Lipopolysaccharide- and lipoteichoic acid-mediated pro-inflammatory cytokine production and modulation of tlr2, tlr4 and myd88 expression in human endometrial cells

Toll-like receptor [TLR]-mediated inflammatory processes are supposed to be involved in pathophysiology of spontaneous abortion and preterm labor. Here, we investigated functional responses of human endometrial stromal cells [ESCs] and whole endometrial cells [WECs] to lipopolysaccharide [LPS] and lipoteichoic acid [LTA] Endometrial tissues were obtained from 15 cycling women who underwent laparoscopic tubal ligation. Modulation of TLR2, TLR4 and MyD88 expression and production of pro-inflammatory cytokines by WECs and ESCs in response to LPS and LTA were assessed. WECs and ESCs expressed significant levels of TLR4 and MyD88 transcripts but, unlike WECs, ESCs failed to express TLR2 gene. Regardless of positive results of Western blotting, ESCs did not express TLR4 at their surface as judged by flow cytometry. Immunofluorescent staining revealed intracellular localization of TLR4 with predominant perinuclear pattern. LPS stimulation marginally increased TLR4 gene expression in both cell types, whereas such treatment significantly upregulated MyD88 gene expression after 8 hr [p<0.05]. At the protein level, however, LPS activation significantly increased TLR4 expression by ESCs [p<0.05]. LTA stimulation of WECs was accompanied with non-significant increase of TLR2 and MyD88 transcripts. LPS and LTA stimulation of WECs caused significant production of IL-6 and IL-8 in a dose-dependent manner [p<0.05]. Similarly, ESCs produced significant amounts of IL-6, IL-8 and also TNF-alpha in response to LPS activation [p<0.05]. Our results provided further evidence of initiation of inflammatory processes following endometrial TLR activation by bacterial components which could potentially be harmful to developing fetus

Conjugation of monoclonal antibodies to super paramagnetic iron oxide nanoparticles for detection of her2/neu antigen on breast cancer cell lines

Conjugation of monoclonal antibodies to super paramagnetic nanoparticles is an effective method for cancer diagnosis and treatment. In this study the humanized anti her2/neu monoclonal antibody- Herceptin- was conjugated to super paramagnetic iron oxide [SPIO] nanoparticles using EDC method. The concentration of the conjugated antibodies was measured by Bradford assay. The antibody-nanoparticle conjugates were incubated with SKBR-3 and T47D human breast carcinoma cell lines and the presence of the conjugates on cell surface was confirmed by Prussian blue iron staining method. Conjugation of Herceptin to SPIO resulted in a precipitate-free conjugate containing 20microg antibody/mg SPIO. Prussian blue iron-staining of cells showed successful binding of the conjugates to the cell surfaces. Conjugation of monoclonal antibodies to SPIO may be a useful method for detection of tumor cells, especially by MRI techniques