Tumorski makrofagi u regulaciji angiogeneze invazivnog duktalnog raka dojke : doktorska disertacija

Ciljevi istraživanja: ciljevi ovog istraživanja bili su odrediti gustoću tumorskih makrofaga (TAM) kao i ispoljenost vaskularnog endotelnog čimbenika rasta (VEGF) i bazičnog fibroblastnog čimbenika rasta (bFGF) u tumorskom parenhimu duktalnog invazivnog karcinoma dojke NOS te usporediti ove parametre međusobno te s maksimalnom gustoćom malih krvnih žila (MVD). Nadalje odredila se je ispoljenost monocitne kemotaktične bjelančevine-I (MCP-1) u tumorskim stanicama ove vrste raka dojke te se usporedila s infiltracijom makrofaga i drugim spomenutim parametrima. Konačno, sposobnost tumora da stvori spomenuti kemokin ispitana je i određivanjem razine glasničke ribonukleinske kiseline (gRNK) za MCP-1. Materijali i metode: 97 duktalnih invazivnih karcinoma dojke NOS imunohistokemijski je obrađeno koristeći "avidin-biotin" metodu kako bi se odredila gustoća TAM, MVD te ispoljenost MCP-1, VEGF i bFGF (analizirano 63 karcinoma dojke) u tumorskim stanicama. gRNK za spomenuti kemokin određena je reakcijom lančane polimeraze kojoj prethodi obrnuta transkripcija (RT-PCR) u 8 uzoraka duktalnog invazivnog raka dojke te kontrolnog normalnog tkiva dojke. Statistička analiza učinjena je na osobnom računalu koristeći hi-kvadrat test, t-test i jednosmjernu analizu varijance. Rezultllti: statistička analiza utvrdila je da karcinomi dojke s pojedinačnim makrofagima u svojoj stromi imaju značajno nižu MVD u usporedbi s onima koji su imali nakupine ili difuzne infiltrate makrofaga u stromi (p=0,005).Također, značajna pozitivna korelacija uočena je između ispoljenosti VEGF u tumorskom parenhimu te MVD (p<0,001), infiltracije makrofaga (p=0,003) i ispoljenosti bFGF (p=0,016). Povezanost između MCP-1 i infiltracije makrofaga nije utvrđena, a rezultati reakcije lančane polimeraze utvrdili su da duktalni invazivni rak dojke, kao i normalno tkivo dojke, stvaraju glasničku ribonukleinsku kiselinu za ovaj kemokin. Zaključci: u duktalnom invazivnom raku dojke NOS- makrofazi ostvaruju angiogenezni učinak, baš kao i VEGF stvoren od strane tumorskog parenhima. Ovaj angiogenezni čimbenik, također, pokazuje kemotaktični učinak na makrofage. bFGF samostalno ne ostvaruje angiogeni učinak u ovoj vrsti raka dojke, a njegova ekspresija mogla bi biti ostvarena istimregulacijskim mehanizmima kao i ekspresija VEGF.Objectives: the aims of this investigation were to determine infiltrations of Tumor Associated Machrophages (TAM) as well as expresion of Vascular Endothelial Growth Factor (VEGF) and basic Fibroblast Growth Factor (bFGF) in the tumr parenchymal of ductal invasive breast carcinomas NOS and correlate these parameters to each other and wth maximal microvessel density (MVD). Also, the expresion of Monocite Chemotactic Protein - 1 (MCP - 1) in the tumr cells of this kind of breast cancer was determined and compared with macrophage infiltrations and above - mentioned parameters. Finally, the tumor´s capability to produce mentioned chemokine was, also, analysed by determination the level of messenger - ribonucleic acid (mRNA) for MCP - 1. Material and methods: 97 ductal invasive breast carcinomas NOS were analysed by immunohistochemistry using "avidin - biotin" method and macrophage infiltration, MVD, expression of MCP - 1, VEGF and FGF (63 samples of breast carcinoma were analysed) in the tumor cells were determined. The expression of messenger - ribonucleic acid for MCP - 1 was examined by reverse transcriptase - polymerase chain reaction (RT - PCR) in the 8 samples of breast carcinomas and control normal breast tissues. Statistical analysis was performed by the personal computer using chi square test, t - test and "one way" analysis of variance. Results: statistical analysis pointed that breast carcinomas with only indivudual stromal macrophages had significantly decreased MVD in the comparation with focally and diffusely infiltrated tumros (p = 0, 005). Also, postive corelations between parenchymal expression of VEGF and MVD (p < 0, 001), macrophage infiltrations (p = 0, 003) and expression of bFGF (p = 0, 016) were found. The corelation between expession of MCP - 1 and macrophages infiltrations was not found and the results of the polymerase chain reaction pointed that breast carcinomas, as well as normal breast tissue can expressed messenger ribonucleic acid for MCP - 1. Conclusion: in the ductal invasive breast carcinomas NOS, TAM promote angiogenesis as well as VEGF produced by tumor parenchymal. Also, this angiogenic factor has chemotactic influence on macrophages. bFGF has no angiogenic properties in this kind of breast cancer and the expression of this angiogenic factor and VEGF can be achieved by the same regulatory mechanisms