Biology subjects

Lam, A.

Publications and source records attributed to Lam, A..

43 records · Page 3Linked to original sources

bioRxiv2021

Inhibition of BKCa channels protects neonatal hearts against myocardial ischemia and reperfusion injury

BKCa channels are large-conductance calcium and voltage-activated potassium channels that are heterogeneously expressed in a wide array of cells. Activation of BKCa channels present in mitochondria of adult ventricular cardiomyocytes is implicated in cardioprotection against ischemia-reperfusion (IR) injury. However, the BKCa channels activity has never been detected in the plasma membrane of adult ventricular cardiomyocytes. In this study, we report the presence of the BKCa channel in the plasma membrane and mitochondria of neonatal murine and rodent cardiomyocytes which protects the heart on inhibition but not activation. Furthermore, K+ currents measured in neonatal cardiomyocyte (NCM) was sensitive to iberiotoxin (IbTx), suggesting the presence of BKCa channels in the plasma membrane. Neonatal hearts subjected to IR when post-conditioned with NS1619 during reoxygenation increased the myocardial infarction whereas IbTx reduced the infarct size. In agreement, isolated NCM also presented increased apoptosis on treatment with NS1619 during hypoxia and reoxygenation, whereas IbTx reduced TUNEL positive cells. In NCMs, activation of BKCa channels increased the intracellular reactive oxygen species post HR injury. Electrophysiological characterization of NCMs indicated that NS1619 increased the beat period, field, and action potential duration, and decreased the conduction velocity and spike amplitude. In contrast, IbTx had no impact on the electrophysiological properties of NCMs. Taken together, our data established that inhibition of plasma membrane BKCa channels in the NCM protects neonatal heart/cardiomyocytes from IR injury. Furthermore, the functional disparity observed towards the cardioprotective activity of BKCa channels in adults compared to neonatal heart could be attributed to their differential localization.

physiology↗

bioRxiv2021

Dietary vitamin B12 reduces amyloid-β proteotoxicity by alleviating oxidative stress and mitochondrial dysfunction

Alzheimers disease (AD) is a devastating neurodegenerative disorder with no effective treatment. Diet, as a modifiable risk factor for AD, could potentially be targeted to slow disease onset and progression. However, complexity of the human diet and indirect effects of the microbiome make it challenging to identify protective nutrients. Multiple factors contribute to AD pathogenesis including amyloid beta (A{beta}) deposition, mitochondrial dysfunction, and oxidative stress. Here we used Caenorhabditis elegans to define the impact of diet on A{beta} proteotoxicity. We discovered that dietary vitamin B12 alleviated mitochondrial fragmentation, bioenergetic defects, and oxidative stress, delaying A{beta}-induced paralysis without affecting A{beta} accumulation. Vitamin B12 had this protective effect by acting as a cofactor for methionine synthase rather than as an antioxidant. Vitamin supplementation of B12 deficient adult A{beta} animals was beneficial, demonstrating potential for vitamin B12 as a therapy to target pathogenic features of AD triggered by both aging and proteotoxic stress.

cell biology↗

bioRxiv2020

A C. elegans genome-wide RNAi screen for altered levamisole sensitivity identifies genes required for muscle function

At the neuromuscular junction (NMJ), postsynaptic ionotropic acetylcholine receptors (AChRs) transduce a chemical signal released from a cholinergic motor neuron into an electrical signal to induce muscle contraction. To identify regulators of postsynaptic function, we conducted a genome-wide RNAi screen for genes required for proper response to levamisole, a pharmacological agonist of ionotropic L-AChRs at the Caenorhabditis elegans NMJ. A total of 117 gene knockdowns were found to cause levamisole hypersensitivity, while 18 resulted in levamisole resistance. Our screen identified conserved genes important for muscle function including some that are mutated in congenital myasthenic syndrome, congenital muscular dystrophy, congenital myopathy, myotonic dystrophy, and mitochondrial myopathy. Of the genes found in the screen, we further investigated those predicted to play a role in endocytosis of cell surface receptors. Loss of the Epsin homolog epn-1 had opposing effects on the levels of postsynaptic L-AChRs and GABAA receptors, resulting in increased and decreased abundance, respectively. This disrupts the balance of postsynaptic excitatory and inhibitory signaling, causing levamisole hypersensitivity. We also examined other genes that resulted in a levamisole hypersensitive phenotype when knocked down including gas-1, which functions in Complex I of the mitochondrial electron transport chain. Consistent with altered ATP synthesis impacting levamisole response, treatment of wild-type animals with levamisole resulted in L-AChR dependent depletion of ATP levels. These results suggest that the paralytic effects of levamisole ultimately lead to metabolic exhaustion.

genetics↗

bioRxiv2019

Plasmid transmitted small multidrug resistant (SMR) efflux pumps differ in gene regulation and enhance tolerance to quaternary ammonium compounds (QAC) when grown as biofilms

Small multidrug resistance (SMR) efflux pump genes are commonly identified from integrons carried by multidrug-resistant (MDR) plasmids. SMR pumps are annotated as qac for their ability to confer resistance to quaternary ammonium compounds (QACs) but few qac are characterized to date. Hence, we have examined SMR sequence diversity, antimicrobial susceptibility, and gene expression from >500 sequenced proteobacterial plasmids. SMR sequence diversity from plasmid database surveys identified 20 unique SMR sequences annotated as qacE/E{Delta}1/F/G/H/I/L, or sugE. Phylogenetic analysis shows Qac sequences are homologous to archetypical SMR member EmrE, and share a single sequence origin. In contrast, SugE sequences are homologous to archetypical member Gdx/SugE and likely originate from different species. SMR genes, qacE, qacE{Delta}1, qacF, qacG, qacH, and sugE(p), were over-expressed in Escherichia coli to determine their QAC antimicrobial susceptibility as planktonic, colony, and biofilms. SMRs (except qacE{Delta}1/sugE) expressed in biofilms significantly increased its QAC tolerance as compared to planktonic and colony growth. Analysis of upstream SMR nucleotide regions indicate sugE(p) genes are regulated by type II guanidinium riboswitches, whereas qacE and qacE{Delta}1 have a conserved class I integron Pq promoter, and qacF/G/H are regulated by integron Pc promoter in variable cassettes region. Beta-galactosidase assays were used to characterize growth conditions regulating Pq and Pc promoters and revealed that Pq and Pc have different expression profiles during heat, peroxide, and QAC exposure. Altogether, this study reveals that biofilm growth methods are optimal for SMR-mediated QAC susceptibility testing and suggests SMR gene regulation on plasmids is similar to chromosomally inherited SMR members.

microbiology↗

bioRxiv2019

Human cardiac-fibrosis-on-a-chip model recapitulates disease hallmarks and can serve as a platform for drug screening

While interstitial fibrosis plays a significant role in heart failure, our understanding of disease progression in humans is limited. To address this limitation, we have engineered a cardiac-fibrosis-on-a-chip model consisting of a microfabricated device with live force measurement capabilities using co-cultured human cardiac fibroblasts and pluripotent stem cell-derived cardiomyocytes. Transforming growth factor-{beta} was used as a trigger for fibrosis. Here, we have reproduced the classic hallmarks of fibrosis-induced heart failure including high collagen deposition, increased tissue stiffness, BNP secretion, and passive tension. Force of contraction was significantly decreased in fibrotic tissues that displayed a transcriptomic signature consistent with human cardiac fibrosis/heart failure. Treatment with an anti-fibrotic drug decreased tissue stiffness and BNP secretion, with corresponding changes in the transcriptomic signature. This model represents an accessible approach to study human heart failure in vitro, and allows for testing anti-fibrotic drugs while facilitating the real-time assessment of cardiomyocyte function.

bioengineering↗

bioRxiv2018

A comprehensive analysis of the usability and archival stability of omics computational tools and resources

Developing new software tools for analysis of large-scale biological data is a key component of advancing modern biomedical research. Scientific reproduction of published findings requires running computational tools on data generated by such studies, yet little attention is presently allocated to the installability and archival stability of computational software tools. Scientific journals require data and code sharing, but none currently require authors to guarantee the continuing functionality of newly published tools. We have estimated the archival stability of computational biology software tools by performing an empirical analysis of the internet presence for 36,702 omics software resources published from 2005 to 2017. We found that almost 28% of all resources are currently not accessible through URLs published in the paper they first appeared in. Among the 98 software tools selected for our installability test, 51% were deemed \"easy to install,\" and 28% of the tools failed to be installed at all due to problems in the implementation. Moreover, for papers introducing new software, we found that the number of citations significantly increased when authors provided an easy installation process. We propose for incorporation into journal policy several practical solutions for increasing the widespread installability and archival stability of published bioinformatics software.

bioinformatics↗

bioRxiv2018

Microtubules Gate Tau Condensation to Spatially Regulate Microtubule Functions

Tau is an abundant microtubule-associated protein in neurons. Tau aggregation into insoluble fibrils is a hallmark of Alzheimers disease and other dementias, yet the physiological state of tau molecules within cells remains unclear. Using single molecule imaging, we directly observe that the microtubule lattice regulates reversible tau self-association, leading to dynamic condensation of tau molecules on the microtubule surface. Tau condensates form selectively permissible barriers, spatially regulating the activity of MT severing enzymes and the movement of molecular motors through their boundaries. We propose that reversible self-association of tau molecules, controlled by the microtubule, is an important mechanism of taus biological functions, and that oligomerization of tau is a common property shared between the physiological and disease forms of the molecule.\n\nOne Sentence SummaryReversible tau oligomerization regulates microtubule functions.

biophysics↗