NeuroBiology & Genetics Group
"To educate and inspire through discoveries"
RESEARCH FOCUS
Neuropathology and Intellectual Disability in Down Syndrome
Down syndrome (DS) is the most common genetic cause of intellectual disability, and its hallmark brain abnormalities include fewer neurons and excess astroglia. Studies of DS brains and mouse models reveal that an extra chromosome 21 disrupts normal neurodevelopment – DS brains have more astrocytes and fewer neurons than usual. Using a DS mouse model (Ts1Cje) and human induced pluripotent stem cell (hiPSC)-derived neural models, our team showed that trisomy causes various neuropathological features and widespread gene expression changes. These changes impair neurogenesis: neural stem cells in DS tend to overproduce glial cells at the expense of neurons, which likely contributes to cognitive deficits. This neuropathology underlies the mild to severe intellectual disability seen in DS individuals and even predisposes them to early-onset Alzheimer’s disease.
Figure 1: In a Down syndrome model (Ts1Cje mouse), neural progenitors produce markedly fewer neurons (red) compared to normal (disomic) controls, contributing to intellectual disability. Data show ~56% reduction in new neurons alongside a slight increase in astrocytes in DS cultures (https://doi.org/10.1371/journal.pone.0011561)
Figure 2: Fluorescent micrographs for immunocytochemical characterisation of iPSC, NSC and neurons. Nuclei were counterstained with DAPI. (A) Positive
nuclear OCT4 staining indicates strong expression of OCT4 as a pluripotent marker in the iPSCs. Some cells that are potentially undergoing mitosis can be detected (yellow arrow), which indicates the actively proliferative characteristic of iPSCs. (B) Cytoplasmic NESTIN staining indicates positive expression of the neural stem/progenitor marker in the NSCs. (C) Positive TUJ1 staining indicates positive expression of the post-mitotic neuronal marker in the neurons. Scale bar: 50 μm (https://doi.org/10.1016/j.neuroscience.2024.12.061)
Organoid Models for Brain Development and Function
Three-dimensional brain organoids derived from human induced pluripotent stem cells (hiPSCs) have transformed the way we investigate human brain development and neurological disorders. In our research, these organoids act as powerful platforms to study cellular and molecular changes in Down syndrome (DS). We demonstrated that DS-derived organoids exhibit delayed neuronal differentiation, altered cortical organization, and metabolic stress, which mirror early developmental abnormalities in the DS brain. Beyond DS, we extended organoid technology to model other complex brain disorders. In Alzheimer’s disease models, early amyloid pathology and synaptic dysfunction were observed within organoids, providing a window into the earliest stages of neurodegeneration. Similarly, organoids modeling bipolar disorder showed abnormal neurodevelopmental trajectories and synaptic network changes, uncovering disease-related mechanisms in a human-relevant system. Together, these studies highlight how organoids bridge the gap between basic neuroscience and translational research, enabling the discovery of mechanisms and the testing of therapeutic strategies in multiple brain disorders.
Figure 3: Signalling pathways and biological functions enriched in the commonly involved REST-targeted DEGs at different ages (PCW = post-conception weeks, M = month, Y = year) and different DS neural models (organoids, NPCs, neurons, and astrocytes) (https://doi.org/10.3390/ijms24129980)
Figure 4: Increased neuronal gene expression and network activity in BD cerebral organoids. (A) Unsupervised hierarchical clustering of genes
differentially expressed between CTR and BD cerebral organoids after 46 days of maturation. Differentially expressed genes (rows) and organoids (columns) were clustered, and gene expression was transformed to a z-score and represented in the heat map. (B) GO biological process (GoBP) groups enriched in differentially expressed genes in BD compared to CTR organoids. (C) Representative image of two live-sections from cerebral organoids at DIV60 (left) and a representative image showing live calcium imaging in live sections. (D) Mean firing rate of neurons that do not participate in synchronous firing events (Un-synchronised) and neurons that participate in synchronous, burst firing (Synchronised). (E, F) Width and Interval in seconds of each firing peak of synchronised neurons derived from data collected in (D). (G) Amplitude of individual firing peaks of synchronised neurons derived from data collected in (D). (https://doi.org/10.1038/s41380-023-02313-7)
Molecular Pathways Underlying Brain Disorders
Understanding the genetic and molecular circuitry of the brain is key to unravelling disorders like DS. Our research explores pathways that regulate brain cell fate and function. One focus is the REST protein – a master gene regulator that normally silences neuron-specific genes. In DS brains, we found that REST levels are abnormally low, leading to the overactivation of many of its target genes. This unleashes downstream pathways such as JAK–STAT immune signalling and HIF-1 oxidative stress responses, which we detected as consistently overactive across DS brain regions. Such dysregulation likely drives excessive inflammation and glial proliferation in DS, exacerbating neurological issues. We also linked DS cognitive decline to brain insulin resistance and overactivation of the PI3K–Akt/mTOR pathway, which is associated with Alzheimer-like changes; we proposed this pathway as a promising therapeutic target to slow or prevent dementia in DS. We also apply genomic tools to map network disruptions, for instance, transcriptomic analyses in DS models have highlighted interferon and immune-related gene networks being upended, linking inflammation to impaired brain development. Beyond protein-coding genes, we investigate non-coding RNAs (like microRNAs) as regulatory players. These small RNAs have emerged as important in cognitive development, and our review of intellectual disability syndromes noted that many microRNAs are misregulated in DS and other forms of hereditary intellectual disability. By charting these molecular pathways and genetic factors, we aim to pinpoint the root causes of neurodevelopmental disorders and identify targets for intervention.
Figure 5: Insulin receptor signalling pathway. Binding insulin to α-subunits on the cell surface triggers a change in the formation of intracellular β-subunits through dimerisation and autophosphorylation. This activates tyrosine kinase (e.g., Tyr1158/1162/1163) and phosphorylates IRS-1 as a docking site for downstream proteins, connecting them to the insulin receptor and IRS-1. Phosphorylated IRS-1 recruits and activates PI3K, which phosphorylates Akt/PKB, a key node in the pathway. Akt then influences downstream cascades such as GSK3β, FOXO1, and mTORC1/2. (https://doi.org/10.1016/j.bbrc.2024.150713)
Figure 6: (a1–a3) The PPI network of DEGs in NPCs, neurons, and astrocytes, respectively, was constructed using Cytoscape. Red represents the REST-targeted upregulated genes, green represents the REST-targeted downregulated genes, yellow represents the non-REST-targeted upregulated genes, and purple represents the non-REST-targeted downregulated genes. The larger the network node degree distribution, the larger the shape. (b1–b3, c1–c3, d1–d3) The top three models obtained using the MCODE plug-in of Cytoscape in neuron DEGs in NPCs, neurons, and astrocytes, respectively. (e1–e3) The GO enrichment’s most significant five terms were shown in each of the top three critical models in neuron DEGs. (https://doi.org/10.3390/ijms24129980)
Figure 7: The illustration depicts the (a) role of microglia and astrocytes response to neuroinflammation and (b) how JAK-STAT signalling pathway are involved in the activation process. (ISBN 9781138487628)
Translational Research: From Bench to Bedside
A core goal of our work is to translate scientific insights into treatments that improve lives. We actively explore therapies that could counteract DS-related brain deficits. For example, we discovered that the drug lithium, commonly used as a mood stabilizer, can restore nuclear REST levels in DS neurons and dramatically reduce their oxidative stress. In lab tests on patient-derived cells, a 24-hour lithium treatment boosted REST in DS neurons back to normal and brought down harmful reactive oxygen species, suggesting lithium repurposing could improve neuronal health in DS. We are also investigating metabolic and signaling pathways as drug targets. On a broader front, our team evaluates interventions at multiple levels, from small molecules to gene therapy. Additionally, we have tested a JAK-STAT inhibitor (Ruxolitinib) in DS mouse models to gauge improvements in neurodevelopmental outcomes. Through such preclinical trials and collaborations, we strive to carry discoveries from the bench to the bedside, inching closer to treatments that could enhance learning, memory, and quality of life for individuals with Down syndrome. Each finding, whether positive or negative, guides us toward more effective therapies and underscores our translational mission.
Figure 7 (Left): Behavioural assessments on young adult mice underwent prenatal transient ruxolitinib treatment. (A) A schematic diagram depicting the prenatal transient ruxolitinib treatment from E7.5 to E20.5 gestational period. After delivery, the pups were allowed to grow and mature into young adulthood without further intervention before behavioural assessments. (B) The measurements of mouse weight, grip strength, forelimb strength (hanging wire test) and locomotor coordination (rotarod). (C) Open field test for explorative behaviour and the general level of hyperactivity/anxiety. Representative mouse locomotion tracking for both the vehicle control and treated groups. The number of entries and time spent in each zone (zones 1–10) within the open field are presented to the right. All comparisons. (D) Novel object recognition assessment for the mouse capability to recognise a familiar object and discriminate a novel object. (E) The bird’s-eye view of the Morris water maze setup. The arrow indicates the hidden escape platform in the middle of the north-east quadrant. (F) The proportion of distance travelled to the north-east quadrant, the number of entries to the north-east quadrant and the escape platform zone during the probe trial on day 7. (https://doi.org/10.1038/s41598-021-83222-z)
Figure 8 (Right): Three pairs of cell lines (C2 vs. DS2, C4 vs. DS3, and C5 vs. DS4) were used to perform ICC. All cells used in this study were NPCs during maintenance. All measurements were normalised by dividing DS values by the corresponding controls. (A, B) Optimisation of safety dosage of lithium carbonate and X5050 treatment in iPSC-derived NPCs through MTT cell viability assay. (C) ICC validation for REST, DCX and NFIA expressed in the Con group (Control; Disomic NPCs), Con+X5050 group (Disomic NPCs with X5050 treatment), DS+Li group (Trisomic NPCs with lithium carbonate treatment), DS+Li+X5050 (Trisomic NPCs treated with both lithium carbonate and X5050), respectively (scale bar = 50µm); (D–F) Fluorescence intensity analysis for REST, DCX and NFIA expressed in different groups. (https://doi.org/10.1038/s41598-025-87314-y)
STATS
- Total no. of grants obtained: 21
- Total funding obtained: RM2,556,116
- Ongoing grants: 3
- Completed grants: 18
ONGOING FUNDING
2024-2026
Putra Grant (GPB), UPM – RM150,000
Targeting REST to mitigate neurodevelopmental and synaptic dysfunction in Down syndrome human-induced pluripotent stem cell-derived cerebral organoids. PI: Prof Dr. Michael Ling
2024-2025
Industrial Grant – RM47,000
Developmental and functional evaluation of CEXCI on Down syndrome human iPSC derived neural models. PI: Prof Dr. Michael Ling
2022-2025
Fundamental Research Grant Scheme, MOHE – RM165,036
Investigation of cell-specific REST expression and its repression on JAK-STAT signalling pathway to revert the neurogenic-to-gliogenic shift in Down syndrome cerebral organoids model. PI: AP Dr. Michael Ling
COMPLETED PROJECTS
2022-2024
Putra Grant IPS, UPM – RM25,000
Investigation of lithium effect on REST expression and reactive astrocyte in Down Syndrome. PI: AP Dr. Michael Ling (Postgraduate: Dr Huang Tan)
2022-2024
Putra Grant (GPB), UPM – RM120,000
Targeting JAK-STAT signalling pathway to revert neurogenic-to-gliogenic shift and connective deficits in the brain of Ts1Cje mouse model for Down syndrome. PI: AP Dr. Pike-See Cheah
2021-2024
Fundamental Research Grant Scheme, MOHE – RM157,000
Investigation of lithium-mediated neuroprotection in Down syndrome via REST restoration. PI: AP Dr. Pike-See Cheah
2021-2023
IBRO Return Home Grant – EURO20,000
The interplay between REST and JAK-STAT in the neurogenic-to-gliogenic shift in Down syndrome. PI: AP Dr. Michael KH Ling
2021-2023
ISN Return Home Grant – USD10,000
Profiling of cell-specific REST expression and its repression on JAK-STAT signaling in Down syndrome cerebral organoids. PI: AP Dr. Michael KH Ling
2017-2018
Putra Grant (IPS), UPM – RM20,000
Identification and validation of a novel human orthologue for miR-3099 known as mds-21. PI: Dr. Michael KH Ling (Postgraduate: Mr. Shahidee Zainal Abidin)
2015-2017
Sciencefund, MOSTI – RM293,523
TargetingJAK-STAT signaling pathwayto revertneurogenic-to-gliogenic shift in the brain of Ts1Cje Mouse Model for Down syndrome. PI: Dr. Michael KH Ling
2015-2017
Fundamental Research Grant Scheme, MOHE – RM139,000
Role of the JAK-STAT signaling pathway during neurogenic-to-gliogenic shift in the brain of Ts1Cje Mouse Model for Down syndrome. PI: Dr. Pike-See Cheah
2014-2016
Putra Grant (Interdisciplinary), UPM – RM273,100
Psychosocial risk factors, BDNF and DAT1 Polymorphism and Risk of Major Depressive Disorder. PI: Joint between Prof. Dr. Normala Ibrahim with Dr. Michael KH Ling
2015-2016
Putra Grant (Postgraduate), UPM – RM15,000
Protein and gene expression profile of skeletal muscles in Ts1Cje Mouse Model of Down Syndrome : an insight to muscle weakness. PI: Dr. Pike-See Cheah (Postgraduate: Mr. Usman Bala)
2013-2015
Putra Grant, UPM – RM198,963
Molecular, metabolic and functional characterization of adult skeletal muscle in Down syndrome mouse model : insights into the muscle weakness seen in human condition. PI: Dr. Pike-See Cheah
2012-2015
Putra Grant, UPM – RM176,994
Identification of molecular mechanism responsible for hypotonia in adult Ts1Cje mouse model of Down syndrome. PI: Dr. Michael KH Ling
2013-2015
Putra Grant (Postgraduate), UPM – RM15,000
The association of CDKL5 and STXBP1 gene mutations in paediatric patients with early-onset epileptic encephalopathy in Malaysia. PI: Dr. Michael KH Ling (Postgraduate: Ms Ameerah Jaafar)
2012-2015
ERGS, MOHE – RM119,000
In vivo analysis of a novel miRNA, M1181, during mouse cerebral corticogenesis using in utero electroporation (IUE) approach. PI: Dr. Pike-See Cheah
2012-2015
FRGS, MOHE – RM81,000
A mouse embryonic stem cell culture system with stable and regulatable expression for a novel microRNA, miR3099: An in vitro approach towards functional genomics study. PI: Dr. Michael KH Ling
2013-2015
RUGS (Postgraduate), UPM – RM10,000
Expression profiling of a novel microRNA, miR3099, during neuronal development using an in vitro system. PI: Dr. Michael KH Ling (Postgraduate: Mr. Wei-Hong Siew)
2012-2014
Sciencefund, MOSTI – RM263,500
Establishment of the first ‘superelectroporator’ platform in Malaysia for the delivery of bicistronic expression vectors with shRNA and GFP open reading frames into the cerebral cortex of mouse model for Down syndrome. PI: Dr. Pike-See Cheah
2011-2013
RUGS, UPM – RM168,000
The identification of disrupted molecular networks involved in brain maturation and function in the Ts1Cje mouse model of Down Syndrome. PI: Dr. Pike-See Cheah