Conditional ablation of macrophages disrupts ovarian vasculature

Macrophages are the most abundant immune cell within the ovary. Their dynamic distribution throughout the ovarian cycle and heterogenic array of functions suggest the involvement in various ovarian processes, but their functional role has yet to be fully established. The aim was to induce conditional macrophage ablation to elucidate the putative role of macrophages in maintaining the integrity of ovarian vasculature. Using the CD11b-diphtheria toxin receptor (DTR) mouse, in which expression of human DTR is under the control of the macrophage-specific promoter sequence CD11b, ovarian macrophages were specifically ablated in adult females by injections of diphtheria toxin (DT). CD11b-DTR mice were given DT treatment or vehicle and ovaries collected at 2, 8, 16, 24 and 48 h. Histochemical stains were employed to characterise morphological changes, immunohistochemistry for F4/80 to identify macrophages and the endothelial cell marker CD31 used to quantify vascular changes. In normal ovaries, macrophages were detected in corpora lutea and in the theca layer of healthy and atretic follicles. As macrophage ablation progressed, increasing amounts of ovarian haemorrhage were observed affecting both luteal and thecal tissue associated with significant endothelial cell depletion, increased erythrocyte accumulation and increased follicular atresia by 16 h. These events were followed by necrosis and profound structural damage. Changes were limited to the ovary, as DT treatment does not disrupt the vasculature of other tissues likely reflecting the unique cyclical nature of the ovarian vasculature and heterogeneity between macrophages within different tissues. These results show that macrophages play a critical role in maintaining ovarian vascular integrity

Human Resistin Is a Systemic Immune-Derived Proinflammatory Cytokine Targeting both Leukocytes and Adipocytes

The characteristics of human resistin (RETN) are unclear and controversial despite intensive adipose-focused research. Its transcriptional and functional similarity with the murine myeloid-specific and CCAAT/enhancer binding protein epsilon (Cebpe)-dependent gene, resistin-like gamma (Retnlg), is unexplored. We examined the human CEBPE-regulatory pathway by unbiased reference and custom gene expression assays. Real-time RT-PCR analysis demonstrated lack of both the transcriptional factor CEBPE and RETN expression in adipose and muscle cells. In contrast, primary myelocytic samples revealed a concerted CEBPE-RETN transcription that was significantly elevated in inflammatory synoviocytes relative to intact peripheral blood mononuclear cells (PBMC). Mouse Cebpe and Retnlg were predictably expressed in macrophages, whereas Retn was abundant in adipocytes. Quite the opposite, a low and inconsistent RETN transcription was seen in some human white adipose tissue (WAT) biopsies without any relationship to body mass index, insulin sensitivity, or fat depot. However, in these cases, RETN was co-detected with CEBPE and the leukocyte-specific marker, EMR1, indicating the presence of inflammatory cells and their possible resistin-mediated effect on adipocytes. Indeed, addition of human resistin to WAT in culture induced, like in PBMC, the inflammatory cytokines IL6, IL8 and TNF. Importantly, the expression of the adipose-specific markers CEBPA, FABP4 and SLC2A4 was unchanged, while the expected inhibitory effect was seen with TNF. Both cytokines increased the mRNA level of CCL2 and MMP3, which may further promote inflammation in WAT. Thus, the myeloid-restricted nature of CEBPE precludes the expression of RETN in human adipocytes which, however, are targeted by this innate immune-derived proinflammatory cytokine

Reconstruction and simulation of neocortical microcircuitry

We present a first-draft digital reconstruction of the microcircuitry of somatosensory cortex of juvenile rat. The reconstruction uses cellular and synaptic organizing principles to algorithmically reconstruct detailed anatomy and physiology from sparse experimental data. An objective anatomical method defines a neocortical volume of 0.29 ± 0.01 mm3 containing ∼31,000 neurons, and patch-clamp studies identify 55 layer-specific morphological and 207 morpho-electrical neuron subtypes. When digitally reconstructed neurons are positioned in the volume and synapse formation is restricted to biological bouton densities and numbers of synapses per connection, their overlapping arbors form ∼8 million connections with ∼37 million synapses. Simulations reproduce an array of in vitro and in vivo experiments without parameter tuning. Additionally, we find a spectrum of network states with a sharp transition from synchronous to asynchronous activity, modulated by physiological mechanisms. The spectrum of network states, dynamically reconfigured around this transition, supports diverse information processing strategies

Pathological pregnancies. Results of amniotic fluid studies and fetal outcome

Late amniocenteses (greater than 20 weeks' gestation) were performed in 114 pregnancies with no a priori genetic risk, but referred because of abnormal clinical and/or ultrasound findings suggesting fetal malformations. Reasons for referral included polyhydramnios (51 cases), oligohydramnios (15 cases), fetal growth retardation (FGR) (16 cases) and abnormal fetal ultrasound findings excluding anencephaly (32 cases). In 42 of these cases, referral was motivated by a combination of the above abnormal findings. When polyhydramnios was the sole anomaly (25 cases), 5 fetuses were malformed (20%), abnormal fetal karyotype and/or elevated amniotic fluid alphafetoprotein (AFP) were demonstrated in 2 cases. Oligohydramnios was the sole anomaly in one case; the infant died of prematurity. Fetal growth retardation was the sole anomaly in 14 cases, 11 otherwise normal newborns were small for date, 2 died at birth and 1 was malformed (1/14, 7%). In this group all fetal karyotypes were normal and in 2 cases amniotic fluid AFP were increased. In the 32 pregnancies without abnormal amniotic fluid volume and/or FGR and with fetal malformation(s) suggested by ultrasound, all malformations except one (ovarian cyst possibly ruptured during birth) were confirmed at birth, amniotic fluid AFP was elevated, and/or karyotype was abnormal in 6 cases. In 42 pregnancies where more than one alarm sign was present, abnormal karyotype and/or elevated amniotic fluid AFP level were recorded in 21 of the 39 cases where amniocentesis was performed, 33 fetuses were malformed (79%) and 13 died in the perinatal period (31%). The high incidence of abnormal results of amniocentesis found in this survey of pathological pregnancies, particularly in those with multiple alarm signs, emphasizes the need for amniocentesis in these situations.Journal ArticleResearch Support, Non-U.S. Gov'tFLWNAinfo:eu-repo/semantics/publishe

Acute molecular effects of pressure-controlled intermittent coronary sinus occlusion in patients with advanced heart failure

Aims Cardiac repair has steered clinical attention and remains an unmet need, because available regenerative therapies lack robust mechanistic evidence. Pressure-controlled intermittent coronary sinus occlusion (PICSO), known to induce angiogenetic and vasoactive molecules as well as to reduce regional ischemia, may activate endogenous regenerative processes in failing myocardium. We aimed to investigate the effects of PICSO in patients with advanced heart failure undergoing cardiac resynchronization therapy. Methods and results Eight out of 32 patients were treated with PICSO, and the remainder served as controls. After electrode testing including left ventricular leads, PICSO was performed for 20 min. To test immediate molecular responses, in both patient groups, coronary venous blood samples were taken at baseline and after 20 min, the time required for the intervention. Sera were tested for microRNAs and growth factors. To test the ability of up-regulated soluble factors on cell proliferation and expression of transcription factors [e.g. Kruppel-like factor 4 (KLF-4)], sera were co-cultured with human cardiomyocytes and fibroblasts. As compared with controls, significant differential expression (differences between pre-values and post-values in relation to both patient cohorts) of microRNA patterns associated with cardiac development was observed with PICSO. Importantly, miR-143 (P < 0.048) and miR-145 (P < 0,047) increased, both targeting a network of transcription factors (including KLF-4) that promote differentiation and repress proliferation of vascular smooth muscle cells. Additionally, an increase of miR-19b (P < 0.019) known to alleviate endothelial cell apoptosis was found, whereas disadvantageous miR-320b (P < 0.023) suspect to impair expression of c-myc, normally provoking cell cycle re-entry in post-mitotic myocytes and miR-25 (P < 0.023), decreased, a target of anti-miR application to improve contractility in the failing heart. Co-cultured post-PICSO sera significantly increased cellular proliferation both in fibroblasts (P < 0.001) and adult cardiomycytes (P < 0.004) sampled from a transplant recipient as compared with controls. Adult cardiomyocytes showed a seven-fold increase of the transcription factor KLF-4 protein when co-cultured with treated sera as compared with controls. Conclusions Here, we show for the first time that PICSO, a trans-coronary sinus catheter intervention, is associated with an increase in morphogens secreted into cardiac veins, normally present during cardiac development, and a significant induction of cell proliferation. Present findings support the notion that epigenetic modifications, that is, haemodynamic stimuli on venous vascular cells, may reverse myocardial deterioration. Further investigations are needed to decipher the maze of complex interacting molecular pathways in failing myocardium and the potential role of PICSO to reinitiate developmental processes to prevent further myocardial decay eventually reaching clinical significance