Regulation of hepatocyte growth factor by basal and stimulated macrophages in women with endometriosis.
BACKGROUND
The different macromolecules as secreted by macrophages in the pelvic environment are believed to enhance the growth of endometriosis. However, the possible mediator that stimulates macrophages for the production of different growth factors is not well described. Therefore, we investigated the possible production of hepatocyte growth factor (HGF) by the basal and lipopolysaccharide (LPS)-stimulated macrophages derived from women with or without endometriosis.


METHODS
Using primary culture and 4-well chamber slides, adherent macrophages immunoreactive to CD68 were isolated from the peritoneal fluid (PF) of 20 infertile women with endometriosis and 12 women without endometriosis. The proliferation of basal and LPS-treated macrophages was investigated by the dimethylthiazole tetrazolioum bromide (MTT) assay. The production of HGF in the culture media of basal and LPS-stimulated macrophages was examined by enzyme-linked immunosorbent assay. The expression of mRNA for HGF and its receptor, c-Met, in the macrophages was investigated by RT-PCR. The effect of HGF on the growth of endometrial cells and macrophages was analysed by bromodeoxyuridine (BrdU) incorporation.


RESULTS
A >100% increase in the proliferation of peritoneal macrophages derived from women with endometriosis, and particularly of those harbouring dominant red lesions, was observed after treatment with LPS (P<0.05). A 4- and 3-fold increase in the production of HGF was observed by the LPS-treated macrophages derived from women with stage I-II endometriosis and stage III-IV endometriosis, respectively, when compared with non-LPS-treated macrophages (P<0.001). At the transcriptional level, we found a 5-fold increase in HGF mRNA expression in LPS-treated macrophages versus basal macrophages in women with endometriosis (P<0.001). The BrdU incorporation study indicates that 10-100 ng/ml of HGF enhanced the growth of endometrial epithelial cells, stroma and macrophages (approximately 50% increase) derived from women with endometriosis (all P<0.05).


CONCLUSIONS
LPS could be an inflammatory mediator of macrophage stimulation in the pelvic microenvironment. Besides mesenchymal cells, HGF is also produced by peritoneal macrophages and is possibly involved in the growth of endometriosis.
Khan
Khaleque Newaz
KN
Department of Obstetrics and Gynecology, Division of Cytokine Signaling, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8501, Japan. nemokhan@net.nagasaki-u.ac.jp
Masuzaki
Hideaki
H
Fujishita
Akira
A
Kitajima
Michio
M
Kohno
Tomoko
T
Sekine
Ichiro
I
Matsuyama
Toshifumi
T
Ishimaru
Tadayuki
T
D000328:Adult; D015703:Antigens, CD / Q000378:metabolism; D015214:Antigens, Differentiation, Myelomonocytic / Q000378:metabolism; D016022:Case-Control Studies; D049109:Cell Proliferation / Q000187:drug effects; D004715:Endometriosis / Q000235:genetics / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D017228:Hepatocyte Growth Factor / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D066298:In Vitro Techniques; D008070:Lipopolysaccharides / Q000494:pharmacology; D008262:Macrophage Activation / Q000187:drug effects; D017737:Macrophages, Peritoneal / Q000166:cytology / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D019859:Proto-Oncogene Proteins c-met / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D000097382:CD68 Molecule
D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D017228:Hepatocyte Growth Factor; D008070:Lipopolysaccharides; D019859:Proto-Oncogene Proteins c-met; D012333:RNA, Messenger; C067980:CD68 antigen, human; D000097382:CD68 Molecule