Combinatorial polymer electrospun matrices promote physiologically-relevant cardiomyogenic stem cell differentiation

Myocardial infarction results in extensive cardiomyocyte death which can lead to fatal arrhythmias or congestive heart failure. Delivery of stem cells to repopulate damaged cardiac tissue may be an attractive and innovative solution for repairing the damaged heart. Instructive polymer scaffolds with a wide range of properties have been used extensively to direct the differentiation of stem cells. In this study, we have optimized the chemical and mechanical properties of an electrospun polymer mesh for directed differentiation of embryonic stem cells (ESCs) towards a cardiomyogenic lineage. A combinatorial polymer library was prepared by copolymerizing three distinct subunits at varying molar ratios to tune the physicochemical properties of the resulting polymer: hydrophilic polyethylene glycol (PEG), hydrophobic poly(ε-caprolactone) (PCL), and negatively-charged, carboxylated PCL (CPCL). Murine ESCs were cultured on electrospun polymeric scaffolds and their differentiation to cardiomyocytes was assessed through measurements of viability, intracellular reactive oxygen species (ROS), α-myosin heavy chain expression (α-MHC), and intracellular Ca(2+) signaling dynamics. Interestingly, ESCs on the most compliant substrate, 4%PEG-86%PCL-10%CPCL, exhibited the highest α-MHC expression as well as the most mature Ca(2+) signaling dynamics. To investigate the role of scaffold modulus in ESC differentiation, the scaffold fiber density was reduced by altering the electrospinning parameters. The reduced modulus was found to enhance α-MHC gene expression, and promote maturation of myocyte Ca(2+) handling. These data indicate that ESC-derived cardiomyocyte differentiation and maturation can be promoted by tuning the mechanical and chemical properties of polymer scaffold via copolymerization and electrospinning techniques.     

Inflammatory Breast Cancer: A Distinct Clinicopathological Entity Transcending Histological Distinction

Introduction

Although well recognized in breast oncology literature, histologic subtypes have not been previously described in inflammatory breast cancer (IBC). The purpose of this study was to describe lobular subtype in IBC and assess the impact of histology on patient outcomes.

Methods

We performed a retrospective analysis of 659 IBC patients at MD Anderson Cancer Center between January 1984 and December 2009. Patients with Invasive Lobular, Mixed Invasive Ductal and Lobular, or Invasive Ducal Carcinomas (ILC, MIC, IDC, respectively) comprise the subject of this report. Patient characteristics and survival estimates were compared by using chi-square test and Kaplan-Meier method with log-rank statistic. Cox proportional hazards models were fit to determine association of histology with outcomes after adjustment for other characteristics.

Results

A total of 30, 37, and 592 patients were seen to have invasive lobular, mixed, or ductal histology, respectively. Grade 3 tumors were more common in the ductal group (78%) than in the lobular (60%) or mixed (61%) group (P = 0.01). The 3-year overall survival rates were 68%, 64%, and 62% in the lobular, mixed, and ductal groups, respectively (P = 0.68). After adjustment, histology did not have a significant effect on death in the lobular group (HR = 0.70, 95% confidence interval [CI]: 0.26–1.94; P = 0.50) or mixed group (HR = 0.53, 95% CI: 0.25–1.13; P = 0.10) compared with the ductal group.

Conclusion

In this cohort of IBC patients, lobular histology was seen in 4.5% cases. Histology does not appear to have a significant effect on survival outcomes in IBC patients, unlike in patients with non-inflammatory breast cancer (n-IBC), indicating the distinct biological behavior of the IBC phenotype.     

Ethanolic fermentation of cane molasses by a highly flocculent yeast

Summary A study of the comparative kinetics of standardS.uvarum ATCC 26602 withS.cerevisiae Y-10 (an isolate) and a highly flocculent strain ofS.uvarum in batch mode has shown that both the isolate and the highly flocculentS. uvarum strain have more desirable characteristics than the standard strains for ethanol production from cane molasses.     

Identification and Characterization of a Compound That Protects Cardiac Tissue from Human Ether-à-go-go-related Gene (hERG)-related Drug-induced Arrhythmias

The human Ether-à-go-go-related gene (hERG)-encoded K⁺ current, I_Kr is essential for cardiac repolarization but is also a source of cardiotoxicity because unintended hERG inhibition by diverse pharmaceuticals can cause arrhythmias and sudden cardiac death. We hypothesized that a small molecule that diminishes I_Kr block by a known hERG antagonist would constitute a first step toward preventing hERG-related arrhythmias and facilitating drug discovery. Using a high-throughput assay, we screened a library of compounds for agents that increase the IC₇₀ of dofetilide, a well characterized hERG blocker. One compound, VU0405601, with the desired activity was further characterized. In isolated, Langendorff-perfused rabbit hearts, optical mapping revealed that dofetilide-induced arrhythmias were reduced after pretreatment with VU0405601. Patch clamp analysis in stable hERG-HEK cells showed effects on current amplitude, inactivation, and deactivation. VU0405601 increased the IC₅₀ of dofetilide from 38.7 to 76.3 nm. VU0405601 mitigates the effects of hERG blockers from the extracellular aspect primarily by reducing inactivation, whereas most clinically relevant hERG inhibitors act at an inner pore site. Structure-activity relationships surrounding VU0405601 identified a 3-pyridiyl and a naphthyridine ring system as key structural components important for preventing hERG inhibition by multiple inhibitors. These findings indicate that small molecules can be designed to reduce the sensitivity of hERG to inhibitors.     

Effect of aqueous leaf extract of Azadirachta indica on the reproductive organs in male mice

Effect of oral administration (50, 100, and 200 mg/kg body weight/day, for 28 days) of aqucous leaf extract of neem (Azadirachta indica) on the male reproductive organs of the Parkes (P) strain mice was investigated. The treatment had no effect on body weight and the reproductive organs weight. In treated mice, testes showed both normal and affected seminiferous tubules in the same sections; the affected seminiferous tubules showed intraepithelial vacuolation, loosening of germinal epithelium, marginal condensation of chromatin in round spermatids, occurrence of giant cells, mixing of germ cell types in stages of spermatogenesis and degenerated appearance of germ cells. In severe cases, the tubules were lined with Sertoli cells only, Sertoli cells and rare germ cells, or with Sertoli cells and several germ cells but without cellular association patterns. Also, the frequency of affected seminiferous tubules in testes of the extract-treated mice was significantly higher than the controls, though this remained unaffected in mice treated at 50 mg/kg body weight of the extract. Doses at 50 or 100 mg/kg body weight of neem leaf extract did not cause appreciable alterations in histological appearance of the epididymis, while a dose of 200 mg/kg body weight caused marked alterations both in histological appearance and the level of sialic acid in the duct. The treatment also had adverse effects on motility, morphology, and number of spermatozoa in the cauda epididymidis, level of fructose in the seminal vesicle, and on litter size. After 42 days of withdrawal of the treatment, the alterations induced in the reproductive organs recovered to control levels. Our results suggested that treatment with neem leaf extract caused reversible alterations in the male reproductive organs of P mice.     

Lipocalin (LCN) 2 Mediates Pro-Atherosclerotic Processes and Is Elevated in Patients with Coronary Artery Disease

Background

Lipocalin (LCN) 2 is associated with multiple acute and chronic inflammatory diseases but the underlying molecular and cellular mechanisms remain unclear. Here, we investigated whether LCN2 is released from macrophages and contributes to pro-atherosclerotic processes and whether LCN2 plasma levels are associated with the severity of coronary artery disease progression in humans.

Methods and Results

In an autocrine-paracrine loop, tumor necrosis factor (TNF)-α promoted the release of LCN2 from murine bone-marrow derived macrophages (BMDM) and vice versa. Moreover, LCN2 stimulation of BMDM led to up-regulation of M1 macrophage markers. In addition, enhanced migration of monocytic J774A.1 cells towards LCN2 was observed. Furthermore, LCN2 increased the expression of the scavenger receptors Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) as well as scavenger receptor class A-1 (SRA-1) and induced the conversion of macrophages to foam cells. In atherosclerotic lesions of low density lipoprotein receptor-deficient (ldlr ^−/−) mice fed a high fat, high cholesterol diet, LCN2 was found to be co-localized with macrophages in the shoulder region of the atherosclerotic plaque. In addition, LCN2 plasma levels were significantly increased in plasma samples of these mice. Finally, LCN2 plasma levels correlated with the severity of coronary artery disease (CAD) in patients as determined by coronary angiography.

Conclusions

Here we demonstrated that LCN2 plays a pivotal role in processes involved in atherogenesis by promoting polarization and migration of monocytic cells and development of macrophages towards foam cells. Moreover, LCN2 may be used as a prognostic marker to determine the status of CAD progression.     

Enhanced heterodimerization of Bax by Bcl-2 mutants improves irradiated cell survival

B Cell Lymphoma-2 (Bcl-2) protein suppresses ionizing radiation-induced apoptosis in hemato-lymphoid system. To enhance the survival of irradiated cells, we have compared the effects and mechanism of Bcl-2 and its functional variants, D34A (caspase-3 resistant) and S70E (mimics phosphorylation on S70). Bcl-2 and its mutants were transfected into hematopoietic cell line and assessed for cell survival, clonogenicity and cell cycle perturbations upon exposure to ionizing radiation. The electrostatic potential of BH3 cleft of Bcl-2/mutants and their heterodimerization with Bcl-2 associated X protein (Bax) were computationally evaluated. Correspondingly, these results were verified by co-immunoprecipitation and western blotting. The mutants afford higher radioprotective effect than Bcl-2 in apoptotic and clonogenic assays at D₀ (radiation dose at which 37 % cell survival was observed). The computational and functional analysis indicates that mutants have higher propensity to neutralize Bax protein by heterodimerization and have increased caspase-9 suppression capability, which is responsible for enhanced survival. This study implies potential of Bcl-2 mutants or their chemical/peptide mimics to elicit radioprotective effect in cells exposed to radiation.     

Determination of tissue contributions to the circulating lipid pool in cold exposure via systematic assessment of lipid profiles

Plasma lipid levels are altered in chronic conditions such as type 2 diabetes and cardiovascular disease as well as during acute stresses such as fasting and cold exposure. Advances in MS-based lipidomics have uncovered a complex plasma lipidome of more than 500 lipids that serve functional roles, including as energy substrates and signaling molecules. This plasma lipid pool is maintained through regulation of tissue production, secretion, and uptake. A major challenge in understanding the lipidome complexity is establishing the tissues of origin and uptake for various plasma lipids, which is valuable for determining lipid functions. Using cold exposure as an acute stress, we performed global lipidomics on plasma and in nine tissues that may contribute to the circulating lipid pool. We found that numerous species of plasma acylcarnitines (ACars) and ceramides (Cers) were significantly altered upon cold exposure. Through computational assessment, we identified the liver and brown adipose tissue as major contributors and consumers of circulating ACars, in agreement with our previous work. We further identified the kidney and intestine as novel contributors to the circulating ACar pool and validated these findings with gene expression analysis. Regression analysis also identified that the brown adipose tissue and kidney are interactors with the plasma Cer pool. Taken together, these studies provide an adaptable computational tool to assess tissue contribution to the plasma lipid pool. Our findings have further implications in understanding the function of plasma ACars and Cers, which are elevated in metabolic diseases.