in a375 ceII.
These results suggest that one of the 5-lipoxygenase metabolites, 5-HETE, plays an
important role in the metastatic progress of melanoma. 5-lipoxygenase inhibitors can
inhibit the process by suPpressing the expression of adhesion molecule through inhibiting
the nuclear factor activation of a375 cell so as to inhibit a375 cell adhesion to
extracel1u1ar matrix and by suPPressing the production of MMP-2 and MMP-9 in a375
cell so as to decrease the invasiveness to through recombinant basement membrane.
That
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53. Nigam Ak, Savage FJ, Boulos BB, et al. Loss of cell-cell and -matrix adhesion molecules in colorectal cancer. Br. J. Cancer, 1993,68(3):507-514
54. Simone F, Frank S, Dirk R, et al. Phorbol ester-induced shedding of intercellular adhesion molecule-1 on erythroleukemic K562 cells. Biochim. Biophys. Acta., 1996, 1312:255-261
55. Araki T, Miki C, Kusunoki M. Biological implications of circulating soluble intercellular adhesion molecule-1 in colorectal cancer patients. Scand J. Gastroenterol, 2001, 36(4):399-404
56. Sunami T, Yashiro M, Chung KH. ICAM-1 gene transfection inhibits lymph node metastasis by human gastric cancer cells. JPN. J Cancer Res., 2000, 91(9):925-933
57．孙婧瑾，周信达，刘银坤，等．肝癌组织中细胞间粘着分子-1的表达与肝癌侵袭转移关系的研究．中国癌症杂志，1997，7(3)：161-164
58．曲增强，吴孟超，谢天培，等．Ⅰ型细胞间粘附分子在肝细胞癌中的表达及其意义．中华病理学杂志，1997，26(2)：82-84
59. Anastassiou G, Schilling H, Stang A, et al. Expression of the cell adhesion molecules ICAM-1, VCAM-1 and NCAM in uveal melanoma: a clinicopathological study. Oncology, 2000, 58(1):83-88
60. Johnson JP. Cell adhesion molecules in the development and progression of malignant melanoma. Cancer Metastasis Rev., 1999, 18(3):345-357
61. Morandini R, Boeynaems JM, Hedley SJ, et al. Modulation of ICAM-1 expression by alpha-MSH in human melanoma cells and melanocytes. J. Cell Physiol. 1998, 175(3):276-282.
62. Natali P, Nicotra MR, Carvaliere R, et al. Differential expression of intercellular adhesion molecule-1 in primary and metastatic melanoma lesions. Cancer Res., 1990, 50:1271-1278
63. Tozawa K, Sakurada S, Kohri K. Effects of anti-nuclear factor κ B reagents in blocking adhesion of human cancer cell to vascular endothelial cells. Cancer Res., 1995, 55:4162-4167
64. Karmann K, Wang M, Fanslow WC, et al. Activation and homologous desensitization of human endothelial cells by CD40 ligand, tumor and interleukin-1. J. Exp. Med., 1996, 184:173-182
65. Finco TS, Baldwin AS. Mechanistic aspects of NF-kappa B regulation: the emerging role of phosphorylation and proteolysis. Immunity, 1995,3(3):263-272
66. Baldwin AS. The NF-κ B and Iκ B protein: new discoveries and insights. Annu. Rev. Immunol, 1996, 14:649-681
67. Sherri L, Katherine AF, Graham CN, et al. Inhibition of 5-lipoxygense blocks IL-1 β-induced vascular adhesion molecule-1 gene expression in human endothelial cells. J. Immun., 1997, 158:3401-3407
68. Brad HR, Jennifer AD, Laura CT, et al. Activation of nuclear factor-κ B correlate with MCP-1 expression by human mesangial cells. Kidney Int., 1995, 48:1263-1271
69. Gou FK, Li YL, Wu SG. Antisense IRAK-2 oligonuleotide blocks IL-1-stimilated NF-κ B activation and ICAM-1 expression in cultured endothelial cells. Inflammation, 1999,23(6):535-543
70. Stetler-Stevenson WG, Aznavoorian S, Liotta LA. Tumor cell interations with the extracelluar matrix during invasion and metastasis. Annu. Rev Cell Biol., 1993, 9:541-573
71. Mignatti P, Rifkin DB. Biology and biochemistry of proteinases in tumor invasion. Physiol. Rev., 1993,73:161-195
72. Reuven R, Erik WT, Yukihide I, et al. Effects of inhibitors of plasminogen activator, serine proteinases, and collagenase Ⅳ on the invasion of basement membranes by metastatic cells. Cancer Res., 1988,48:3307-3312
73. Albini A, Iwamoto Y, Kleinman HK, et al. A rapid in vitro assay for quantitating the invasive potential of tumor cells. Cancer Res., 1987, 47:3239-3245
74. Fadwa AA, Patricia MF, Ashani TW, et al. Inhibitors of prostaglandin synthesis inhibit human prostate tumor cell invasiveness and reduce the release of matrix metalloproteinases. Cancer Res., 2000, 60:4629-4637
75. Aznavoorian S, Stetler-Stevenson WG, Liotta LA. Molecular aspects of tumor cell invasion and metastasis. Cancer, 1993,71 (4): 1368-1383.
76. Stetler-Stevenson WG, Liotta LA, Kleiner DE, et al. Role of matrix metalloproteinase in tumor invasion and metastasis. FASEB J., 1993, 7:1434-1441
77. Davis B, Miles DW, Happerfield LC, et al. Activity of type Ⅳ collagenase in benign and malignant breast disease. Br. J. Cancer, 1993, 67:1126-1131
78．卜文，汤钊猷，叶胜龙，等．Ⅳ型胶原酶与肝细胞癌侵袭转移性的关系．中华消化杂志，1999，19(1)：13-15
79．李红梅，方伟岗，郑杰，等．不同转移潜能的人肿瘤细胞系金属蛋白酶活性分析．中华病理学杂志．1998，27(5)：341-343
80. Celentano DC, Frishman WH. Matrix metalloproteinases and coronary artery disease: a novel therapeutic target. J. Clin. Pharmacol, 1997, 37(11):991-1000
81. Edwards DR, Beaudry PP, Laing TD, et al. The roles of tissue inhibitors of metalloproteinases in tissue remodelling and cell growth. Int. J. Obes. Relat. Metab. Disord., 1996, 20(3):9-15
82．曾仲．基质金属蛋白酶组织抑制物的生物学功能．国外医学分子生物学分册，2000，22(2)：84-87
83. Conway JG, Trexler SJ, Wakefiels JA, et al. Effect of matrix metalloproteinase inhibitors on tumor growth and spontaneous metastasis. Clin. Exp. Metastasis, 1996, 14:115-124
84. Yeow KM, Phillips BW, Beaudry PP, et al. Expression of MMPs and TIMPs in mammalian cells. Methods Mol Biol., 2001, 151:181-189
85．史宏男，何荣根，周晓健．金属蛋白酶及其组织抑制剂在涎腺腺样囊性癌细胞株表达的分子生物学研究．中华口腔医学杂志，1997，32(6)：323-325
86．丁挺波，杨渝珍，韩玲，等．基质金属蛋白酶活性测定的改良底物胶电泳法．同济医科大学学报，2000，29(1)：43-48
87．卜文，黄晓武，汤钊猷．基质金属蛋白酶-2与肝细胞癌侵袭转移性的关系．中华医学杂志，1997，77(9)：661-663
88. David WA, Janica M, Bernard HD. Lipoxygenase inhibitors block PDGF-induced mitogenesis: a MARK-independent mechanism that blocks fos and egr. Am J. Physiol., 1995,268(37):604-610
89. Musser JH, Kreft AF. 5-lipoxygenase: properties, pharmacology, and the quinolinyl (bridged)aryl class of inhibitors. J. Med. Chem., 1992, 35(14):2501-2524
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40．田臻，张伟国．粘附机制与口腔粘膜鳞状细胞癌侵袭和转移关系的研究进展．口腔额面外科杂志，2001，11(1)：53-56
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51. Hynes RO, Lander AD. Contact and adhesive specificites in the associations, migration and targeting of cells and axons. Cell, 1992, 68(2):303-322
52. Staunton DE, Marlin SD, Stratowa C, et al. Primary structure of ICAM-1 demonstrates interaction between members of the immunoglobulin and integrin supergene families. Cell, 1988, 52:925-933
53. Nigam Ak, Savage FJ, Boulos BB, et al. Loss of cell-cell and -matrix adhesion molecules in colorectal cancer. Br. J. Cancer, 1993,68(3):507-514
54. Simone F, Frank S, Dirk R, et al. Phorbol ester-induced shedding of intercellular adhesion molecule-1 on erythroleukemic K562 cells. Biochim. Biophys. Acta., 1996, 1312:255-261
55. Araki T, Miki C, Kusunoki M. Biological implications of circulating soluble intercellular adhesion molecule-1 in colorectal cancer patients. Scand J. Gastroenterol, 2001, 36(4):399-404
56. Sunami T, Yashiro M, Chung KH. ICAM-1 gene transfection inhibits lymph node metastasis by human gastric cancer cells. JPN. J Cancer Res., 2000, 91(9):925-933
57．孙婧瑾，周信达，刘银坤，等．肝癌组织中细胞间粘着分子-1的表达与肝癌侵袭转移关系的研究．中国癌症杂志，1997，7(3)：161-164
58．曲增强，吴孟超，谢天培，等．Ⅰ型细胞间粘附分子在肝细胞癌中的表达及其意义．中华病理学杂志，1997，26(2)：82-84
59. Anastassiou G, Schilling H, Stang A, et al. Expression of the cell adhesion molecules ICAM-1, VCAM-1 and NCAM in uveal melanoma: a clinicopathological study. Oncology, 2000, 58(1):83-88
60. Johnson JP. Cell adhesion molecules in the development and progression of malignant melanoma. Cancer Metastasis Rev., 1999, 18(3):345-357
61. Morandini R, Boeynaems JM, Hedley SJ, et al. Modulation of ICAM-1 expression by alpha-MSH in human melanoma cells and melanocytes. J. Cell Physiol. 1998, 175(3):276-282.
62. Natali P, Nicotra MR, Carvaliere R, et al. Differential expression of intercellular adhesion molecule-1 in primary and metastatic melanoma lesions. Cancer Res., 1990, 50:1271-1278
63. Tozawa K, Sakurada S, Kohri K. Effects of anti-nuclear factor κ B reagents in blocking adhesion of human cancer cell to vascular endothelial cells. Cancer Res., 1995, 55:4162-4167
64. Karmann K, Wang M, Fanslow WC, et al. Activation and homologous desensitization of human endothelial cells by CD40 ligand, tumor and interleukin-1. J. Exp. Med., 1996, 184:173-182
65. Finco TS, Baldwin AS. Mechanistic aspects of NF-kappa B regulation: the emerging role of phosphorylation and proteolysis. Immunity, 1995,3(3):263-272
66. Baldwin AS. The NF-κ B and Iκ B protein: new discoveries and insights. Annu. Rev. Immunol, 1996, 14:649-681
67. Sherri L, Katherine AF, Graham CN, et al. Inhibition of 5-lipoxygense blocks IL-1 β-induced vascular adhesion molecule-1 gene expression in human endothelial cells. J. Immun., 1997, 158:3401-3407
68. Brad HR, Jennifer AD, Laura CT, et al. Activation of nuclear factor-κ B correlate with MCP-1 expression by human mesangial cells. Kidney Int., 1995, 48:1263-1271
69. Gou FK, Li YL, Wu SG. Antisense IRAK-2 oligonuleotide blocks IL-1-stimilated NF-κ B activation and ICAM-1 expression in cultured endothelial cells. Inflammation, 1999,23(6):535-543
70. Stetler-Stevenson WG, Aznavoorian S, Liotta LA. Tumor cell interations with the extracelluar matrix during invasion and metastasis. Annu. Rev Cell Biol., 1993, 9:541-573
71. Mignatti P, Rifkin DB. Biology and biochemistry of proteinases in tumor invasion. Physiol. Rev., 1993,73:161-195
72. Reuven R, Erik WT, Yukihide I, et al. Effects of inhibitors of plasminogen activator, serine proteinases, and collagenase Ⅳ on the invasion of basement membranes by metastatic cells. Cancer Res., 1988,48:3307-3312
73. Albini A, Iwamoto Y, Kleinman HK, et al. A rapid in vitro assay for quantitating the invasive potential of tumor cells. Cancer Res., 1987, 47:3239-3245
74. Fadwa AA, Patricia MF, Ashani TW, et al. Inhibitors of prostaglandin synthesis inhibit human prostate tumor cell invasiveness and reduce the release of matrix metalloproteinases. Cancer Res., 2000, 60:4629-4637
75. Aznavoorian S, Stetler-Stevenson WG, Liotta LA. Molecular aspects of tumor cell invasion and metastasis. Cancer, 1993,71 (4): 1368-1383.
76. Stetler-Stevenson WG, Liotta LA, Kleiner DE, et al. Role of matrix metalloproteinase in tumor invasion and metastasis. FASEB J., 1993, 7:1434-1441
77. Davis B, Miles DW, Happerfield LC, et al. Activity of type Ⅳ collagenase in benign and malignant breast disease. Br. J. Cancer, 1993, 67:1126-1131
78．卜文，汤钊猷，叶胜龙，等．Ⅳ型胶原酶与肝细胞癌侵袭转移性的关系．中华消化杂志，1999，19(1)：13-15
79．李红梅，方伟岗，郑杰，等．不同转移潜能的人肿瘤细胞系金属蛋白酶活性分析．中华病理学杂志．1998，27(5)：341-343
80. Celentano DC, Frishman WH. Matrix metalloproteinases and coronary artery disease: a novel therapeutic target. J. Clin. Pharmacol, 1997, 37(11):991-1000
81. Edwards DR, Beaudry PP, Laing TD, et al. The roles of tissue inhibitors of metalloproteinases in tissue remodelling and cell growth. Int. J. Obes. Relat. Metab. Disord., 1996, 20(3):9-15
82．曾仲．基质金属蛋白酶组织抑制物的生物学功能．国外医学分子生物学分册，2000，22(2)：84-87
83. Conway JG, Trexler SJ, Wakefiels JA, et al. Effect of matrix metalloproteinase inhibitors on tumor growth and spontaneous metastasis. Clin. Exp. Metastasis, 1996, 14:115-124
84. Yeow KM, Phillips BW, Beaudry PP, et al. Expression of MMPs and TIMPs in mammalian cells. Methods Mol Biol., 2001, 151:181-189
85．史宏男，何荣根，周晓健．金属蛋白酶及其组织抑制剂在涎腺腺样囊性癌细胞株表达的分子生物学研究．中华口腔医学杂志，1997，32(6)：323-325
86．丁挺波，杨渝珍，韩玲，等．基质金属蛋白酶活性测定的改良底物胶电泳法．同济医科大学学报，2000，29(1)：43-48
87．卜文，黄晓武，汤钊猷．基质金属蛋白酶-2与肝细胞癌侵袭转移性的关系．中华医学杂志，1997，77(9)：661-663
88. David WA, Janica M, Bernard HD. Lipoxygenase inhibitors block PDGF-induced mitogenesis: a MARK-independent mechanism that blocks fos and egr. Am J. Physiol., 1995,268(37):604-610
89. Musser JH, Kreft AF. 5-lipoxygenase: properties, pharmacology, and the quinolinyl (bridged)aryl class of inhibitors. J. Med. Chem., 1992, 35(14):2501-2524