Diabetic Impairment of C-Kit+ Bone Marrow Stem Cells Involves the Disorders of Inflammatory Factors, Cell Adhesion and Extracellular Matrix Molecules

Bone marrow stem cells from diabetes mellitus patients exhibit functional impairment, but the relative molecular mechanisms responsible for this impairment are poorly understood. We investigated the mechanisms responsible for diabetes-related functional impairment of bone marrow stem cells by extensively screening the expression levels of inflammatory factors, cell cycle regulating molecules, extracellular matrix molecules and adhesion molecules. Bone marrow cells were collected from type 2 diabetic (db/db) and healthy control (db/m+) mice, and c-kit+ stem cells were purified (purity>85%) for experiments. Compared with the healthy control mice, diabetic mice had significantly fewer c-kit+ stem cells, and these cells had a lower potency of endothelial differentiation; however, the production of the angiogenic growth factor VEGF did not differ between groups. A pathway-focused array showed that the c-kit+ stem cells from diabetic mice had up-regulated expression levels of many inflammatory factors, including Tlr4, Cxcl9, Il9, Tgfb1, Il4, and Tnfsf5, but no obvious change in the expression levels of cell cycle molecules. Interestingly, diabetes-related alterations of the extracellular matrix and adhesion molecules were varied; Pecam, Mmp10, Lamc1, Itgb7, Mmp9, and Timp4 were up-regulated, but Col11a1, Fn1, Admts2, and Itgav were down-regulated. Some of these changes were also confirmed at the protein level by flow cytometry analysis. In conclusion, c-kit+ bone marrow stem cells from diabetic mice exhibited an extensive enhancement of inflammatory factors and disorders of the extracellular matrix and adhesion molecules. Further intervention studies are required to determine the precise role of each molecule in the diabetes-related functional impairment of c-kit+ bone marrow stem cells

A Computational Model of the Ionic Currents, Ca2+ Dynamics and Action Potentials Underlying Contraction of Isolated Uterine Smooth Muscle

Uterine contractions during labor are discretely regulated by rhythmic action potentials (AP) of varying duration and form that serve to determine calcium-dependent force production. We have employed a computational biology approach to develop a fuller understanding of the complexity of excitation-contraction (E-C) coupling of uterine smooth muscle cells (USMC). Our overall aim is to establish a mathematical platform of sufficient biophysical detail to quantitatively describe known uterine E-C coupling parameters and thereby inform future empirical investigations of physiological and pathophysiological mechanisms governing normal and dysfunctional labors. From published and unpublished data we construct mathematical models for fourteen ionic currents of USMCs: currents (L- and T-type), current, an hyperpolarization-activated current, three voltage-gated currents, two -activated current, -activated current, non-specific cation current, - exchanger, - pump and background current. The magnitudes and kinetics of each current system in a spindle shaped single cell with a specified surface area∶volume ratio is described by differential equations, in terms of maximal conductances, electrochemical gradient, voltage-dependent activation/inactivation gating variables and temporal changes in intracellular computed from known fluxes. These quantifications are validated by the reconstruction of the individual experimental ionic currents obtained under voltage-clamp. Phasic contraction is modeled in relation to the time constant of changing . This integrated model is validated by its reconstruction of the different USMC AP configurations (spikes, plateau and bursts of spikes), the change from bursting to plateau type AP produced by estradiol and of simultaneous experimental recordings of spontaneous AP, and phasic force. In summary, our advanced mathematical model provides a powerful tool to investigate the physiological ionic mechanisms underlying the genesis of uterine electrical E-C coupling of labor and parturition. This will furnish the evolution of descriptive and predictive quantitative models of myometrial electrogenesis at the whole cell and tissue levels

Transcription analysis of the myometrium of labouring and non-labouring women

An incomplete understanding of the molecular mechanisms that initiate normal human labour at term seriously hampers the development of effective ways to predict, prevent and treat disorders such as preterm labour. Appropriate analysis of large microarray experiments that compare gene expression in non-labouring and labouring gestational tissues is necessary to help bridge these gaps in our knowledge. In this work, gene expression in 48 (22 labouring, 26 non-labouring) lower-segment myometrial samples collected at Caesarean section were analysed using Illumina HT-12 v4.0 BeadChips. Normalised data were compared between labouring and non-labouring groups using traditional statistical methods and a novel network graph approach. We sought technical validation with quantitative real-time PCR, and biological replication through inverse variance-weighted meta-analysis with published microarray data. We have extended the list of genes suggested to be associated with labour: Compared to non-labouring samples, labouring samples showed apparent higher expression at 960 probes (949 genes) and apparent lower expression at 801 probes (789 genes) (absolute fold change ≥1.2, rank product percentage of false positive value (RP-PFP) <0.05). Although half of the women in the labouring group had received pharmaceutical treatment to induce or augment labour, sensitivity analysis suggested that this did not confound our results. In agreement with previous studies, functional analysis suggested that labour was characterised by an increase in the expression of inflammatory genes and network analysis suggested a strong neutrophil signature. Our analysis also suggested that labour is characterised by a decrease in the expression of muscle-specific processes, which has not been explicitly discussed previously. We validated these findings through the first formal meta-analysis of raw data from previous experiments and we hypothesise that this represents a change in the composition of myometrial tissue at labour. Further work will be necessary to reveal whether these results are solely due to leukocyte infiltration into the myometrium as a mechanism initiating labour, or in addition whether they also represent gene changes in the myocytes themselves. We have made all our data available at www.ebi.ac.uk/arrayexpress/ (accession number E-MTAB-3136) to facilitate progression of this work