IQCELL: A platform for predicting the effect of gene perturbations on developmental trajectories using single-cell RNA-seq data

The increasing availability of single-cell RNA-sequencing (scRNA-seq) data from various developmental systems provides the opportunity to infer gene regulatory networks (GRNs) directly from data. Herein we describe IQCELL, a platform to infer, simulate, and study executable logical GRNs directly from scRNA-seq data. Such executable GRNs allow simulation of fundamental hypotheses governing developmental programs and help accelerate the design of strategies to control stem cell fate. We first describe the architecture of IQCELL. Next, we apply IQCELL to scRNA-seq datasets from early mouse T-cell and red blood cell development, and show that the platform can infer overall over 74% of causal gene interactions previously reported from decades of research. We will also show that dynamic simulations of the generated GRN qualitatively recapitulate the effects of known gene perturbations. Finally, we implement an IQCELL gene selection pipeline that allows us to identify candidate genes, without prior knowledge. We demonstrate that GRN simulations based on the inferred set yield results similar to the original curated lists. In summary, the IQCELL platform offers a versatile tool to infer, simulate, and study executable GRNs in dynamic biological systems.     

Production of acetone,butanol and ethanol from palm oil waste by Clostridium saccharoperbutylacetonicum N1-4

Summary A study of the feasibility of utilizing palm oil waste as a substrate for acetonebutanol-ethanol fermentation by Clostridium saccharoperbutylacetonicum N1-4 and, hence, to reduce the biochemical oxygen demand (BOD) of waste disposal was carried out. Among the two types of palm oil waste tested, separator sludge was the better substrate and it was able to support production of solvents by C. saccharoperbutylacetonicum N1-4 without a need for any mineral supplements. Enzymatic hydrolysis catalyzed by cellulase prior to fermentation was found to increase the yield of butanol by 75% (from 2.47g/l to 4.37g/l) and decreased the BOD by 66% (from 26,500 ppm to 8,900 ppm).     

Cardiomyocyte specific ablation of p53 is not sufficient to block doxorubicin induced cardiac fibrosis and associated cytoskeletal changes

Doxorubicin (Dox) is an anthracycline used to effectively treat several forms of cancer. Unfortunately, the use of Dox is limited due to its association with cardiovascular complications which are manifested as acute and chronic cardiotoxicity. The pathophysiological mechanism of Dox induced cardiotoxicity appears to involve increased expression of the tumor suppressor protein p53 in cardiomyocytes, followed by cellular apoptosis. It is not known whether downregulation of p53 expression in cardiomyocytes would result in decreased rates of myocardial fibrosis which occurs in response to cardiomyocyte loss. Further, it is not known whether Dox can induce perivascular necrosis and associated fibrosis in the heart. In this study we measured the effects of acute Dox treatment on myocardial and perivascular apoptosis and fibrosis in a conditional knockout (CKO) mouse model system which harbours inactive p53 alleles specifically in cardiomyocytes. CKO mice treated with a single dose of Dox (20 mg/kg), did not display lower levels of myocardial apoptosis or reactive oxygen and nitrogen species (ROS/RNS) compared to control mice with intact p53 alleles. Interestingly, CKO mice also displayed higher levels of interstitial and perivascular fibrosis compared to controls 3 or 7 days after Dox treatment. Additionally, the decrease in levels of the microtubule protein α-tubulin, which occurs in response to Dox treatment, was not prevented in CKO mice. Overall, these results indicate that selective loss of p53 in cardiomyocytes is not sufficient to prevent Dox induced myocardial ROS/RNS generation, apoptosis, interstitial fibrosis and perivascular fibrosis. Further, these results support a role for p53 independent apoptotic pathways leading to Dox induced myocardial damage and highlight the importance of vascular lesions in Dox induced cardiotoxicity.     

Role of D-type cyclins in heart development and disease

A defining feature of embryonic cardiomyocytes is their relatively high rates of proliferation. A gradual reduction in proliferative capacity throughout development culminates in permanent cell cycle exit by the vast majority of cardiomyocytes around the perinatal period. Accordingly, the adult heart has severely limited capacity for regeneration in response to injury or disease. The D-type cyclins (cyclin D1, D2, and D3) along with their catalytically active partners, the cyclin dependent kinases, are positive cell cycle regulators that play important roles in regulating proliferation of cardiomyocytes during normal heart development. While expression of D-type cyclins is generally low in the adult heart, expression levels are augmented in association with cardiac hypertrophy, but are uncoupled from myocyte cell division. Accordingly, re-activation of D-type cyclin expression in the adult heart has been implicated in pathophysiological processes via mechanisms distinct from those that drive proliferation during cardiac development. Growth factors and other exogenous agents regulate D-type cyclin production and activity in embryonic and adult cardiomyocytes. Understanding differences in the precise intracellular mediators downstream from these signalling molecules in embryonic versus adult cardiomyocytes could prove valuable for designing strategies to reactivate the cell cycle in cardiomyocytes in the setting of cardiovascular disease in the adult heart.     

A mouse model of inherited choline kinase β-deficiency presents with specific cardiac abnormalities and a predisposition to arrhythmia

The CHKB gene encodes choline kinase β, which catalyzes the first step in the biosynthetic pathway for the major phospholipid phosphatidylcholine. Homozygous loss-of-function variants in human CHKB are associated with a congenital muscular dystrophy. Dilated cardiomyopathy is present in some CHKB patients and can cause heart failure and death. Mechanisms underlying a cardiac phenotype due to decreased CHKB levels are not well characterized. We determined that there is cardiac hypertrophy in Chkb^−/− mice along with a decrease in left ventricle size, internal diameter, and stroke volume compared with wildtype and Chkb^+/− mice. Unlike wildtype mice, 60% of the Chkb^+/− and all Chkb^−/− mice tested displayed arrhythmic events when challenged with isoproterenol. Lipidomic analysis revealed that the major change in lipid level in Chkb^+/− and Chkb^−/− hearts was an increase in the arrhythmogenic lipid acylcarnitine. An increase in acylcarnitine level is also associated with a defect in the ability of mitochondria to use fatty acids for energy and we observed that mitochondria from Chkb^−/− hearts had abnormal cristae and inefficient electron transport chain activity. Atrial natriuretic peptide (ANP) is a hormone produced by the heart that protects against the development of heart failure including ventricular conduction defects. We determined that there was a decrease in expression of ANP, its receptor NPRA, as well as ventricular conduction system markers in Chkb^+/− and Chkb^−/− mice.     

DNA Detection Based on Localized Surface Plasmon Resonance Spectroscopy of Ag@Au Biocomposite Nanoparticles

Noble metal nanoparticles (NPs) have attracted much attention due to their unique physical and chemical properties such as tunable surface plasmonics, high-efficiency electrochemical sensing, and enhanced fluorescence. We produced two biosensor chips consisting of Ag@Au bimetallic nanoparticles (BNPs) on a carbon thin film by simple RF-sputtering and RF-plasma-enhanced chemical vapor co-deposition. We deposited Au NPs with average size of 4 nm (Au₁ NPs) or 11 nm (Au₂ NPs) on a sensor chip consisting of Ag NPs with mean size of 15 nm, and we investigated the effect of shell size (Au NPs) on the chemical activities of the resulting Ag@Au₁ BNPs and Ag@Au₂ BNPs. We estimated the average size and morphology of Ag@Au BNPs by scanning electron microscopy (SEM) and atomic force microscopy (AFM) images. X-ray diffraction (XRD) patterns revealed that Ag NPs and Au NPs had face-centered cubic (FCC) structure. We studied aging of the biosensor chips consisting of Ag@Au BNPs by localized surface plasmon resonance (LSPR) spectroscopy for up to 3 months. UV–visible aging of the prepared samples indicated that Ag@Au₁ BNPs, which corresponded to Ag NPs covered with smaller Au NPs, were more chemically active than Ag@Au₂ BNPs. Furthermore, we evaluated changes in the LSPR absorption peaks of Ag@Au₁ BNPs and bare Ag NPs in the presence of a DNA primer decamer at fM concentrations, to find that Ag@Au₁ BNPs were more sensitive biosensor chips within a short response time as compared to bare Ag NPs.