Publications - 2024

Lost in translation: challenges of current pharmacotherapy for sarcopenia
Tsai Shih-Yin
Trends in Molecular Medicine, Volume 0, Issue 0
Abstract
A healthy lifespan relies on independent living, in which active skeletal muscle is a critical element. The cost of not recognizing and acting earlier on unhealthy or aging muscle could be detrimental, since muscular weakness is inversely associated with all-cause mortality. Sarcopenia is characterized by a decline in skeletal muscle mass and strength and is associated with aging. Exercise is the only effective therapy to delay sarcopenia development and improve muscle health in older adults. Although numerous interventions have been proposed to reduce sarcopenia, none has yet succeeded in clinical trials. This review evaluates the biological gap between recent clinical trials targeting sarcopenia and the preclinical studies on which they are based, and suggests an alternative approach to bridge the discrepancy.

The role of artificial intelligence in knowledge management for medical students and doctors
W.A. Nathasha V. Luke & Shing Chuan Hooi
Medical Teacher
Abstract
The skill of knowledge management empowers practitioners to efficiently integrate and apply knowledge in the clinical context. Current medical knowledge is vast which is beyond the human capacity to retain. Hence, modern-day learning is not just knowledge acquisition, but the organization of knowledge in a retrievable manner. Advancing technology in digital learning spaces and artificial intelligence enables the development of personalized knowledge platforms. Clinicians should find their own ideal spaces for knowledge management from the early stages of their careers, and develop it into a lifelong learning platform, which will eventually lead to better patient care.

Neurokinin1 − cholinergic receptor mechanisms in the medial Septum-Dorsal hippocampus axis mediates experimental neuropathic pain
Mohammed Zacky Ariffin, Si Yun Ng, Hamzah Nadia, Darrel Koh, Natasha Loh, Naomi Michiko, Sanjay Khanna
Neurobiology of Pain, Volume 16, 2024, 100162, ISSN 2452-073X. 
Abstract
The neurokinin-1 receptors (NK1Rs) in the forebrain medial septum (MS) region are localized exclusively on cholinergic neurons that partly project to the hippocampus and the cingulate cortex (Cg), regions implicated in nociception. In the present study, we explored the hypothesis that neurotransmission at septal NK1R and hippocampal cholinergic mechanisms mediate experimental neuropathic pain in the rodent chronic constriction injury model (CCI). Our investigations showed that intraseptal microinjection of substance P (SP) in rat evoked a peripheral hypersensitivity (PH)-like response in uninjured animals that was attenuated by systemic atropine sulphate, a muscarinic-cholinergic receptor antagonist. Conversely, pre-emptive destruction of septal cholinergic neurons attenuated the development of PH in the CCI model that also prevented the expression of cellular markers of nociception in the spinal cord and the forebrain. Likewise, anti-nociception was evoked on intraseptal microinjection of L-733,060, an antagonist at NK1Rs, and on bilateral or unilateral microinjection of the cholinergic receptor antagonists, atropine or mecamylamine, into the different regions of the dorsal hippocampus (dH) or on bilateral microinjection into the Cg. Interestingly, the effect of L-733,060 was accompanied with a widespread decreased in levels of CCI-induced nociceptive cellular markers in forebrain that was not secondary to behaviour, suggesting an active modulation of nociceptive processing by transmission at NK1R in the medial septum. The preceding suggest that the development and maintenance of neuropathic nociception is facilitated by septal NK1R-dH cholinergic mechanisms which co-ordinately affect nociceptive processing in the dH and the Cg. Additionally, the data points to a potential strategy for pain modulation that combines anticholinergics and anti-NKRs.

Synapses tagged, memories kept: synaptic tagging and capture hypothesis in brain health and disease
Mohammad Zaki Bin Ibrahim, Zijun Wang and Sreedharan Sajikumar
Phil. Trans. R. Soc. B 379:20230237.
Abstract
The synaptic tagging and capture (STC) hypothesis lays the framework on the synapse-specific mechanism of protein synthesis-dependent long-term plasticity upon synaptic induction. Activated synapses will display a transient tag that will capture plasticity-related products (PRPs). These two events, tag setting and PRP synthesis, can be teased apart and have been studied extensively—from their electrophysiological and pharmacological properties to the molecular events involved. Consequently, the hypothesis also permits interactions of synaptic populations that encode different memories within the same neuronal population—hence, it gives rise to the associativity of plasticity. In this review, the recent advances and progress since the experimental debut of the STC hypothesis will be shared. This includes the role of neuromodulation in PRP synthesis and tag integrity, behavioural correlates of the hypothesis and modelling in silico. STC, as a more sensitive assay for synaptic health, can also assess neuronal aberrations. We will also expound how synaptic plasticity and associativity are altered in ageing-related decline and pathological conditions such as juvenile stress, cancer, sleep deprivation and Alzheimer’s disease.

This article is part of a discussion meeting issue ‘Long-term potentiation: 50 years on’.

PHF2 regulates genome topology and DNA replication in neural stem cells via cohesin
Jia Feng, You Heng Chuah, Yajing Liang, Nadia Omega Cipta, Yingying Zeng, Tushar Warrier, Gamal Ahmed Rashed Elsayed Elfar, Jeehyun Yoon, Oleg V Grinchuk, Emmy Xue Yun Tay, Ker-Zhing Lok, Zong-Qing Zheng, Zi Jian Khong, Zheng-Shan Chong, Jackie Teo, Emma May Sanford, Cheryl Jia Yi Neo, Hsin Yao Chiu, Jia Yu Leung, Loo Chien Wang, Yan Ting Lim, Tianyun Zhao, Radoslaw M Sobota, Karen Carmelina Crasta, Vinay Tergaonkar, Reshma Taneja, Shi-Yan Ng, Chit Fang Cheok, Shuo-Chien Ling, Yuin-Han Loh, Derrick Sek Tong Ong
Nucleic Acids Research, Volume 52, Issue 12, 8 July 2024, Pages 7063–7080
Abstract
Cohesin plays a crucial role in the organization of topologically-associated domains (TADs), which influence gene expression and DNA replication timing. Whether epigenetic regulators may affect TADs via cohesin to mediate DNA replication remains elusive. Here, we discover that the histone demethylase PHF2 associates with RAD21, a core subunit of cohesin, to regulate DNA replication in mouse neural stem cells (NSC). PHF2 loss impairs DNA replication due to the activation of dormant replication origins in NSC. Notably, the PHF2/RAD21 co-bound genomic regions are characterized by CTCF enrichment and epigenomic features that resemble efficient, active replication origins, and can act as boundaries to separate adjacent domains. Accordingly, PHF2 loss weakens TADs and chromatin loops at the co-bound loci due to reduced RAD21 occupancy. The observed topological and DNA replication defects in PHF2 KO NSC support a cohesin-dependent mechanism. Furthermore, we demonstrate that the PHF2/RAD21 complex exerts little effect on gene regulation, and that PHF2’s histone-demethylase activity is dispensable for normal DNA replication and proliferation of NSC. We propose that PHF2 may serve as a topological accessory to cohesin for cohesin localization to TADs and chromatin loops, where cohesin represses dormant replication origins directly or indirectly, to sustain DNA replication in NSC.

A novel MTORC2-AKT-ROS axis triggers mitofission and mitophagy-associated execution of colorectal cancer cells upon drug-induced activation of mutant KRAS
Kartini Iskandar, Jonathan Foo, Angeline Qiu Xia Liew, Haiyuxin Zhu, Deepika Raman, Jayshree L. Hirpara, Yan Yi Leong, Maria V. Babak, Anna A. Kirsanova, Anne-Sophie Armand, Franck Oury, Gregory Bellot & Shazib Pervaiz
Autophagy 2024, Vol. 20, No. 6, 1418–1441
Abstract
RAS is one of the most commonly mutated oncogenes associated with multiple cancer hallmarks. Notably, RAS activation induces intracellular reactive oxygen species (ROS) generation, which we previously demonstrated as a trigger for autophagy-associated execution of mutant KRAS-expressing cancer cells. Here we report that drug (merodantoin; C1)-induced activation of mutant KRAS promotes phospho-AKT S473-dependent ROS-mediated S616 phosphorylation and mitochondrial localization of DNM1L/DRP1 (dynamin 1 like) and cleavage of the fusion-associated protein OPA1 (OPA1 mitochondrial dynamin like GTPase). Interestingly, accumulation of the outer mitochondrial membrane protein VDAC1 (voltage dependent anion channel 1) is observed in mutant KRAS-expressing cells upon exposure to C1. Conversely, silencing VDAC1 abolishes C1-induced mitophagy, and gene knockdown of either KRAS, AKT or DNM1L rescues ROS-dependent VDAC1 accumulation and stability, thus suggesting an axis of mutant active KRAS-phospho-AKT S473-ROS-DNM1L-VDAC1 in mitochondrial morphology change and cancer cell execution. Importantly, we identified MTOR (mechanistic target of rapamycin kinsase) complex 2 (MTORC2) as the upstream mediator of AKT phosphorylation at S473 in our model. Pharmacological or genetic inhibition of MTORC2 abrogated C1-induced phosphorylation of AKT S473, ROS generation and mitophagy induction, as well as rescued tumor colony forming ability and migratory capacity. Finally, increase in thermal stability of KRAS, AKT and DNM1L were observed upon exposure to C1 only in mutant KRAS-expressing cells. Taken together, our work has unraveled a novel mechanism of selective targeting of mutant KRAS-expressing cancers via MTORC2-mediated AKT activation and ROS-dependent mitofission, which could have potential therapeutic implications given the relative lack of direct RAS-targeting strategies in cancer.

Is ChatGPT ‘ready’ to be a learning tool for medical undergraduates and will it perform equally in different subjects? Comparative study of ChatGPT performance in tutorial and case-based learning questions in physiology and biochemistry
W. A. Nathasha V. Luke, Lee Seow Chong, Kenneth H. Ban, Amanda H. Wong, Chen Zhi Xiong, Lee Shuh Shing, Reshma Taneja, Dujeepa D. Samarasekera and Celestial T. Yap
Medical Teacher, 1–7.
Abstract
Generative AI will become an integral part of education in future. The potential of this technology in different disciplines should be identified to promote effective adoption. This study evaluated the performance of ChatGPT in tutorial and case-based learning questions in physiology and biochemistry for medical undergraduates. Our study mainly focused on the performance of GPT-3.5 version while a subgroup was comparatively assessed on GPT-3.5 and GPT-4 performances.

BRD4 isoforms have distinct roles in tumour progression and metastasis in rhabdomyosarcoma
Dipanwita Das, Jia Yu Leung, Shivaranjani Balamurugan, Vinay Tergaonkar, Amos Hong Pheng Loh, Cheng-Ming Chiang and Reshma Taneja
Abstract
BRD4, a bromodomain and extraterminal (BET) protein, is deregulated in multiple cancers and has emerged as a promising drug target. However, the function of the two main BRD4 isoforms (BRD4-L and BRD4-S) has not been analysed in parallel in most cancers. This complicates determining therapeutic efficacy of pan-BET inhibitors. In this study, using functional and transcriptomic analysis, we show that BRD-L and BRD4-S isoforms play distinct roles in fusion negative embryonal rhabdomyosarcoma. BRD4-L has an oncogenic role and inhibits myogenic differentiation, at least in part, by activating myostatin expression. Depletion of BRD4-L in vivo impairs tumour progression but does not impact metastasis. On the other hand, depletion of BRD4-S has no significant impact on tumour growth, but strikingly promotes metastasis in vivo. Interestingly, BRD4-S loss results in the enrichment of BRD4-L and RNA Polymerase II at integrin gene promoters resulting in their activation. In fusion positive alveolar rhabdomyosarcoma, BRD4-L is unrestricted in its oncogenic role, with no evident involvement of BRD4-S. Our work unveils isoform-specific functions of BRD4 in rhabdomyosarcoma.

PHF2 regulates genome topology and DNA replication in neural stem cells via cohesin
Derrick Sek Tong Ong
Nucleic Acids Res. 2024 Jul 8;52(12):7063-7080. doi: 10.1093/nar/gkae457.
Abstract
Cohesin plays a crucial role in the organization of topologically-associated domains (TADs), which influence gene expression and DNA replication timing. Whether epigenetic regulators may affect TADs via cohesin to mediate DNA replication remains elusive. Here, we discover that the histone demethylase PHF2 associates with RAD21, a core subunit of cohesin, to regulate DNA replication in mouse neural stem cells (NSC). PHF2 loss impairs DNA replication due to the activation of dormant replication origins in NSC. Notably, the PHF2/RAD21 co-bound genomic regions are characterized by CTCF enrichment and epigenomic features that resemble efficient, active replication origins, and can act as boundaries to separate adjacent domains. Accordingly, PHF2 loss weakens TADs and chromatin loops at the co-bound loci due to reduced RAD21 occupancy. The observed topological and DNA replication defects in PHF2 KO NSC support a cohesin-dependent mechanism. Furthermore, we demonstrate that the PHF2/RAD21 complex exerts little effect on gene regulation, and that PHF2's histone-demethylase activity is dispensable for normal DNA replication and proliferation of NSC. We propose that PHF2 may serve as a topological accessory to cohesin for cohesin localization to TADs and chromatin loops, where cohesin represses dormant replication origins directly or indirectly, to sustain DNA replication in NSC.

A novel MTORC2-AKT-ROS axis triggers mitofission and mitophagy-associated execution of colorectal cancer cells upon drug-induced activation of mutant KRAS
Shazib Pervaiz
Autophagy. 2024 Jun;20(6):1418-1441. doi: 10.1080/15548627.2024.2307224.
Abstract
RAS is one of the most commonly mutated oncogenes associated with multiple cancer hallmarks. Notably, RAS activation induces intracellular reactive oxygen species (ROS) generation, which we previously demonstrated as a trigger for autophagy-associated execution of mutant KRAS-expressing cancer cells. Here we report that drug (merodantoin; C1)-induced activation of mutant KRAS promotes phospho-AKT S473-dependent ROS-mediated S616 phosphorylation and mitochondrial localization of DNM1L/DRP1 (dynamin 1 like) and cleavage of the fusion-associated protein OPA1 (OPA1 mitochondrial dynamin like GTPase). Interestingly, accumulation of the outer mitochondrial membrane protein VDAC1 (voltage dependent anion channel 1) is observed in mutant KRAS-expressing cells upon exposure to C1. Conversely, silencing VDAC1 abolishes C1-induced mitophagy, and gene knockdown of either KRAS, AKT or DNM1L rescues ROS-dependent VDAC1 accumulation and stability, thus suggesting an axis of mutant active KRAS-phospho-AKT S473-ROS-DNM1L-VDAC1 in mitochondrial morphology change and cancer cell execution. Importantly, we identified MTOR (mechanistic target of rapamycin kinsase) complex 2 (MTORC2) as the upstream mediator of AKT phosphorylation at S473 in our model. Pharmacological or genetic inhibition of MTORC2 abrogated C1-induced phosphorylation of AKT S473, ROS generation and mitophagy induction, as well as rescued tumor colony forming ability and migratory capacity. Finally, increase in thermal stability of KRAS, AKT and DNM1L were observed upon exposure to C1 only in mutant KRAS-expressing cells. Taken together, our work has unraveled a novel mechanism of selective targeting of mutant KRAS-expressing cancers via MTORC2-mediated AKT activation and ROS-dependent mitofission, which could have potential therapeutic implications given the relative lack of direct RAS-targeting strategies in cancer.

Secretome from Magnetically Stimulated Muscle Exhibits Anticancer Potency: Novel Preconditioning Methodology Highlighting HTRA1 Action
Alfredo Franco-Obregón
Bioengineering 2024, 11(7), 637;
Abstract
Briefly (10 min) exposing C2C12 myotubes to low amplitude (1.5 mT) pulsed electromagnetic fields (PEMFs) generated a conditioned media (pCM) that was capable of mitigating breast cancer cell growth, migration, and invasiveness in vitro, whereas the conditioned media harvested from unexposed myotubes, representing constitutively released secretome (cCM), was less effective. Administering pCM to breast cancer microtumors engrafted onto the chorioallantoic membrane of chicken eggs reduced tumor volume and vascularity. Blood serum collected from PEMF-exposed or exercised mice allayed breast cancer cell growth, migration, and invasiveness. A secretome preconditioning methodology is presented that accentuates the graded anticancer potencies of both the cCM and pCM harvested from myotubes, demonstrating an adaptive response to pCM administered during early myogenesis that emulated secretome-based exercise adaptations observed in vivo. HTRA1 was shown to be upregulated in pCM and was demonstrated to be necessary and sufficient for the anticancer potency of the pCM; recombinant HTRA1 added to basal media recapitulated the anticancer effects of pCM and antibody-based absorption of HTRA1 from pCM precluded its anticancer effects. Brief and non-invasive PEMF stimulation may represent a method to commandeer the secretome response of muscle, both in vitro and in vivo, for clinical exploitation in breast and other cancers.

Synergistic Cellular Responses Conferred by Concurrent Optical and Magnetic Stimulation Are Attenuated by Simultaneous Exposure to Streptomycin: An Antibiotic Dilemma
Alfredo Franco-Obregón
Bioengineering 2024, 11(7), 637;
Abstract
Concurrent optical and magnetic stimulation (COMS) combines extremely low-frequency electromagnetic and light exposure for enhanced wound healing. We investigated the potential mechanistic synergism between the magnetic and light components of COMS by comparing their individual and combined cellular responses. Lone magnetic field exposure produced greater enhancements in cell proliferation than light alone, yet the combined effects of magnetic fields and light were supra-additive of the individual responses. Reactive oxygen species were incrementally reduced by exposure to light, magnetics fields, and their combination, wherein statistical significance was only achieved by the combined COMS modality. By contrast, ATP production was most greatly enhanced by magnetic exposure in combination with light, indicating that mitochondrial respiratory efficiency was improved by the combination of magnetic fields plus light. Protein expression pertaining to cell proliferation was preferentially enhanced by the COMS modality, as were the protein levels of the TRPC1 cation channel that had been previously implicated as part of a calcium–mitochondrial signaling axis invoked by electromagnetic exposure and necessary for proliferation. These results indicate that light facilitates functional synergism with magnetic fields that ultimately impinge on mitochondria-dependent developmental responses. Aminoglycoside antibiotics (AGAs) have been previously shown to inhibit TRPC1-mediated magnetotransduction, whereas their influence over photomodulation has not been explored. Streptomycin applied during exposure to light, magnetic fields, or COMS reduced their respective proliferation enhancements, whereas streptomycin added after the exposure did not. Magnetic field exposure and the COMS modality were capable of partially overcoming the antagonism of proliferation produced by streptomycin treatment, whereas light alone was not. The antagonism of photon-electromagnetic effects by streptomycin implicates TRPC1-mediated calcium entry in both magnetotransduction and photomodulation. Avoiding the prophylactic use of AGAs during COMS therapy will be crucial for maintaining clinical efficacy and is a common concern in most other electromagnetic regenerative paradigms.

Telomere shortening induces aging-associated phenotypes in hiPSC-derived neurons and astrocytes
Derrick Sek Tong Ong
Biogerontology. 2024 Apr;25(2):341-360.
doi: 10.1007/s10522-023-10076-5. Epub 2023 Nov 21.
Abstract
Telomere shortening is a well-established hallmark of cellular aging. Telomerase reverse transcriptase (TERT) plays a crucial role in maintaining the length of telomeres, which are specialised protective caps at the end of chromosomes. The lack of in vitro aging models, particularly for the central nervous system (CNS), has impeded progress in understanding aging and age-associated neurodegenerative diseases. In this study, we aimed to explore the possibility of inducing aging-associated features in cell types of the CNS using hiPSC (human induced pluripotent stem cell) technology. To achieve this, we utilised CRISPR/Cas9 to generate hiPSCs with a loss of telomerase function and shortened telomeres. Through directed differentiation, we generated motor neurons and astrocytes to investigate whether telomere shortening could lead to age-associated phenotypes. Our findings revealed that shortened telomeres induced age-associated characteristics in both motor neurons and astrocytes including increased cellular senescence, heightened inflammation, and elevated DNA damage. We also observed cell-type specific age-related morphology changes. Additionally, our study highlighted the fundamental role of TERT and telomere shortening in neural progenitor cell (NPC) proliferation and neuronal differentiation. This study serves as a proof of concept that telomere shortening can effectively induce aging-associated phenotypes, thereby providing a valuable tool to investigate age-related decline and neurodegenerative diseases.

Are Aminoglycoside Antibiotics TRPing Your Metabolic Switches?
Alfredo Franco-Obregón
Cells 2024, 13(15), 1273;
Abstract
Transient receptor potential (TRP) channels are broadly implicated in the developmental programs of most tissues. Amongst these tissues, skeletal muscle and adipose are noteworthy for being essential in establishing systemic metabolic balance. TRP channels respond to environmental stimuli by supplying intracellular calcium that instigates enzymatic cascades of developmental consequence and often impinge on mitochondrial function and biogenesis. Critically, aminoglycoside antibiotics (AGAs) have been shown to block the capacity of TRP channels to conduct calcium entry into the cell in response to a wide range of developmental stimuli of a biophysical nature, including mechanical, electromagnetic, thermal, and chemical. Paradoxically, in vitro paradigms commonly used to understand organismal muscle and adipose development may have been led astray by the conventional use of streptomycin, an AGA, to help prevent bacterial contamination. Accordingly, streptomycin has been shown to disrupt both in vitro and in vivo myogenesis, as well as the phenotypic switch of white adipose into beige thermogenic status. In vivo, streptomycin has been shown to disrupt TRP-mediated calcium-dependent exercise adaptations of importance to systemic metabolism. Alternatively, streptomycin has also been used to curb detrimental levels of calcium leakage into dystrophic skeletal muscle through aberrantly gated TRPC1 channels that have been shown to be involved in the etiology of X-linked muscular dystrophies. TRP channels susceptible to AGA antagonism are critically involved in modulating the development of muscle and adipose tissues that, if administered to behaving animals, may translate to systemwide metabolic disruption. Regenerative medicine and clinical communities need to be made aware of this caveat of AGA usage and seek viable alternatives, to prevent contamination or infection in in vitro and in vivo paradigms, respectively.
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