Transcription profiling of PBMCS - PIF modulated gene expression

ABSTRACT: Pregnancy is an immune paradox: where the mother accepts a semi-allograft/allograft (donor embryo), while avoiding maternal immune suppression. Indeed, impaired development of maternal immune tolerance towards the conceptus may be an important cause of implantation failure and pregnancy loss. Successful implantation rates for allogenic donor embryos are unexpectedly high, while abnormal embryos may be unable to initiate maternal tolerance, suggesting that embryo-specific compounds can play a vital role in this process. Both a tolerant T-suppressor (TH2), and inflammatory, T-helper 1 (TH1) cytokine profiles are an integral part of pregnancy and the viable embryo/fetus plays a critical role in creating this delicate balance. Maternal recognition of pregnancy occurs prior to implantation. There were several embryo derived factors reported. However, we previously found that preimplantation factor (PIF) is distinct from the other non-pregnancy-specific embryo-derived factors. Herein we aimed to characterize PIF and examine its role in early pregnancy events by observing the isolated novel peptide's effect on human Peripheral Blood Mononuclear Cell (PBMC). We did affymetrix, GeneChip Human Genome U133 plus 2.0 array on RNA isolated from normal human PBMC cultured for 24hrs in the presence or absence of PIF to study the genes modulated by PIF. Experiment Overall Design: Blood was collected from a male volunteer and PBMC were separated using the Ficoll Hypaque method. Cells were cultured in RPMI culture media for 24 hours in four different groups in duplicate viz., Media alone, Media + 50 nM PIF, Media + 50nM PIF +CD3 (10µg/ml)+ CD28 (10µg/ml) and Media + CD3 (10µg/ml) +CD28 (10µg/ml). At the end of incubation cells were removed from the media, washed and collected in RNAlater (Qiagen) and transferred to Biocodon Sciences (Tx, USA) for Gene array analysis on dry ice. Samples were analyzed using the Agilent Bioanalyzer Process found to be adequate. Each 100 µl contained 2.7-5.4 µg RNA (Nanodrop analyzer). Each sample was tested integrity. RNA from duplicate samples was combined and array was performed on an Affymetrix chip (Human Genome U 133 Plus 2.0 Array). Subsequently, arrays were processed, scanned, data was extracted and statistical evaluation was carried out comparing the various groups. Data generated were categorized as robust changes or mild changes based on differences in expression.

ORGANISM(S): Homo sapiens

SUBMITTER: Eytan Barnea 

PROVIDER: E-GEOD-18291 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress