February 26

Hot Off the Press: Andrographolide attenuates LPS-stimulated up-regulation of C-C and C-X-C motif chemokines in rodent cortex and primary astrocytes

Andro chemokine brain


Andrographolide is the major bioactive compound isolated from Andrographis paniculata, a native South Asian herb used medicinally for its anti-inflammatory properties. In this study, we aimed to assess andrographolide’s potential utility as an anti-neuroinflammatory therapeutic. Therefore, we conducted an invitro study where the effects of andrographolide on lipopolysaccharide (LPS)-Andro chemokine schematicinduced chemokine up-regulation both in mouse cortex and in cultured primary astrocytes were measured, including cytokine profiling, gene expression, and, in cultured astrocytes, activation of putative signaling regulators. We found that orally administered andrographolide significantly attenuated mouse cortical chemokine levels from the C-C and C-X-C subfamilies. Similarly, andrographolide abrogated a range of LPS-induced chemokines as well as tumor necrosis factor (TNF)-α in astrocytes. In astrocytes, the inhibitory actions of andrographolide on chemokine and TNF-α up-regulation appeared to be mediated by nuclear factor-κB (NF-κB) or c-Jun N-terminal kinase (JNK) activation. These results suggest that andrographolide may be useful as a therapeutic for neuroinflammatory diseases, especially those characterized by chemokine dysregulation.


Wong SY, Tan MG, Banks WA, Wong WS, Wong PT, Lai MK (This paper).

February 26

Hot Off the Press: An isoform-specific role of FynT tyrosine kinase in Alzheimer’s disease

Fyn isoform AD

Recent studies suggest that Fyn tyrosine kinase forms part of a toxic triad with β-amyloid and tau in the disease process. However, it is not known whether Fyn is associated with the pathological features of AD in an isoform-specific manner. In this study, we identified selective up-regulation of the alternative-spliced FynT isoform with no change in FynB in the AD neocortex. Furthermore, gene ontology term enrichment analyses and cell type-specific localization of FynT immunoreactivity suggest that FynT up-regulation was associated with neurofibrillary degeneration and reactive astrogliosis. Interestingly, significantly increased FynT in NFT-bearing neurons was concomitant to decreased FynB immunoreactivity, suggesting an involvement of alternative splicing in NFT formation. Furthermore, cultured cells of astrocytic origin have higher FynT to FynB ratio compared to those of neuronal origin. Lastly, primary rat mixed neuron-astrocyte cultures treated with Aβ25-35 showed selective up-regulation of FynT expression in activated astrocytes. Our findings point to an isoform-specific role of FynT in modulating neurofibrillary degeneration and reactive astrogliosis in AD. Fyn kinase is known to interact with β-amyloid and tau, and contributes to Alzheimer’s disease pathogenesis. In this study, it is shown that the alternatively spliced FynT isoform is specifically up-regulated in the AD neocortex, with no change in FynB isoform. The increased FynT correlated with markers of neurofibrillary degeneration and reactive astrogliosis. In primary mixed cultures, treatment with amyloid peptides specifically up-regulated FynT in activated astrocytes. This study points to altered alternative splicing as a potential pathogenic mechanism in AD.

Fyn pTau


Lee C, Low CY, Francis PT, Attems J, Wong PT, Lai MK, Tan MG (This paper).

February 25

Hot Off the Press: Altered relaxin family receptors RXFP1 and RXFP3 in the neocortex of depressed Alzheimer’s disease patients


The G-protein-coupled relaxin family receptors RXFP1 and RXFP3 are widely expressed in the cortex and are involved in stress responses and memory and emotional processing. However, the identification of these receptors in human cortex and their status in Alzheimer’s disease (AD), which is characterized by both cognitive impairments and neuropsychiatric behaviours, have not been reported. In this study, we characterized RXFP receptors for immunoblotting and measured RXFP1 and RXFP3 immunoreactivities in the postmortem neocortex of AD patients longitudinally assessed for depressive symptoms. RXFP1 and RXFP3 antibodies were characterized by immunoblotting with lysates from transfected HEK cells and preadsorption with RXFP3 peptides. Also, postmortem neocortical tissues from behaviourally assessed AD and age-matched controls were processed for immunoblotting with RXFP1 and RXFP3 antibodies. Compared to controls, putative RXFP1 immunoreactivity was reduced in parietal cortex of non-depressed AD patients but unchanged in depressed patients. Furthermore, RXFP3 immunoreactivity was increased only in depressed AD patients. RXFP1 levels in the parietal cortex also correlated with severity of depression symptoms. In contrast, RXFP1 and RXFP3 levels did not correlate with dementia severity or β-amyloid burden. Our study suggests that alterations of RXFP1 and RXFP3 may be neurochemical markers of depression in AD, and relaxin family receptors warrant further preclinical investigations as possible therapeutic targets for neuropsychiatric symptoms in dementia.


Lee JH, Koh SQ, Guadagna S, Francis PT, Esiri MM, Chen CP, Wong PT, Dawe GS, Lai MK (This paper).

October 6

Hot Off the Press: Extracellular vesicles are rapidly purified from human plasma by PRotein Organic Solvent PRecipitation (PROSPR)


Extracellular vesicles (EVs) such as exosomes and microvesicles mediate intercellular communication and regulate a diverse range of crucial biological processes. Host cells that are damaged, infected or transformed release biomarker-containing EVs into the peripheral circulation, where they can be readily accessed for use in diagnostic or prognostic testing. However, current methods of EV isolation from blood plasma are complex and often require relatively large sample volumes, hence are inefficient for widespread use in clinical settings. In work led by collaborator Newman Sze, we report a novel and inexpensive method of rapidly isolating EVs from small volumes of human blood plasma by PRotein Organic Solvent PRecipitation (PROSPR). PROSPR encompasses a rapid three-step protocol to remove soluble proteins from plasma via precipitation in cold acetone, leaving the lipid-encapsulated EVs behind in suspension. This generates higher purity EVs that can then be obtained from filtration or classical ultracentrifugation methods. We foresee that PROSPR-based purification of EVs will significantly accelerate the discovery of new disease biomarkers and the characterization of EVs with potential for clinical applications.


Gallart-Palau X, Serra A, Wong AS, Sandin S, Lai MK, Chen CP, Kon OL, Sze SK (This paper).

May 19

Hot Off the Press: Cystathionine β-Synthase Inhibition Is a Potential Therapeutic Approach to Treatment of Ischemic Injury

CBS ASNneuro

Cystathionine β-synthase (CBS) is the central nervous system’s major producer of hydrogen sulfide (H2S), which has been reported to exacerbate stroke outcome in experimental models. This study led by Chan Su Jing and Peter Wong showed that when SH-SY5Y cells were transfected to overexpress CBS, the cells were able to synthesize H2S when exposed to high levels of enzyme substrates but not substrate concentrations that may reflect normal physiological conditions. At the same time, these cells demonstrated exacerbated cell death when subjected to oxygen and glucose deprivation (OGD) together with high substrate concentrations, indicating that H2S production has a detrimental effect on cell survival. This effect could be abolished by CBS inhibition. The same effect was observed with primary astrocytes exposed to OGD and high substrates or sodium hydrosulfide. In addition, CBS was upregulated and activated by truncation in primary astrocytes subjected to OGD. When rats were subjected to permanent middle cerebral artery occlusion, CBS activation was also observed. These results imply that in acute ischemic conditions, CBS is upregulated and activated by truncation causing an increased production of H2S, which exacerbate the ischemic injuries. Therefore, CBS inhibition may be a viable approach to stroke treatment.


Chan SJ, Chai C, Lim TW, Yamamoto M, Lo EH, Lai MK, Wong PT (This paper).

September 29

Hot Off the Press: Differential Alterations of Neocortical GluN Receptor Subunits in Patients with Mixed Subcortical Ischemic Vascular Dementia and Alzheimer’s Disease

Ezan GluN VaD MixD Fig1Glutamatergic deficits are well-established neurochemical findings in Alzheimer’s disease (AD) and are thought to underlie both cognitive and behavioral symptoms of the disease. However, it is unclear whether subcortical ischemic vascular dementia (SIVD) and mixed SIVD/AD (MixD) manifest similar changes in the glutamatergic system. In this study, we measured the immunoreactivities of NMDA receptor GluN1, GluN2A, and GluN2B subunits in SIVD and MixD in postmortem neocortical tissues from a cohort of well-characterized, longitudinally followed-up patients with SIVD and MixD, together with age-matched controls. We found a significant reduction of GluN1 only in MixD, while significant increases of GluN2A and GluN2B were found only in SIVD. Furthermore, GluN1 loss and GluN2A/2B upregulation was associated respectively with higher Braak stages and lacunar infarct scores. Our data therefore suggest that the differential alterations of GluN subunits in SIVD and MixD may result from separate, interacting disease processes, and point to the potential utility of glutamatergic approaches for pharmacotherapy.


Mohamed NE, Lee JH, Francis PT, Esiri MM, Chen CP, Lai MK (This paper).

August 22

Hot Off the Press: Autoantibodies to GM1 and GQ1bα are not Biological Markers of Alzheimer’s Disease

Uncovering novel biomarkers which lead to better diagnosis, prognosis and treatment is undoubtedly an important endeavor, but sometimes, finding what is not a biomarker may be worthwhile as well. Gangliosides are complex ceramide-glycosphingolipids known to play important roles in neuronal development. The GM1 ganglioside binds to β-amyloid and prevents β-sheet conformational changes (Yanagisawa et al., 1995), while GQ1bα and GT1aα are cholinergic neuron-specific gangliosides found to be increased in murine AD models (Ariga et al. 2010; Ariga et al. 2013a). Antibodies to gangliosides have been detected in AD, but the differences compared to controls were too low to be useful for diagnosis (Chapman et al. 1988). Other studies suffered from small patient numbers (n = 4, Hatzinfilippou et al. 2008), or low cut-off values resulting in low specificity (Ariga et al. 2013b).

autoantibodies AD F1

In this study led by Prof. Nobuhiro Yuki, we measured serum levels of anti-ganglioside antibodies in patients with clinically diagnosed AD (n = 22) and vascular dementia (n = 14) and compared them to sera of patients with Guillain-Barré syndrome (GBS) and multifocal motor neuropathy (MMN), diseases where the antibodies are thought to be pathogenic; as well as to normal controls. We found that titers of IgG and IgM anti-GM1, anti-GQ1bα, and anti-GT1aα antibodies did not differ among AD, vascular dementia, and normal controls, and were remarkably lower than those in GBS and MMN. Our results suggest that the anti-ganglioside antibodies are unlikely to be pathogenic in AD or useful as biomarkers.


Miura Y, Miyaji K, Chai YL, Chen CL, Lai MK, Yuki N (This paper).

Ariga T, Yanagisawa M, Wakade C, Ando S, Buccafusco JJ, McDonald MP, Yu RK (2010) Ganglioside metabolism in a transgenic mouse model of Alzheimer’s disease: Expression of Chol-1α antigens in the brain. ASN Neuro 2: e00044.

Ariga T, ItokazuY, McDonald MP, Hirabayashi Y, Ando S, Yu RK (2013a) Brain gangliosides of a transgenic mouse model of Alzheimer’s disease with deficiency in GD3-synthase: Expression of elevated levels of a cholinergic-specific ganglioside, GT1aα. ASN Neuro 5: 141-148.

Ariga T, Kubota M, Nakane M, Oguro K, Yu RK, Ando S (2013b) Anti-Chol-1 antigen, GQ1bα, antibodies are associated with Alzheimer’s disease. PLoS One 8: e63326.

Chapman J, Sela BA, Wertman E, Michaelson DM (1988) Antibodies to ganglioside GM1 in patients with Alzheimer’s disease. Neurosci Lett 86: 235-240.

Hatzifilippou E, Koutsouraki E, Banaki T, Traka M, Costa VG, Baloyannis SJ (2008) Antibodies against GM1 in demented patients. Am J Alzheimers Dis Other Demen 23: 274-279.

Yanagisawa K, Odaka A, Suzuki N, Ihara Y (1995) GM1 ganglioside-bound amyloid beta-protein (Aβ): A possible form of preamyloid in Alzheimer’s disease. Nat Med 1: 1062-1066.

June 24

Hot Off the Press: Andrographolide attenuates interleukin-1β-stimulated upregulation of chemokine CCL5 and glial fibrillary acidic protein in astrocytes

NR-D-14-10829R1 Fig 1Andrographolide is a bioactive molecule isolated from Andrographis paniculata with anticancer and anti-inflammatory activities. In this study, we tested the effects of andrographolide on astrocyte-mediated neuroinflammatory responses. Cultured rat primary astrocytes were treated with proinflammatory cytokine interleukin 1β with or without pretreatment with andrographolide, and then processed for measurements of chemokine C-C motif ligand 5 (CCL5) and glial fibrillary acidic protein. The activation status of nuclear factor-κB activation that may underlie CCL5 upregulation was also measured. Andrographolide pretreatment was found to attenuate the upregulation of CCL5 and glial fibrillary basic protein as well as reduce the phosphorylation of nuclear factor-κB p65 and IκBα after interleukin 1β stimulation. These data suggest that andrographolide should be evaluated further as a therapeutic for central nervous system diseases characterized by astrocyte-mediated neuroinflammatory processes. NR-D-14-10829R1 Fig 2


Wong SY, Chan SJ, Wong WS, Wong PT, Lai MK (This paper).

May 6

Hot Off the Press: The brain lipidomes of subcortical ischemic vascular dementia and mixed dementia

VaD MIX lipidome

 Despite its importance as the leading cause of vascular dementia in Asia (Chen 2004), the primary pathogenic mechanisms in subcortical ischemic vascular dementia (SIVD) have remained elusive. Because of the lack of approved therapeutic agents for SIVD, there is a pressing need to identify novel therapeutic targets. In work led by collaborators Guanghou Shui and Markus Wenk, we performed comparative lipidomic analyses of SIVD and mixed dementia (i.e., SIVD and Alzheimer’s disease, MixD) which may confer new insights pertaining to the possible interaction between neurodegenerative and vascular mechanisms in the pathogenesis of dementia. Liquid chromatography coupled to mass spectrometry was used to comprehensively analyze the lipidomes of white and gray matter from the temporal cortex of nondemented controls, SIVD, and MixD subjects. Detailed molecular profiles highlighted the pathologic relevance of gray matter sphingolipid fatty acyl chain heterogeneity in dementia. In addition, the levels of sulfatides and lysobisphosphatidic acids were progressively increased in the temporal cortex gray matter from control to SIVD to MixD. White matter phospholipid profiles indicated possible adaptive mechanisms (i.e., increased unsaturation) to chronic ischemia in SIVD and elevated membrane degradation in MixD.


Chen CP (2004) Transcultural expression of subcortical vascular disease. J Neurol Sci 226:45-47.

Lam SM, Wang Y, Duan X, Wenk MR, Kalaria RN, Chen CP, Lai MK, Shui G (This paper).

February 11

Hot Off the Press: Novel pathophysiological markers are revealed by iTRAQ-based quantitative clinical proteomics approach in vascular dementia

In research led by collaborator Newman Sze, we used an iTRAQ-2D-LC-MS/MS strategy for quantitative analysis of pooled lysates from the neocortex of 21 pathologically confirmed cases of vascular dementia (VaD) and matched non-neurological controls. A total of 144 differentially perturbed proteins out of 2284 confidently identified proteins (false discovery rate=0.3%) were shortlisted for bioinformatics analysis. Western blot analysis of selected proteins using samples from individual patients (n=10 per group) showed significant increases in the abundance of SOD1 and NCAM and reduced ATP5A in VaD. This suggests a state of hypometabolism and vascular insufficiency along with an inflammatory condition during VaD. Elevation of SOD1 and increasing trend for iron-storage proteins (FTL, FTH1) may be indicative of an oxidative imbalance that is accompanied by an aberrant iron metabolism. The synaptic proteins did not exhibit a generalized decrease in abundance (e.g. syntaxin) in the VaD subjects. This reported proteome offers a reference data set for future basic or translational studies on VaD.

VaD Proteomics PNG


Datta A, Qian J, Chong R, Kalaria RN, Francis P, Lai MK, Chen CP, Sze SK. (This paper).