Master, Shantou University, Biochemistry and molecular biology (2011)
Silicone Oil-Induced Glaucomatous Neurodegeneration in Rhesus Macaques.
International journal of molecular sciences
2022; 23 (24)
Previously, we developed a simple procedure of intracameral injection of silicone oil (SO) into mouse eyes and established the mouse SOHU (SO-induced ocular hypertension under-detected) glaucoma model with reversible intraocular pressure (IOP) elevation and significant glaucomatous neurodegeneration. Because the anatomy of the non-human primate (NHP) visual system closely resembles that of humans, it is the most likely to predict human responses to diseases and therapies. Here we tried to replicate the mouse SOHU glaucoma model in rhesus macaque monkeys. All six animals that we tested showed significant retinal ganglion cell (RGC) death, optic nerve (ON) degeneration, and visual functional deficits at both 3 and 6 months. In contrast to the mouse SOHU model, however, IOP changed dynamically in these animals, probably due to individual differences in ciliary body tolerance capability. Further optimization of this model is needed to achieve consistent IOP elevation without permanent damage of the ciliary body. The current form of the NHP SOHU model recapitulates the severe degeneration of acute human glaucoma, and is therefore suitable for assessing experimental therapies for neuroprotection and regeneration, and therefore for translating relevant findings into novel and effective treatments for patients with glaucoma and other neurodegenerations.
View details for DOI 10.3390/ijms232415896
View details for PubMedID 36555536
Longitudinal in vivo Ca2+ imaging reveals dynamic activity changes of diseased retinal ganglion cells at the single-cell level.
Proceedings of the National Academy of Sciences of the United States of America
2022; 119 (48): e2206829119
Retinal ganglion cells (RGCs) are heterogeneous projection neurons that convey distinct visual features from the retina to brain. Here, we present a high-throughput in vivo RGC activity assay in response to light stimulation using noninvasive Ca2+ imaging of thousands of RGCs simultaneously in living mice. Population and single-cell analyses of longitudinal RGC Ca2+ imaging reveal distinct functional responses of RGCs and unprecedented individual RGC activity conversions during traumatic and glaucomatous degeneration. This study establishes a foundation for future in vivo RGC function classifications and longitudinal activity evaluations using more advanced imaging techniques and visual stimuli under normal, disease, and neural repair conditions. These analyses can be performed at both the population and single-cell levels using temporal and spatial information, which will be invaluable for understanding RGC pathophysiology and identifying functional biomarkers for diverse optic neuropathies.
View details for DOI 10.1073/pnas.2206829119
View details for PubMedID 36409915
Maprotiline restores ER homeostasis and rescues neurodegeneration via Histamine Receptor H1 inhibition in retinal ganglion cells.
2022; 13 (1): 6796
When the protein or calcium homeostasis of the endoplasmic reticulum (ER) is adversely altered, cells experience ER stress that leads to various diseases including neurodegeneration. Genetic deletion of an ER stress downstream effector, CHOP, significantly protects neuron somata and axons. Here we report that three tricyclic compounds identified through a small-scale high throughput screening using a CHOP promoter-driven luciferase cell-based assay, effectively inhibit ER stress by antagonizing their common target, histamine receptor H1 (HRH1). We further demonstrated that systemic administration of one of these compounds, maprotiline, or CRISPR-mediated retinal ganglion cell (RGC)-specific HRH1 inhibition, delivers considerable neuroprotection of both RGC somata and axons and preservation of visual function in two mouse optic neuropathy models. Finally, we determine that maprotiline restores ER homeostasis by inhibiting HRH1-mediated Ca2+ release from ER. In this work we establish maprotiline as a candidate neuroprotectant and HRH1 as a potential therapeutic target for glaucoma.
View details for DOI 10.1038/s41467-022-34682-y
View details for PubMedID 36357388
Single-cell transcriptome analysis of regenerating RGCs reveals potent glaucoma neural repair genes.
Axon regeneration holds great promise for neural repair of CNS axonopathies, including glaucoma. Pten deletion in retinal ganglion cells (RGCs) promotes potent optic nerve regeneration, but only a small population of Pten-null RGCs are actually regenerating RGCs (regRGCs); most surviving RGCs (surRGCs) remain non-regenerative. Here, we developed a strategy to specifically label and purify regRGCs and surRGCs, respectively, from the same Pten-deletion mice after optic nerve crush, in which they differ only in their regeneration capability. Smart-Seq2 single-cell transcriptome analysis revealed novel regeneration-associated genes that significantly promote axon regeneration. The most potent of these, Anxa2, acts synergistically with its ligand tPA in Pten-deletion-induced axon regeneration. Anxa2, its downstream effector ILK, and Mpp1 dramatically protect RGC somata and axons and preserve visual function in a clinically relevant model of glaucoma, demonstrating the exciting potential of this innovative strategy to identify novel effective neural repair candidates.
View details for DOI 10.1016/j.neuron.2022.06.022
View details for PubMedID 35952672
In Vivo Evaluation of Naive and Diseased RGC Activities at Single-Cell Level
ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2022
View details for Web of Science ID 000844401305210
NMNAT2 and NAD(+) are Downregulated in Glaucomatous RGCs and Overexpression of NMNAT2 Rescues Glaucomatous Neurodegeneration
ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2022
View details for Web of Science ID 000844401303120
Neuroprotection of SARM1 Inhibition in Traumatic and Glaucomatous but not in EAE Optic Neuropathies
ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2022
View details for Web of Science ID 000844401303121
NMNAT2 Is Downregulated in Glaucomatous RGCs and RGC-Specific Gene Therapy Rescues Neurodegeneration and Visual Function.
Molecular therapy : the journal of the American Society of Gene Therapy
The lack of neuroprotective treatments for retinal ganglion cells (RGCs) and optic nerve (ON) is a central challenge for glaucoma management. Emerging evidence suggests that redox factor NAD+ decline is a hallmark of aging and neurodegenerative diseases. Supplementation with NAD+ precursors and overexpression of NMNAT1, the key enzyme in the NAD+ biosynthetic process, have significant neuroprotective effects. We first profile the translatomes of RGCs in naive mice and mice with silicone oil-induced ocular hypertension (SOHU)/glaucoma by RiboTag mRNA sequencing. Intriguingly, only NMNAT2, but not NMNAT1 or NMNAT3, is significantly decreased in SOHU glaucomatous RGCs, which we confirm by in situ hybridization. We next demonstrate that AAV2 intravitreal injection-mediated overexpression of long half-life NMNAT2 mutant driven by RGC-specific mouse gamma-synuclein (mSncg) promoter restores decreased NAD+ levels in glaucomatous RGCs and ONs. Moreover, this RGC-specific gene therapy strategy delivers significant neuroprotection of both RGC soma and axon and preservation of visual function in the traumatic ON crush model and the SOHU glaucoma model. Collectively, our studies suggest that the weakening of NMNAT2 expression in glaucomatous RGCs contributes to a deleterious NAD+ decline and that modulating RGC intrinsic NMNAT2 levels by AAV2-mSncg vector is a promising gene therapy for glaucomatous neurodegeneration.
View details for DOI 10.1016/j.ymthe.2022.01.035
View details for PubMedID 35114390
Osteopontin drives retinal ganglion cell resiliency in glaucomatous neuropathy
ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2021
View details for Web of Science ID 000690761400585
OCRL regulates lysosome positioning and mTORC1 activity through SSX2IP-mediated microtubule anchoring.
Lysosomal positioning and mTOR (mammalian target of rapamycin) signaling coordinate cellular responses to nutrient levels. Inadequate nutrient sensing can result in growth delays, a hallmark of Lowe syndrome. OCRL mutations cause Lowe syndrome, but the role of OCRL in nutrient sensing is unknown. Here, we show that OCRL is localized to the centrosome by its ASH domain and that it recruits microtubule-anchoring factor SSX2IP to the centrosome, which is important in the formation of the microtubule-organizing center. Deficiency of OCRL in human and mouse cells results in loss of microtubule-organizing centers and impaired microtubule-based lysosome movement, which in turn leads to mTORC1 inactivation and abnormal nutrient sensing. Centrosome-targeted PACT-SSX2IP can restore microtubule anchoring and mTOR activity. Importantly, boosting the activity of mTORC1 restores the nutrient sensing ability of Lowe patients' cells. Our findings highlight mTORC1 as a novel therapeutic target for Lowe syndrome.
View details for DOI 10.15252/embr.202052173
View details for PubMedID 33987909
Chronic mild and acute severe glaucomatous neurodegeneration derived from silicone oil-induced ocular hypertension.
2021; 11 (1): 9052
Recently, we established silicone oil-induced ocular hypertension (SOHU) mouse model with significant glaucomatous neurodegeneration. Here we characterize two additional variations of this model that simulate two distinct glaucoma types. The first is a chronic model produced by high frequency (HF) pupillary dilation after SO-induced pupillary block, which shows sustained moderate IOP elevation and corresponding slow, mild glaucomatous neurodegeneration. We also demonstrate that although SO removal quickly returns IOP to normal, the glaucomatous neurodegeneration continues to advance to a similar degree as in the HF group without SO removal. The second, an acute model created by no pupillary dilation (ND), shows a greatly elevated IOP and severe inner retina degeneration at an early time point. Therefore, by a straightforward dilation scheme, we extend our original SOHU model to recapitulate phenotypes of two major glaucoma forms, which will be invaluable for selecting neuroprotectants and elucidating their molecular mechanisms.
View details for DOI 10.1038/s41598-021-88690-x
View details for PubMedID 33907301
Delineating the organization of projection neuron subsets in primary visual cortex with multiple fluorescent rabies virus tracing.
Brain structure & function
The impressive functions of the brain rely on an extensive connectivity matrix between specific neurons, the architecture of which is frequently characterized by one brain nucleus/region connecting to multiple targets, either via collaterals of the same projection neuron or several, differentially specified neurons. Delineating the fine architecture of projection neuron subsets in a specific brain region could greatly facilitate its circuit, computational, and functional resolution. Here, we developed multiple fluorescent rabies viruses (RV) to delineate the fine organization of corticothalamic projection neuron subsets in the primary visual cortex (V1). By simultaneously retrograde labeling multiple distinct subsets of corticothalamic projection neurons in V1 from their target nuclei in thalamus (dLGN, LP, LD), we observed that V1-dLGN corticothalamic projection neurons were densely concentrated in layer VI, except for several sparsely scattered neurons in layer V, while V1-LP and V1-LD corticothalamic projection neurons were localized to both layers V and VI. Meanwhile, we observed a fraction of V1 corticothalamic projection neurons targeting two thalamic nuclei, which was further confirmed by fMOST whole-brain imaging. The multiple fluorescent RV tracing tools can be extensively applied to resolve the architecture of projection neuron subsets in certain brain regions, with a strong potential to delineate the computational and functional organization of these brain regions.
View details for DOI 10.1007/s00429-021-02250-7
View details for PubMedID 33710409
Post-translational modification of Sox11 regulates RGC survival and axon regeneration.
The failure of adult CNS neurons to survive and regenerate their axons after injury or in neurodegenerative disease remains a major target for basic and clinical neuroscience. Recent data demonstrated in the adult mouse that exogenous expression of Sry-related high-mobility-box 11 (Sox11) promotes optic nerve regeneration after optic nerve injury, but exacerbates the death of a subset of retinal ganglion cells, alpha-RGCs. During development, Sox11 is required for RGC differentiation from retinal progenitor cells (RPCs), and we found that mutation of a single residue to prevent sumoylation at lysine 91 (K91) increased nuclear localization and RGC differentiation in vitro Here we explored whether this Sox11 manipulation similarly has stronger effects on RGC survival and optic nerve regeneration. In vitro, we found that non-SUMOylatable Sox11K91A leads to RGC death and suppresses axon outgrowth in primary neurons. We furthermore found that Sox11K91A more strongly promotes axon regeneration but also increases RGC death after optic nerve injury in vivo in adult mouse. RNA sequence data showed that Sox11 and Sox11K91A increase the expression of key signaling pathway genes associated with axon growth and regeneration but downregulated Spp1 and Opn4 expression in RGC cultures, consistent with negatively regulating the survival of alpha-RGCs and ipRGCs. Thus Sox11 and its sumoylation site at K91 regulate gene expression, survival and axon growth in RGCs and may be explored further as potential regenerative therapies for optic neuropathy.Significance Statement Sox11 expression promotes optic nerve regeneration but also increases RGC death after optic nerve injury. Here we demonstrate that mutation of a single SUMOylation site on Sox11 (Sox11K91A) leads to stronger effects in vivo RNA sequencing analysis reveals that Sox11 and Sox11K91A differentially regulate downstream gene expression related to axon growth and guidance. Understanding these effects of post-translational modification of Sox11 in regulating regeneration in vivo suggests a potent therapeutic strategy for vision restoration in optic neuropathies.
View details for DOI 10.1523/ENEURO.0358-20.2020
View details for PubMedID 33441400
Neuronal NMNAT2 Overexpression Does Not Achieve Significant Neuroprotection in Experimental Autoimmune Encephalomyelitis/Optic Neuritis.
Frontiers in cellular neuroscience
2021; 15: 754651
Optic neuritis, inflammation, and demyelination of the optic nerve (ON), is one of the most common clinical manifestations of multiple sclerosis; affected patients suffer persistent visual symptoms due to ON degeneration and secondary retinal ganglion cell (RGC) death. The mouse experimental autoimmune encephalomyelitis (EAE) model replicates optic neuritis and significant RGC soma and axon loss. Nicotinamide mononucleotide adenylyltransferases (NMNATs) are NAD+-synthetic enzymes that have been shown to be essential for axon integrity, activation of which significantly delays axonal Wallerian degeneration. NMNAT2, which is enriched in axons, has been proposed as a promising therapeutic target for axon injury-induced neurodegeneration. We therefore investigated whether activation of NMNAT2 can be used as a gene therapy strategy for neuroprotection in EAE/optic neuritis. To avoid the confounding effects in inflammatory cells, which play important roles in EAE initiation and progression, we used an RGC-specific promoter to drive the expression of the long half-life NMNAT2 mutant in mouse RGCs in vivo. However, optical coherence tomography in vivo retina imaging did not reveal significant protection of the ganglion cell complex, and visual function assays, pattern electroretinography, and optokinetic response also showed no improvement in mice with NMNAT2 overexpression. Postmortem histological analysis of retina wholemounts and semithin sections of ON confirmed the in vivo results: NMNAT2 activation in RGCs does not provide significant neuroprotection of RGCs in EAE/optic neuritis. Our studies suggest that a different degenerative mechanism than Wallerian degeneration is involved in autoimmune inflammatory axonopathy and that NMNAT2 may not be a major contributor to this mechanism.
View details for DOI 10.3389/fncel.2021.754651
View details for PubMedID 34707482
Longitudinal Morphological and Functional Assessment of RGC Neurodegeneration After Optic Nerve Crush in Mouse.
Frontiers in cellular neuroscience
2020; 14: 109
The mouse optic nerve crush (ONC) model has been widely used to study optic neuropathies and central nervous system (CNS) axon injury and repair. Previous histological studies of retinal ganglion cell (RGC) somata in retina and axons in ON demonstrate significant neurodegeneration after ONC, but longitudinal morphological and functional assessment of RGCs in living animals is lacking. It is essential to establish these assays to provide more clinically relevant information for early detection and monitoring the progression of CNS neurodegeneration. Here, we present in vivo data gathered by scanning laser ophthalmoscopy (SLO), optical coherence tomography (OCT), and pattern electroretinogram (PERG) at different time points after ONC in mouse eyes and corresponding histological quantification of the RGC somata and axons. Not surprisingly, direct visualization of RGCs by SLO fundus imaging correlated best with histological quantification of RGC somata and axons. Unexpectedly, OCT did not detect obvious retinal thinning until late time points (14 and 28-days post ONC) and instead detected significant retinal swelling at early time points (1-5 days post-ONC), indicating a characteristic initial retinal response to ON injury. PERG also demonstrated an early RGC functional deficit in response to ONC, before significant RGC death, suggesting that it is highly sensitive to ONC. However, the limited progression of PERG deficits diminished its usefulness as a reliable indicator of RGC degeneration.
View details for DOI 10.3389/fncel.2020.00109
View details for PubMedID 32410964
View details for PubMedCentralID PMC7200994
Mouse gamma-Synuclein Promoter-Mediated Gene Expression and Editing in Mammalian Retinal Ganglion Cells.
The Journal of neuroscience : the official journal of the Society for Neuroscience
Optic neuropathies are a group of optic nerve (ON) diseases caused by various insults including glaucoma, inflammation, ischemia, trauma and genetic deficits, which are characterized by retinal ganglion cell (RGC) death and ON degeneration. An increasing number of genes involved in RGC intrinsic signaling have been found to be promising neural repair targets that can potentially be modulated directly by gene therapy, if we can achieve RGC specific gene targeting. To address this challenge, we first used adeno associated virus (AAV)-mediated gene transfer to perform a low throughput in vivo screening in both male and female mouse eyes and identified the mouse γ-synuclein (mSncg) promoter, which specifically and potently sustained transgene expression in mouse RGCs and also works in human RGCs. We further demonstrated that gene therapy that combines AAV-mSncg promoter with CRISPR/Cas9 gene editing can knockdown pro-degenerative genes in RGCs and provide effective neuroprotection in optic neuropathies.Significance Statement:Here we present an RGC-specific promoter, mouse γ-synuclein (mSncg) promoter, and perform extensive characterization and proof-of-concept studies of mSncg promoter-mediated gene expression and CRISPR/Cas9 gene editing in RGCs in vivo To our knowledge, this is the first report demonstrating in vivo neuroprotection of injured RGCs and optic nerve by AAV-mediated CRISPR/Cas9 inhibition of genes that are critical for neurodegeneration. It represents a powerful tool to achieve RGC-specific gene modulation, and also opens up a promising gene therapy strategy for optic neuropathies, the most common form of eye diseases that cause irreversible blindness.
View details for DOI 10.1523/JNEUROSCI.0102-20.2020
View details for PubMedID 32300046
In Vivo Two-photon Calcium Imaging in Dendrites of Rabies Virus-labeled V1 Corticothalamic Neurons
Monitoring neuronal activity in vivo is critical to understanding the physiological or pathological functions of the brain. Two-photon Ca2+ imaging in vivo using a cranial window and specific neuronal labeling enables real-time, in situ, and long-term imaging of the living brain. Here, we constructed a recombinant rabies virus containing the Ca2+ indicator GCaMP6s along with the fluorescent protein DsRed2 as a baseline reference to ensure GCaMP6s signal reliability. This functional tracer was applied to retrogradely label specific V1-thalamus circuits and detect spontaneous Ca2+ activity in the dendrites of V1 corticothalamic neurons by in vivo two-photon Ca2+ imaging. Notably, we were able to record single-spine spontaneous Ca2+ activity in specific circuits. Distinct spontaneous Ca2+ dynamics in dendrites of V1 corticothalamic neurons were found for different V1-thalamus circuits. Our method can be applied to monitor Ca2+ dynamics in specific input circuits in vivo, and contribute to functional studies of defined neural circuits and the dissection of functional circuit connections.
View details for DOI 10.1007/s12264-019-00452-y
View details for Web of Science ID 000500858000002
View details for PubMedID 31808041
A Reversible Silicon Oil-Induced Ocular Hypertension Model in Mice.
Journal of visualized experiments : JoVE
Elevated intraocular pressure (IOP) is a well-documented risk factor for glaucoma. Here we describe a novel, effective method for consistently inducing stable IOP elevation in mice that mimics the post-operative complication of using silicone oil (SO) as a tamponade agent in human vitreoretinal surgery. In this protocol, SO is injected into the anterior chamber of the mouse eye to block the pupil and prevent inflow of aqueous humor. The posterior chamber accumulates aqueous humor and this in turn increases the IOP of the posterior segment. A single SO injection produces reliable, sufficient, and stable IOP elevation, which induces significant glaucomatous neurodegeneration. This model is a true replicate of secondary glaucoma in the eye clinic. To further mimic the clinical setting, SO can be removed from the anterior chamber to reopen the drainage pathway and allow inflow of aqueous humor, which is drained through the trabecular meshwork (TM) at the angle of the anterior chamber. Because IOP quickly returns to normal, the model can be used to test the effect of lowering IOP on glaucomatous retinal ganglion cells. This method is straightforward, does not require special equipment or repeat procedures, closely simulates clinical situations, and may be applicable to diverse animal species. However, minor modifications may be required.
View details for DOI 10.3791/60409
View details for PubMedID 31789319
A novel inducible and reversible mouse glaucoma model: Silicone Oil-Induced Ocular Hypertension Under-detected (SOHU)
ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488628101260
Neuroprotection of Retinal Ganglion Cells by AAV2-gamma-Synuclein Promoter-Mediated CRISPR/Cas9 Gene Editing
ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488628105194
Stem cell-derived retinal ganglion cell differentiation and its transplantation
ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488628108093
Loss of optineurin C-terminus causes significant retinal ganglion cell degeneration
ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488800702030
Memantine can relieve the neuronal impairment caused by neurotropic virus infection
JOURNAL OF MEDICAL VIROLOGY
2019; 91 (6): 935–40
View details for DOI 10.1002/jmv.25396
View details for Web of Science ID 000465087100005
Silicone oil-induced ocular hypertension and glaucomatous neurodegeneration in mouse
View details for DOI 10.7554/eLife.45881
View details for Web of Science ID 000468863200001
The anatomy and metabolome of the lymphatic system in the brain in health and disease.
Brain pathology (Zurich, Switzerland)
Recent studies have demonstrated that the brain is equipped with a lymphatic drainage system that is actively involved in parenchymal waste clearance, brain homeostasis and immune regulation. However, the exact anatomic drainage routes of brain lymph fluid (BLF) remain elusive, hampering the physiological study and clinical application of this system. In this study, we systematically dissected the anatomy of the BLF pathways in a rat model. Moreover, we developed a protocol to collect BLF from the afferent lymphatic vessels of deep cervical lymph nodes (dcLNs) and cerebrospinal fluid (CSF) from the fourth ventricle. Nuclear magnetic resonance spectroscopy showed that BLF contains more metabolites than CSF, suggesting that BLF might be a more sensitive indicator of brain dynamics under physiological and pathological conditions. Finally, we identified several metabolites as potential diagnostic biomarkers for glioma, Parkinson's disease and CNS infectious diseases. Together, these data may provide insight into the physiology of the lymphatic system in the brain and into the clinical diagnosis of CNS disorders.
View details for DOI 10.1111/bpa.12805
View details for PubMedID 31747475
Digestion-ligation-only Hi-C is an efficient and cost-effective method for chromosome conformation capture
2018; 50 (5): 754-+
Chromosome conformation capture (3C) technologies can be used to investigate 3D genomic structures. However, high background noise, high costs, and a lack of straightforward noise evaluation in current methods impede the advancement of 3D genomic research. Here we developed a simple digestion-ligation-only Hi-C (DLO Hi-C) technology to explore the 3D landscape of the genome. This method requires only two rounds of digestion and ligation, without the need for biotin labeling and pulldown. Non-ligated DNA was efficiently removed in a cost-effective step by purifying specific linker-ligated DNA fragments. Notably, random ligation could be quickly evaluated in an early quality-control step before sequencing. Moreover, an in situ version of DLO Hi-C using a four-cutter restriction enzyme has been developed. We applied DLO Hi-C to delineate the genomic architecture of THP-1 and K562 cells and uncovered chromosomal translocations. This technology may facilitate investigation of genomic organization, gene regulation, and (meta)genome assembly.
View details for DOI 10.1038/s41588-018-0111-2
View details for Web of Science ID 000431394900019
View details for PubMedID 29700467
In vivo fiber photometry of neural activity in response to optogenetically manipulated inputs in freely moving mice
JOURNAL OF INNOVATIVE OPTICAL HEALTH SCIENCES
2017; 10 (5)
View details for DOI 10.1112/S1793515817130015
View details for Web of Science ID 000412481200006
Rabies viruses leader RNA interacts with host Hsc70 and inhibits virus replication
2017; 8 (27): 43822–37
Viruses have been shown to be equipped with regulatory RNAs to evade host defense system. It has long been known that rabies virus (RABV) transcribes a small regulatory RNA, leader RNA (leRNA), which mediates the transition from viral RNA transcription to replication. However, the detailed molecular mechanism remains enigmatic. In the present study, we determined the genetic architecture of RABV leRNA and demonstrated its inhibitory effect on replication of wild-type rabies, DRV-AH08. The RNA immunoprecipitation results suggest that leRNA inhibits RABV replication via interfering the binding of RABV nucleoprotein with genomic RNA. Furthermore, we identified heat shock cognate 70 kDa protein (Hsc70) as a leRNA host cellular interacting protein, of which the expression level was dynamically regulated by RABV infection. Notably, our data suggest that Hsc70 was involved in suppressing RABV replication by leader RNA. Finally, our experiments imply that leRNA might be potentially useful as a novel drug in rabies post-exposure prophylaxis. Together, this study suggested leRNA in concert with its host interacting protein Hsc70, dynamically down-regulate RABV replication.
View details for DOI 10.18632/oncotarget.16517
View details for Web of Science ID 000405498000019
View details for PubMedID 28388579
View details for PubMedCentralID PMC5546443
Induction of HepG2 cell apoptosis by Irgarol 1051 through mitochondrial dysfunction and oxidative stresses
TOXICOLOGY IN VITRO
2013; 27 (6): 1771–79
In this study, HepG2 cells were exposed to 0.04-40 mg/L Irgarol 1051. Results show that Irgarol 1051 can damage cell morphology and cause a significant decrease in cell viability. Positive staining by Annexin V, caspase-3 activity enhancement, and the damage in cell ultrastructure indicated an apoptotic mode of cell death for 4.0mg/L Irgarol 1051 treatment. At the same time, caspase-9 was also significantly induced by 0.4 and 4.0mg/L Irgarol 1051 at 72 h, which suggests that the intrinsic mitochondria pathway was involved in the apoptosis. The mitochondrial membrane potential decreased significantly after the HepG2 cells were exposed to Irgarol 1051 for 6 and 72 h. Especially, the translocation of cytochrome c from mitochondria to cytosol was recorded, supporting the idea that the mitochondrial pathway was involved in the apoptosis signal pathways induced by Irgarol 1051. The significantly increased levels of intracellular reactive oxygen species (ROS) and an immediate ROS burst were also recorded. The results here may imply that Irgarol 1051 induces HepG2 cell apoptosis through mitochondrial dysfunction and oxidative stresses. Although it is possible that this chemical has no detrimental effects on human health at the environmentally relevant concentration, it may cause problems to top coastal predators due to bio-accumulation through the food chain.
View details for DOI 10.1016/j.tiv.2013.05.006
View details for Web of Science ID 000324847800021
View details for PubMedID 23722069
Enhanced proliferation and differentiation of neural stem cells grown on PHA films coated with recombinant fusion proteins
2013; 9 (8): 7845–54
Polyhydroxyalkanoates (PHAs) belong to a family of copolyesters with demonstrated biocompatibility. We hypothesize that genetically fusing evolutionarily preserved cell binding motifs, such as RGD or IKVAV, to the PHA-binding protein phasin (PhaP) for surface functionalization of PHA materials could better support the growth and differentiation of neural stem cells (NSCs). This hypothesis is tested on three polyester materials of the same aliphatic family: poly(L-lactic acid) (PLA) and two PHB copolymers, poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) and poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-3-hydroxyhexanoate) (PHBVHHx). Experimental results indicate that surface coating of the two fusion proteins, PhaP-RGD and PhaP-IKVAV, provides short-term advantages in promoting the adhesion, proliferation and neural differentiation of rat NSCs compared to the PhaP-coated or uncoated material. Among the tested samples, the combination of coating PhaP-IKVAV on an PHBVHHx surface yields the highest levels in cell adhesion and proliferation, while the PLA film coated with PhaP-IKVAV promotes better neural differentiation and neurite outgrowth in the early stage. Because both PhaP-RGD and PhaP-IKVAV could be produced in an inexpensive manner, our data suggest that PhaP-IKVAV is an ideal nonspecific coating agent to functionalize hydrophobic biomaterials in the application of neural tissue engineering.
View details for DOI 10.1016/j.actbio.2013.04.038
View details for Web of Science ID 000322207700025
View details for PubMedID 23639778