Eva Sweeney

Mesenchymal stem cell inhibition of T-helper 17 cell- differentiation is triggered by cell–cell contact and mediated by prostaglandin E2 via the EP4 receptor

Mesenchymal stem cells (MSCs) inhibit T‐cell activation and proliferation but their effects on individual T‐cell‐effector pathways and on memory versus naïve T cells remain unclear. MSC influence on the differentiation of naïve and memory CD4+ T cells toward the Th17 phenotype was examined. CD4+ T cells exposed to Th17‐skewing conditions exhibited reduced CD25 and IL‐17A expression following MSC co‐culture. Inhibition of IL‐17A production persisted upon re‐stimulation in the absence of MSCs. These effects were attenuated when cell–cell contact was prevented. Th17 cultures from highly purified naïve‐ and memory‐phenotype responders were similarly inhibited. Th17 inhibition by MSCs was reversed by indomethacin and a selective COX‐2 inhibitor. Media from MSC/Th17 co‐cultures contained increased prostaglandin E2 (PGE2) levels and potently suppressed Th17 differentiation in fresh cultures. MSC‐mediated Th17 inhibition was reversed by a selective EP4 antagonist and was mimicked by synthetic PGE2 and a selective EP4 agonist. Activation‐induced IL‐17A secretion by naturally occurring, effector‐memory Th17 cells from a urinary obstruction model was also inhibited by MSC co‐culture in a COX‐dependent manner. Overall, MSCs potently inhibit Th17 differentiation from naïve and memory T‐cell precursors and inhibit naturally‐occurring Th17 cells derived from a site of inflammation. Suppression entails cell‐contact‐dependent COX‐2 induction resulting in direct Th17 inhibition by PGE2 via EP4.

Engraftment, migration and differentiation of neural stem cells in the rat spinal cord following contusion injury

BACKGROUND AIMS Spinal cord injury is a devastating injury that impacts drastically on the victims quality of life. Stem cells have been proposed as a therapeutic strategy. Neural stem (NS) cells have been harvested from embryonic mouse forebrain and cultured as adherent cells. These NS cells express markers of neurogenic radial glia. METHODS Mouse NS cells expressing green fluorescent protein (GFP) were transplanted into immunosupressed rat spinal cords following moderate contusion injury at T9. Animals were left for 2 and 6 weeks then spinal cords were fixed, cryosectioned and analyzed. Stereologic methods were used to estimate the volume and cellular environment of the lesions. Engraftment, migration and differentiation of NS cells were also examined. RESULTS NS cells integrated well into host tissue and appeared to migrate toward the lesion site. They expressed markers of neurons, astrocytes and oligodendrocytes at 2 weeks post-transplantation and markers of neurons and astrocytes at the 6-week time-point. NS cells appeared to have a similar morphologic phenotype to radial glia, in particular at the pial surface. CONCLUSIONS Although no functional recovery was observed using the Basso Beattie Bresnahan (BBB) locomotor rating scale, NS cells are a potential cellular therapy for treatment of injured spinal cord. They may be used as delivery vehicles for therapeutic proteins because they show an ability to migrate toward the site of a lesion. They may also be used to replace lost or damaged neurons and oligodendrocytes.

Stereology of human myometrium in pregnancy: influence of maternal body mass index and age

OBJECTIVE Knowledge of the stereology of human myometrium in pregnancy is limited. Uterine contractile performance may be altered in association with maternal obesity and advanced maternal age. The aim of this study was to investigate the stereology of human myometrium in pregnancy, and to evaluate a potential influence of maternal body mass index (BMI) and age. STUDY DESIGN Biopsies of human myometrium were obtained from 57 women at cesarean section (n = 26, n = 13, n = 18 normal, overweight and obese BMI, respectively), and volume fractions of smooth muscle and extracellular matrix were assessed using stereologic techniques. RESULTS The smooth muscle constituted 65.2% ± 8.9% (standard deviation) and the extracellular matrix 32.6% ± 7.7% (standard deviation) (n = 57). There was no correlation observed between maternal BMI, age, or parity with the fractional volumes of either smooth muscle or extracellular matrix. CONCLUSION These results outline the stereology of human myometrium in pregnancy. Putative functional differences in contractility, pertaining to obese or older mothers, are not related to smooth muscle content.

The influence of smooth muscle content and orientation in dissected human pregnant myometrial strips on contractility measurements.

This study examined the hypothesis that the force generated by myometrial strips from pregnant women is influenced by the smooth muscle content and fibre orientation of the strips and that correcting for these structural variables will provide a more accurate measure of contractility. Myometrial strips (n=72) were contracted by exposure to KCl, oxytocin, U44619 and phenylephrine and maximum responses were recorded. Morphological techniques were used to determine the cross-sectional area of the strips, the area occupied by smooth muscle and the area occupied by smooth muscle longitudinal in the strip. Maximum responses to contractile agents were expressed in terms of these three variables. The mean cross sectional area of strips was 2.01 ± 0.06 mm(2), of which 50% was smooth muscle, and 18% was smooth muscle longitudinal in the strip (n=72). There was much heterogeneity in responses, smooth muscle content and fibre orientation. Correction for morphological variability did not improve the heterogeneity in responses where coefficients of variation among strips from the same donor ranged from 43% to 63% when expressed in relation to longitudinal smooth muscle cross-sectional area. The standard method of preparation of myometrial strips for in vitro recording results in samples that are not structurally uniform. Correcting for the known structural variables does not provide a more accurate measure of maximum contractile responses. Because of the heterogeneity shown here, experiments that are dependent upon accurate estimation of maximum contractile responses require a large number of replicates to reach meaningful conclusions.

The Effects of Extracellular Calcium-Sensing Receptor Ligands on the Contractility of Pregnant Human Myometrium In Vitro

Ligands for extracellular calcium-sensing (CaS) receptors inhibit oxytocin-induced contractions of the rat’s uterus. In this study, we investigated whether the CaS receptor ligands calindol, cinacalcet, and calhex 231 have similar effects on pregnant human myometrium. We compared their effects to those of the calcium-channel blocker nifedipine. In conventional concentration–effect experiments, both the mean contractile force (MCF) and the maximum amplitude of contractions induced by 1 nmol/L oxytocin were inhibited by nifedipine. Calindol and cinacalcet were ineffective as inhibitors, while calhex-231 produced partial inhibition. When single 10 μmol/L doses were applied calhex-231 produced a slowly developing inhibition, reducing the MCF to 38%, and amplitude to 34%, of vehicle controls after 1 hour. In similar experiments, calindol was ineffective while cinacalcet weakly inhibited only the amplitude. Immunohistochemistry revealed sparse expression of CaS receptors in pregnant human myometrium.