转化生长因子-β和干扰素-γ对黑色素瘤细胞增殖迁移和侵袭的影响作用研究

˖ڍᐸც˙ࣽ 2010Ꭰじ37ࢣじ3᱋ 䕀࣪⫳䭓಴ᄤ�£੠ᑆᡄ㋴�¤ᇍ咥㡆㋴⯸㒚㚲 ๲Ⅺ䖕⿏੠։㺁ⱘᕅડ԰⫼ⷨお� 㨷ᄺᯧķ স ᔎ ᄭ ⍯ 䍉 ἴ 䍉⾔݄ ׾᯹⫳ 䔺 ࿰ ᄭֱᄬĸ ๕≑ࡧ⻽๓႒⫱⤲᪅ⵀა 喋๕≑ጮ ������喌 ᦄ㺭 Ⱊ⮰喝㻮ᄋ䒘ࡂ⩋䪫ఌၼ-Ų喋TGF-Ų喌স᎞វ㉌-ų喋IFN-ų喌ᄥ叽㞞㉌⭐㏲㘊⮰්⃂ȟ䓭⼧সӡ㷙㘩߇⮰ ᒝ৹Ƞ᫥∁喝ѿโഥڧ叽㞞㉌⭐㏲㘊喏ᒱ㏲㘊䪫ݜ 80%㲹ऴᏒᬢ喏ߌڑ TGF-Ų喋≿Ꮢͦ 5ng/mL喌স IFN-ų喋≿Ꮢͦ 10ng/mL喌ఌၼ喏ᄲ叽㞞㉌⭐㏲㘊ܲͦᄥ⚓㏰ȟTGF-Ų㏰ȟIFN-ų㏰সTGF-Ų+IFN-ų㏰喏♢ऺर㏰ܲݗ䛳⩔⸦䚜㒃ͥᬺ B喋SRB喌් ⃂≷჆∁㻮ᄋ叽㞞㉌⭐㏲㘊ഥڧऺ8hȟ16hȟ24hȟ32hȟ40hস48hᬢ䬠◥⮰්⃂ᗱۡ喏κ540nm।ٵᏒ̷℀ 㞞䄧ं।ٵᏒ喋A540喌Ბ㶔⹦㏲㘊⮰්⃂ऄࡂȠ䛳⩔200żL।๠ᄥर㏰㏲㘊⮰㶔䲎䔇㵸ܾ⬁ะ⤲喏䮻ऺκ12h㻮ᄋ ܾ⬁ͷ䬠䌉⻧喋࢟Џ㶔㗫⭐㏲㘊䓭⼧⮰㘩߇喌⮰ऄࡂȠ䛳⩔≷Transwell㈧㐋ႀโ㛈̶㏲㘊⮰595nm।ٵᏒ喋A595喌 Ბ㶔⹦㗫⭐㏲㘊⮰ӡ㷙㘩߇Ƞ䛳⩔ᬺ㘢䚢䅝∁㻮ᄋर㏰㗫⭐㏲㘊⮰MMP-2ȟMMP-9≧ᕓⅠ᎟喏⩔ప׻ܲ᲼䒛Т≷ ჆㖆ͅ☛䚜㘦܉㘢͙MMP-2ȟMMP-9⮰᲍ፒ▜Ꮢ喋Џ㶔MMPs⮰≧ᕓⅠ᎟喌Ƞ㏿᳈喝TGF-ŲछБ්ᑦ叽㞞㉌⭐㏲㘊 ⮰්⃂ȟ䓭⼧সӡ㷙㘩߇喋P<0.05喌喏IFN-ų݅ួݢ叽㞞㉌⭐㏲㘊⮰්⃂ȟ䓭⼧সӡ㷙㘩߇喋P<0.05喌喏θ㔱सᬢะ⤲݅ ᄥ叽㞞㉌⭐㏲㘊⮰ᒝ৹҈⩔ᓚᑝᝂ⊴๝喋P>0.05喌Ƞ㏿䃦喝ѿโ჊侸͙喏TGF-ŲछБ᎞វ IFN-ųᄥ叽㞞㉌⭐㏲㘊⮰් ⃂ȟ䓭⼧সӡ㷙㘩߇⮰ួݢ҈⩔喏᱘ⵀ⾢छͦ►⫳স㗫⭐⮰Ⱔξڟ㈧ⵀ⾢ຌ჆჊侸ദ⵬Ƞ ڟ䩚䃹 叽㞞㉌⭐ 䒘ࡂ⩋䪫ఌၼ-Ų ᎞វ㉌-ų ්⃂ 䓭⼧ ӡ㷙 doi:10.3969/j.issn.1000-8179.2010.03.004 &GGFDUT PG 5('�Ų BOE *'/�ų PO UIF 1SPMJGFSBUJPO .JHSBUJPO BOE *OWBTJPO PG .FMBOPNB $FMMT DONG Xueyi, GU Qiang, SUN Tao, ZHAO Nan, ZHAO Xiulan, NI Chunsheng, CHE Na, SUN Baocun Corresponding author: SUN Baocun, E-mail: sunbaocun@yahoo.com.cn 1Department of Pathology, Tianjin Medical University, Tianjin 300070, China 2Department of Pathology, Cancer Institute and Hospital of Tianjin Medical University, Tianjin 300060, China Grant support: National Natural Science Foundation of China (30830049) and International Cooperation of China and Sweden (09ZCZDSF04400) "CTUSBDU 0CKFDUJWF� To investigate the influence of TGF-ȕ and IFN-Ȗ on the proliferation, migration and invasion of melanoma cells. .FUIPET� Melanoma cells were cultured in vitro. When tumor cells were confluent about 80% degree, cytokines were added into cell culture media. The concentration of TGF-ȕ and IFN-Ȗ was 5ng/mL and 10ng/mL, respectively. Melanoma cells were divided into free-cytokine group, TGF-ȕ group, IFN-Ȗ group, TGF-ȕ and IFN-Ȗ group. Tumor cells in each group were then incubated for 8h, 16h, 24h, 32h, 40h and 48h, respectively. After incubation, fixing and staining with SRB, the optical densities and percentage viability were then determined by absorption at 540 nm (A 540). The scarification of tumor cells in each group on the surface was created by a 200ȝL pipette tube. The motility of tumor cells in each group was assessed by measuring the distance between scarifications. The speed of the scuffing closure was monitored after 12h. The invasive ability of melanoma cells was observed by transwell cultivation. The tumor cells that invaded through the Matrigel and adhered to the bottom of the outside membrane were determined by absorption at 595 nm (A595). Gelatin zymography assay was used to examine the levels of matrix metalloproteinases-2 (MMP-2) and matrix metalloproteinases-9 (MMP-9) activity when the tumor cells were treated with cytokines after 24h. MMP-2 and MMP-9 activity was demonstrated by gradation in the sodium dodecyl sulfate polyacryl- amide gel electrophoresis (SDS-PAGE) gelatin. MMP-2 and MMP-9 activity was determined by Image analy- ७ ᱘᪳䄪䷄ः఩ტ㜖♢⻽႒ദ䛽喋㑂ण喝30830049喌স͙⦊఩䭱ऴ҈䶥Ⱊദ䛽䉰ߕ喋㑂ण喝09ZCZDSF04400喌 Ŗ ๕≑ࡧ⻽๓႒౔䄧ⵀ⾢⩋ ŗ ๕≑ࡧ⻽๓႒䭰ᆊ㗫⭐ࡧ䮎 䕆䃛҈㔱喝ႅ ԉႄ sunbaocun@yahoo.com.cn kk134 2010Ꭰじ37ࢣじ3᱋ ►⫳স►⫳ఌၼ̺㗫⭐⮰ࣽ⩋ȟࣽᆁڣᰵჲܳ ⮰ڟ㈧喾�嗀Ƞᬕ౔����Ꭰ7JSDIPXጞ㏻᣼ܦ㗫⭐छ㘩 䊣⎼κᚎᕓ►⫳喏ຮ϶►⫳স㗫⭐ڟ㖀ᕓ⮰ⵀ⾢Ϲ ♢ःݜڟ∔喾�嗀Ƞ䄪䷄㏰ݹ᱋጑҈ᐦ⿷β㢣⭐ߔ➕⮰ ᠭ㐙►⫳Ὅಷ喾�嗀Ƞ㏻ょ䔵ࣽ⣜►⫳▢ܲ⇸⮰►⫳ఌ ၼ᎞វ㉌�¥喋*'/�¥喌স㗫⭐㏲㘊ܲ⇸⮰䒘ࡂ⩋䪫 ఌၼ�¤喋5('�¤喌౔䄑►⫳�㗫⭐Ὅಷ͙ڣᰵ䛹㺭 ᘻ͵喏᱘ⵀ⾢ݕ⩔ѿโ㏲㘊Ὅಷ䔇̬ₑ㻮ᄋ*'/�¥ স5('�¤ᄥ㗫⭐㏲㘊්⃂ȟ䓭⼧সӡ㷙⮰҈⩔喏㏿ ᳈㶔ᬺ *'/�¥ᄥ㗫⭐㏲㘊ڣᰵួݢ҈⩔喏㔸 5('�¤݅छ᠚ៃ*'/�¥㔸ҫ㗫⭐㏲㘊ᖎ්ู⃂ȟ 䓭⼧সӡ㷙㘩߇Ƞ � ᴤ᭭Ϣᮍ⊩ ��� ㏲㘊㈧ȟ㏲㘊ఌၼস㏲㘊ഥڧ᲍Т 叽㞞㉌⭐㏲㘊㈧#��'��喋๕≑⣛⎂ࡧ䮎㏲㘊ა ᑌ᪳⇧㔭ጴ亴䊌喏⎼㜖"5$$喌喏㏲㘊ഥڧκ⌧ߌ ���㗺➇㵬⌱喋'#4喌⮰31.*����ഥڧദ喋)ZDMPOF喌喏 ഥڧκ��čȟ�� $0�⮰ᕾ⎫ഥڧポ͙Ƞ5('�¤䉙 㜖 1FQUFDI ژत喏ҫ⩔≿Ꮢͦ �OH�N-喞*'/�¥䉙㜖 1FQUFDIژत喏ҫ⩔≿Ꮢͦ��OH�N-Ƞ ��� ㏲㘊්⃂ȟ䓭⼧ȟӡ㷙჊侸 ㏲㘊්⃂⢳⮰ܲ᲼䛳⩔⸦䚜㒃ͥᬺ᳿㞞∁ܲ ᲼喋43#∁喌喏ьЏ්⃂㏲㘊㜟���ᣑ㼒ⅳऴ喏䮻࢟Б �f���͖�N-⮰≿Ꮢᣑ⻹κ��ႀᲫ喋$PTUBS喌Ƞܲ㏰喝 ⏢ݮᄥ⚓㏰ȟ5('�¤ ㏰喋5喌ȟ*'/�¥ ㏰喋*喌স 5('�¤�*'/�¥㏰喋5�*喌喏र㏰ܲݗκ�I喏��I喏��I喏 ��I喏��Iস��Iᬢ䬠◥ߌڑ̵⅛΅䚤ద჆喏䮻ऺᄥ㏲ 㘊䔇㵸43#᳿㞞喏≃㙝᳿᫅κ���ON।ٵᏒ̷℀㞞 䄧ं।ٵᏒ喋"���喌Ƞ㏲㘊䓭⼧䕆䓳ܾ⬁჊侸䔇㵸 ܲ᲼喏㏲㘊ьЏᎢᣑ⻹κ�ႀᲫڱ喏ᒱ���Б̶ᣑ㼒 ⅳऴऺ喏⩔���®-।๠ᄥ㏲㘊㶔䲎䔇㵸ܾ⬁ะ⤲喏 ᰠᢎഥڧദᎢᠵ⚓ܲ㏰ߌڑ̹स⮰㢛➕Ƞܾ⬁ᢋ ѐᘴऴ⮰䕋Ꮢ࢟㶔ᬺβ㏲㘊䓭⼧㘩߇⮰ᔗᚎ喏κ ��Iऺᄥ㻮ᄋ◥ܾ⬁ͷ䬠⮰䌉⻧䔇㵸≷჆喏Б䄑◥ �I݉໷䌉⻧䔇㵸ᴳ۲ࡂऺ䔇㵸㐋䃍႒ܲ᲼Ƞӡ㷙 ჊侸䛳⩔5SBOTXFMM㈧㐋喋*OWJUSPHFO喌喏ႀᒰͦ�®NȠ 仂ٴऽᄥ5SBOXFMMڱႀ㛈࠱㷗.BSUSJHFMϦ጑ദᏁ㛈 喋#%喌喏ऽڱႀڱᣑ⻹≿Ꮢͦ�f���͖�N-⮰㏲㘊স ᬌ㵬⌱ഥڧദ喏Ꭲߌڑ̹स⮰㢛➕Ƞโႀڱ݅ߌڑ ॗ���'#4⮰ഥڧദ҈ͦ䊷ࡂ⎼Ƞഥڧ��Iऺ喏⩔ ᷵オᨒࣧႀڱ㛈̶㏲㘊喏ᄥႀโ㛈̶㏲㘊䔇㵸㏿ᮢ ㉗᳿㞞喏���΅䚤≃㙝ऺ䄧ं���ON।ٵᏒ喋"���喌Ƞ ��� ᬺ㘢䚢䅝ܲ᲼ ܲݗ⩔̹स㢛➕ะ⤲㏲㘊��I喏ᩢ 䯲ഥڧദ⩔κ ᬺ㘢䚢䅝ܲ᲼ȠБॗ����ᬺ㘢⮰���㖆ͅ☛䚜㘦܉ 㘢䔇㵸4%4�1"(&喏⩡∟㜟⏠䚆ڜ䔇ڑ䭟Ჭ㐿۞⋞ͦ ₎喏ܲ ⻧܉㘢κ����5SJUPO�9���≃㘢�⁍喏⃻⁍��NJOȠ ᄲ㘢Ⴘڔ⊤ڑᬺ㘢䚢Ⴁ㗞⋞喋��NNPM�- Q)��� 5SJT�)$M喏 ��NNPM�- $B$M�喏���NPM�- /B$M喏�®NPM�- ;O$M�喌��č Ⴁ㗞��I喏፤㻰㔯侘᫛Ϛ㨉᳿㞞喏㙝㞞⋞㙝㞞㜟ู᳿ፒ ⌱ᮜȠ㏿᳈ݐ჆喝ప׻ܲ᲼㈧㐋≷჆܉㘢͙..1T⮰ ᲍ፒ▜Ꮢ喋࢟Џ㶔β..1T⮰≧ᕓⅠ᎟喌喏⩡∟܉㘢౔ ▜䭢Ὅᐻ̷ភ᣻喏Ꮐ⩔1IPSFUJY �%ܲ᲼䒛Т喏䃪჆࣮㔯 ը喏䄧ं᲍ፒ䲎⼛স䚢㼏᲍ፒ▜Ꮢ喏䚢ॗ䛻�᲍ፒ䲎⼛f 喋᲍ፒ▜Ꮢ�㗸ᮛ▜Ꮢ喌Ƞᄲᵣ᱘⮰䚢ॗ䛻ը̺ⰤᏀ⮰ ᄥ⚓㏰ֆ℀ը喏჆ͦₐᵣ᱘⮰䚢≧ᕓըȠ ��� 㐋䃍႒᫥∁ ڔ䘔᪜ᢚБ YiT㶔⹦喏ݕ⩔㐋䃍႒䒛Т4144 ����䔇㵸ࢁఌ㉌᫥ጚܲ᲼喋"/07"喌Ƞ � 㒧ᵰ ��� 5('�¤স*'/�¥ᄥ叽㞞㉌⭐⩋䪫স්⃂⮰ᒝ ৹҈⩔ 䕆䓳 43# ∁ 㻮ᄋ ��OH�N- *'/�¥ ȟ�OH�N- 5('�¤সθ㔱㖀⩔ᄥ#��㏲㘊ᒝ৹҈⩔⮰ᬢᩴڟ ㈧喏छ㻭5('�¤㏰㗫⭐㏲㘊ႄ≧⢳倄κᄥ⚓㏰喏ጚ ᐮڣᰵ㐋䃍႒ᘻ 喋͵U������喏1������喌喞㔸*'/�¥㏰ 㗫⭐㏲㘊ႄ≧⢳Ѻκᄥ⚓㏰喏ጚᐮڣᰵ㐋䃍႒ᘻ͵ 喋U������喏1������喌喞सᬢ㐅δ *'/�¥স5('�¤ะ ⤲ᬢ喏㗫⭐㏲㘊ႄ≧⢳̺݅ᄥ⚓㏰ᣑ䓽喏ጚᐮᬌ㐋 䃍႒ᘻ 喋͵U������喏1������喌Ƞ㐅δ̹स㏲㘊ఌၼะ ⤲ᬢ喏䮻㏲㘊ႄ≧⢳⮰ऄࡂ喏㗫⭐㏲㘊㶔⣜̹ͦस ⮰ᒎᔭ喝㐅δ5('�¤ะ⤲ᬢ喏㏲㘊ᒎᔭ႒̶ͧ㺭㶔 ⣜ͦᷙᒎ喞*'/�¥㏰݅ܦ⣜㏲㘊叻䭰߇⮰̷䭹喏㏲ 㘊ᒎᔭ႒̶㶔⣜ͦᗘ⊚⤯ᒎᝂๆ䓥ᒎ喏๝ࣧ₏፤ #��㏲㘊ᒎᔭ႒➥◥喞सᬢ㐅δ*'/�¥স5('�¤ ᬢ喏㏲㘊ᖎู#��㏲㘊ᒎᔭ႒➥◥Ƞ㻭ప�喏�Ƞ sis Software. 3FTVMUT� TGF-ȕ promoted the proliferation, migration and invasion of melanoma cells (P<0.05). However, IFN-Ȗ inhibited the proliferation, migration and invasion of melanoma cells (P<0.05). The effect weakened or disappeared when both of them were used (P>0.05). $PODMVTJPO� In vitro, TGF-ȕ may affect the inhibitory effect of IFN-Ȗ on the proliferation, migration and invasion of melanoma cells. 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