Placental Expression of CD100, CD72 and CD45 Is Dysregulated in Human Miscarriage

Abstract

Context and ObjectiveThe etiology of miscarriage is often multifactorial. One major cause, immunological rejection of the fetus, has not been clearly elucidated. Our aim was to establish whether the semaphorin CD100, its natural receptor CD72, and the glycoprotein CD45, implicated in immune mechanisms, are involved in pregnancy loss by examining their placental expression with real-time PCR, immunohistochemistry and western blotting techniques.PatientsPlacenta tissue from 72 Caucasian women undergoing surgical uterine evacuation due to early spontaneous pregnancy loss between the 8th and 12th week of gestation was divided into four groups based on miscarriage number. Gestational age-matched placentas from 18 healthy women without a history of miscarriage undergoing voluntary pregnancy termination were the control group. Placenta from 6 Caesarean deliveries performed at 38–40 weeks of gestation was also studied.ResultsCD100, CD72 and CD45 were expressed in placenta and exhibited different mRNA and protein levels in normal pregnancy and miscarriage. In particular, protein levels were highly dysregulated around 10 weeks of gestation in first and second miscarriage placentas. The CD100 soluble form was produced and immediately shed from placental tissue in all samples.ConclusionsFetal CD100, CD72 and CD45 seem to play a role in miscarriage. The present data support the involvement of the fetal immune system in pregnancy maintenance as well as failure.

HtrA1 in human urothelial bladder cancer: A secreted protein and a potential novel biomarker

“…RNA extraction, reverse transcription and real‐time PCR were performed as previously described . The sequences of the Q‐PCR primers targeting the HtrA1 gene are reported in Supporting Information Table .…”

“…Lysates were obtained as previously described . Protein concentrations of the extracts were assessed with the Bradford protein assay (Bio‐Rad Laboratories, Milan, Italy) …”

“…RNA extraction, reverse transcription and real-time PCR were performed as previously described. 23 The sequences of the Q-PCR primers targeting the HtrA1 gene are reported in Supporting Information Table 2. The reference genes SDHA (Succinate Dehydrogenase Complex Subunit A) and TBP (TATA Binding Protein) were used as the housekeeping genes for data normalization, to correct for variations in RNA quality and quantity.…”

“…Lysates were obtained as previously described. 23 Protein concentrations of the extracts were assessed with the Bradford protein assay (Bio-Rad Laboratories, Milan, Italy). 25 Samples were immunoprecipitated for analysis of the expression of the 50 kDa HtrA1 form, whereas they were directly processed by western blotting for the quantitative determination of the autocatalytic form.…”

Human Intestinal Epithelial Cells Express Interleukin-10 through Toll-Like Receptor 4-Mediated Epithelial-Macrophage Crosstalk

“…The amplification was done by 30 sec denaturation at 98°C and 8 sec annealing at 60.5°C for 35 cycles. Primers and amplification protocols for CD45 and β-actin have been described previously [25, 26]; CD45 sense 5’-CTGACATCATCACCTAGCAG-3’ and antisense 5’-TGCTGTAGTCAATCCAGTGG-3’ (product size 257 bp) and β-actin sense 5’-CATCCTCACCCTGAAGTACC-3’ and antisense 5’-GCTCATTGTAGAAGGTGTGG-3’ (product size 92 bp). Other primers used were human TLR4 sense 5’-GCATTTAACTCACTCTCCAGTC-3’ and antisense 5’-CATCCTGGCTTGAGTAGATAAC-3’ (product size 511 bp) and human PPARg sense 5'-TCTCTCCGTAATGGAAGACC-3' and antisense 5'-GCATTATGAGACATCCCCAC-3' (product size 482 bp).…”