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The molecular link between amyloid-β plaques and neurofibrillary tangles, the two pathological hallmarks of Alzheimer’s disease, is still unclear. Increasing evidence suggests that amyloid-β peptide activates multiple regulators of cell cycle pathways, including transcription factors CDKs and E2F1, leading to hyperphosphorylation of tau protein. However, the exact pathways downstream of amyloid-β-induced cell cycle imbalance are unknown. Here, we show that PAX6, a transcription factor essential for eye and brain development which is quiescent in adults, is increased in the brains of patients with Alzheimer’s disease and in APP transgenic mice, and plays a key role between amyloid-β and tau hyperphosphorylation. Downregulation of PAX6 protects against amyloid-β peptide-induced neuronal death, suggesting that PAX6 is a key executor of the amyloid-β toxicity pathway. Mechanistically, amyloid-β upregulates E2F1, followed by the induction of PAX6 and c-Myb, while Pax6 is a direct target for both E2F1 and its downstream target c-Myb. Furthermore, PAX6 directly regulates transcription of GSK-3β, a kinase involved in tau hyperphosphorylation and neurofibrillary tangles formation, and its phosphorylation of tau at Ser356, Ser396 and Ser404. In conclusion, we show that signalling pathways that include CDK/pRB/E2F1 modulate neuronal death signals by activating downstream transcription factors c-Myb and PAX6, leading to GSK-3β activation and tau pathology, providing novel potential targets for pharmaceutical intervention.

Alzheimer's disease (AD) is the most common form of neurodegenerative disorder characterized by progressive loss of memory and cognitive functions. There are two pathological hallmarks, including accumulation of amyloid plaques composed of β-amyloid peptide (Aβ) and deposits of neurofibrillatory tangles (NFT). Cyclin-dependent kinase 5 (CDK5), a serine/threonine kinase, plays an important role in synaptic plasticity and cognitive behavior. Sulforaphene (SF) has been demonstrated to exert anti-AD activity in AD rat model. In this study, we aimed to evaluate the cognitive deficits improving effects of SF on in TgCRND8 mice and to elucidate the underlying molecular mechanisms.TgCRND8 mice were intragastrically treated with SF (25 and 50 mg/kg) for 4 months from 3-month-old. The cognitive functions were assessed using Morris Water Maze Test. Cultured primary mouse neurons were pre-treated with SF, followed by co-treatment with Aβ1-42 oligomers. CDK5 inhibitor (roscovitine) was used to determine the involvement of CDK5/p25 pathway in the anti-AD effects of SF in primary neurons.Our results showed that SF treatment significantly ameliorated the cognitive deficits in TgCRND8 mice and protected primary mouse neurons against Aβ1-42 induced neurotoxicity. SF could modulate the expression of Aβ production related markers, and suppress the phosphorylation of tau protein at specific sites in the TgCRND8 mice. In addition, SF enhanced the expressions of synaptic plasticity related markers and CDK5. SF also markedly suppressed the CDK5/p25 activity.SF is a potent CDK5 inhibitor and a potential therapeutic agent for treatment and prevention of AD. Moreover, SF inhibited the overexpression of CDK5 in primary neurons of mouse.

Beta amyloid (Aβ) is a key component of parenchymal Aβ plaques and vascular Aβ fibrils, which lead to cerebral amyloid angiopathy (CAA) in Alzheimer’s disease (AD). Recent studies have revealed that Aβ contained in the cerebrospinal fluid (CSF) can re-enter into brain through paravascular spaces. However, whether Aβ in CSF may act as a constant source of pathogenic Aβ in AD is still unclear. This study aimed to examine whether Aβ pathology could be worsened when CSF Aβ level was enhanced by intra-cisternal infusion of aged brain extract containing abundant Aβ in TgCRND8 host mice. TgCRND8 mouse is an AD animal model which develops predominant parenchymal Aβ plaques in the brain at as early as 3 months of age. Here, we showed that single intracisternal injection of Aβ seeds into TgCRND8 mice before the presence of Aβ pathology induced robust prion-like propagation of CAA within 90 days. The induced CAA is mainly distributed in the cerebral cortex, hippocampus and thalamus of TgCRND8 mice. Surprisingly, despite the robust increase in CAA levels, the TgCRND8 mice had a marked decrease in parenchymal Aβ plaques and the plaques related neuroinflammation in the brains compared with the control mice. These results amply indicate that Aβ in CSF may act as a source of Aβ contributing to the growth of vascular Aβ deposits in CAA. Our findings provide experimental evidence to unravel the mechanisms of CAA formation and the potential of targeting CSF Aβ for CAA.

This study aims to evaluate the photodynamic efficacy of purpurin 18 (pu-18) on triple negative breast cancer both in vitro and in vivo. Two states of 4T1 cells, 2D culture and 3D spheroids, were used to evaluate the photodynamic action of pu-18 in vitro. The in vitro study results indicated that for the 4T1 2D cell culture, the photodynamic therapy (PDT) treatment showed significant photocytotoxicity at low pu-18 concentrations following light irradiation. Pu-18 was found to distribute on the lysosomes, mitochondria, Golgi apparatus, and endoplasmic reticulum. After irradiation, pu-18 can generate ROS to destroy the mitochondrial membrane potential (MMP) and eventually induce apoptosis in the 2D 4T1 cells. Light-activated pu-18 could also induce the destruction of the 3D 4T1 cell spheroids. The in vivo study was conducted by using a subcutaneous 4T1 breast cancer animal model. The results demonstrated that pu-18 could remain in the tumor for more than 4 days by direct intra-tumoral injection. The PDT treatment was performed every 2 days for a total of 3 times. The results showed that PDT treatment could significantly inhibit tumor growth in vivo, indicating a good photodynamic efficacy of pu-18 in the mouse breast cancer model, without influencing weight and major organ function. The survival pattern results showed that PDT treatment could largely extend the survival time of mice with breast cancer. The preliminary conclusion is that photodynamic treatment using pu-18 is effective at preventing the growth of triple negative breast cancer cells both in vitro and in vivo. A combination of light irradiation and pu-18 could therefore be a worthwhile approach for the treatment of triple negative breast cancer.

The fluid-filled cystic cavity sealed by a dense scar developed following traumatic spinal cord injury (SCI) has been a major obstacle to neural regeneration and functional recovery. Here the transected lesion is bridged using a functional self-assembling peptide (F-SAP) hydrogel loaded with membrane-permeable intracellular sigma peptide (ISP) and intracellular LAR peptide (ILP), targeted at perturbing chondroitin sulfate proteoglycan (CSPG) inhibitory signaling. As compared to F-SAP hydrogel loaded with chondroitinase ABC, the F-SAP+ISP/ILP promotes a beneficial anti-inflammatory response via manipulation of microglia/macrophages infiltration and assembly of extracellular matrix (ECM) molecules into fibrotic matrix rather than scarring tissues. The remodeled ECM creates a permissive environment that supports axon regrowth and the formation of synaptic connections with neurons derived from endogenous neural stem cells. The remodeled networks contribute to functional recovery, as demonstrated by improved hind limb movements and electrophysiological properties. This work proposes a unique mechanism that ECM remodeling induced by CSPG-manipulation-based anti-inflammation can construct a permissive environment for neural regeneration, and shed light on the advancement of manipulation of cascading cellular and molecular events potential for endogenous repair of SCI.

Brachial plexus avulsion (BPA) occurs when the spinal nerve roots are pulled away from the surface of the spinal cord and disconnects neuronal cell body from its distal downstream axon, which induces massive motoneuron death, motor axon degeneration and de-innervation of targeted muscles, thereby resulting in permanent paralysis of motor functions in the upper limb. Avulsion injury triggers oxidative stress and intense local neuroinflammation at the lesioned site, leading to the death of most motoneurons. Berberine (BBR), a natural isoquinoline alkaloid derived from medicinal herbs of Berberis and Coptis species, has been reported to possess neuro-protective, anti-inflammatory and anti-oxidative effects in various animal models of central nervous system (CNS)-related disorders. In this study, we aimed to investigate the effect of BBR on motoneuron survival and axonal regeneration following spinal root avulsion plus re-implantation in rats. Our results indicated BBR significantly accelerated motor function recovery in the forelimb as revealed by the increased Terzis grooming test score, facilitated motor axon regeneration as evidenced by the elevated number of Fluoro-Gold-labeled and P75-positive regenerative motoneurons. The survival of motoneurons was notably promoted by BBR administration presented with boosted ChAT-immunopositive and neutral red-stained neurons. BBR treatment efficiently alleviated muscle atrophy, attenuated functional motor endplates loss in biceps and prevented the reduction of motor axons in the musculocutaneous nerve. Additionally, BBR treatment markedly mitigated the avulsion-induced neuroinflammation via inhibiting microglial and astroglial reactivity, up-regulated the expression of antioxidative indicator Cu/Zn SOD, and down-regulated the levels of nNOS, 3-NT, lipid peroxidation and NF-κB, as well as promoted SIRT1, PI3K and Akt activation. Collectively, BBR might be a promising therapy to assist re-implantation surgery for the treatment of BPA.

Axonopathy manifested by axon swellings might constitute one of the earliest pathological features of Alzheimer's disease. It has been proposed that axonopathy might be associated with the origin of Aβ plaques. However, how axonopathy leads to Aβ plaque pathogenesis remains elusive. Our previous studies have shown that Aβ neuropathology (mainly diffuse plaques) selectively occurred in the regions of corticospinal tract (CST) pathway and its innervated region in the spinal cord of TgCRND8 mice. In this study, we investigated the occurrence and progression of axonopathy and the possible implication in Aβ plaque pathogenesis in the spinal cord of TgCRND8 mice. By anterograde labeling of CST system with a neuroanatomical tracer, we found that dilated corticospinal axons started to appear at 7 months, then exhibited an age-dependent increase. These abnormal structures appear before any plaque deposits are visible in the spinal cord of the mice. Importantly, they colocalized with Aβ plaques in either the white matter or gray matter of the spinal cord at later stages, suggesting that these axonal swellings might represent the initial stages of Aβ plaque formation, and could play a role in Aβ plaque pathogenesis. Furthermore, using ultrastructural analysis we demonstrated that intracellular contents in the axonal dystrophies such as various dense vesicles leaked out into the extracellular matrix under a condition of axon swelling rupture in CST pathways of spinal cord. This provided precise structural evidence that how the Aβ leaks out from the axonal dystrophies into extracellular matrix and how an axonal swelling might serve as a nidus of amyloid plaque formation.

Ethnopharmacological relevance Chrysanthemum indicum Linne (C. indicum), a healthy food and folk medicine in China for thousands of years, has been reported to exert heat-clearing and detoxifying effects and extensively applied to treat various symptoms such as inflammation diseases, hepatitis and headache. Aim of this study The purpose of the present study was to investigate the protective effect of the supercritical carbon dioxide fluid extract from flowers and buds of C. indicum (CISCFE) on D -galactose-induced brain and liver damage during aging process and to illuminate the underlying mechanisms. Materials and methods Mice were orally administrated with CISCFE (100, 150 and 300 mg/kg) after injection with D -galactose. 24 h after the last administration, the blood samples, whole brain and liver tissues were collected for biochemical analysis, histological examination and western blot analysis. The body weight, spleen and thymus indexes, alanine transaminase (ALT), aspartate transaminase (AST), total antioxidant capacity (T-AOC), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), malondialdehyde (MDA) in brain and liver, interleukin-1β (IL-1β), interleukin-6 (IL-6), and necrosis factor-α (TNF-α) were detected. Besides, the expressions of Bax, Bcl-2 and cleaved caspase-3 were determined by western blot assay. Results The results indicated that CISCFE effectively increased the suppressed body weight, attenuated the decline of thymus and spleen indexes, and reduced the elevated levels of ALT and AST induced by D -gal. Furthermore, CISCFE might notably alleviate D -gal-induced abnormal alterations in structure and function of brain and liver dose-dependently via renewing normal antioxidant enzymes activities (SOD, CAT, GSH-Px), reducing MDA accumulation, decreasing inflammatory cytokines productions (IL-1β, IL-6, TNF-α), as well as attenuating the increase of Bax/Bcl-2 ratio and cleaved caspase-3 activation in the liver and brain. Conclusions Taken together, our present results suggested that CISCFE treatment could effectively mitigate the D -gal-induced hepatic and cerebral injury, and the underlying mechanism might be tightly related to the decreased oxidative stress, inflammation and apoptosis, indicating CISCFE might be an alternative and promising agent for the treatment of aging and age-associated brain and liver diseases.

Graphical abstract, Highlights • In the same content, Nano-HO could effectively enhance the bioactivity of HO remarkably and prolonged its circulation time in rats. • Nano-HO improved cognitive deficits in TgCRND8 mice. • Nano-HO suppressed neuroinflammatory response (TNF-α, IL-Iβ, IL-6) inhibited the activation of microglia, astrocyte and A-β plaque burdens in TgCRND8 mice. • Nano-HO ameliorated AD through regulating APP processing, preventing tau hyperphosphorylation and modulating JNK/CDK5/GSK-3β pathway. • Nano-HO regulated the composition and structure of gut microbiota to protect the gut microflora and its stability., Introduction Honokiol (HO) exerts neuroprotective effects in several animal models of Alzheimer’s disease (AD), but the poor dissolution hampers its bioavailability and therapeutic efficacy. Objectives A novel honokiol nanoscale drug delivery system (Nano-HO) with smaller size and excellent stability was developed in this study to improve the solubility and bioavailability of HO. The anti-AD effects of Nano-HO was determined. Methods Male TgCRND8 mice were daily orally administered Nano-HO or HO at the same dosage (20 mg/kg) for 17 consecutive weeks, followed by assessment of the spatial learning and memory functions using the Morris Water Maze test (MWMT). Results Our pharmacokinetic study indicated that the oral bioavailability was greatly improved by Nano-HO. In addition, Nano-HO significantly improved cognitive deficits and inhibited neuroinflammation via suppressing the levels of TNF-α, IL-6 and IL-1β in the brain, preventing the activation of microglia (IBA-1) and astrocyte (GFAP), and reducing β-amyloid (Aβ) deposition in the cortex and hippocampus of TgCRND8 mice. Moreover, Nano-HO was more effective than HO in modulating amyloid precursor protein (APP) processing via suppressing β-secretase, as well as enhancing Aβ-degrading enzymes like neprilysin (NEP). Furthermore, Nano-HO more markedly inhibited tau hyperphosphorylation via decreasing the ratio of p-Tau (Thr 205)/tau and regulating tau-related apoptosis proteins (caspase-3 and Bcl-2). In addition, Nano-HO more markedly attenuated the ratios of p-JNK/JNK and p-35/CDK5, while enhancing the ratio of p-GSK-3β (Ser9)/GSK-3β. Finally, Nano-HO prevented the gut microflora dysbiosis in TgCRND8 mice in a more potent manner than free HO. Conclusion Nano-HO was more potent than free HO in improving cognitive impairments in TgCRND8 mice via inhibiting Aβ deposition, tau hyperphosphorylation and neuroinflammation through suppressing the activation of JNK/CDK5/GSK-3β signaling pathway. Nano-HO also more potently modulated the gut microbiota community to protect its stability than free HO. These results suggest that Nano-HO has good potential for further development into therapeutic agent for AD treatment.

Background Honokiol (HO) exerts neuroprotective effects in several animal models of Alzheimer’s disease (AD), but the poor dissolution hampers its bioavailability and therapeutic efficacy. A novel honokiol nanoscale drug delivery system (Nano-HO) with smaller size and excellent stability was developed in this study to improve the solubility and bioavailability of HO. Methods Male TgCRND8 mice were administered with Nano-HO or HO at the same dosage (20 mg/kg) by oral gavage daily for 17 consecutive weeks, followed by assessment of the spatial learning and memory functions with the Morris Water Maze test (MWMT). Results Nano-HO and HO could significantly improve cognitive deficits and inhibit neuroinflammation via suppressing the levels of tumor necrosis factor (TNF-α), interleukin 6 (IL-6) and IL-1β in the brain, preventing the activation of microglia (IBA-1) and astrocyte (GFAP), and reducing β-amyloid (Aβ) deposition in the cortex and hippocampus of TgCRND8 mice. In addition, Nano-HO and HO could modulate amyloid precursor protein (APP) processing and phosphorylation via suppressing β-secretase including β-site APP cleaving enzyme-1 (BACE-1) and phosphorylated APP (Thr 668), inhibiting γ-secretase including presenilin-1 (PS-1) and anterior pharynx-defective-1 (APH-1), as well as enhancing Aβ-degrading enzymes such as insulin degrading enzyme (IDE) and neprilysin (NEP). Moreover, Nano-HO remarkably inhibited tau hyperphosphorylation via decreasing the levels of p-tau (Thr 205) and p-tau (Ser 404), as well as regulating tau-related apoptosis proteins including caspase-3 and Bcl-2. Furthermore, Nano-HO and HO markedly attenuated the ratios of p-JNK/JNK and p-35/CDK5, while enhancing the ratio of p-GSK-3β (Ser9)/GSK-3β. On the other hand, Nano-HO and HO prevented the alterations on the composition of gut microbiota in TgCRND8 mice. Conclusions Nano-HO was more effective than regular HO in improving cognitive impairments in TgCRND8 mice via inhibiting Aβ deposition, tau hyperphosphorylation and neuroinflammation through suppressing the activation of JNK/CDK5/GSK-3β signaling pathway. Nano-HO was also more potently modulate the gut microbiota community to protect its stability as compared with that of regular HO. Our results amply indicated that HO with nano-sized drug delivery system has good potential for further development into therapeutic agent for AD treatment.

This study aims to evaluate the photodynamic efficacy of purpurin 18 (pu-18) on triple negative breast cancer both in vitro and in vivo. Two states of 4T1 cells, 2D culture and 3D spheroids, were used to evaluate the photodynamic action of pu-18 in vitro. The in vitro study results indicated that for the 4T1 2D cell culture, the photodynamic therapy (PDT) treatment showed significant photocytotoxicity at low pu-18 concentrations following light irradiation. Pu-18 was found to distribute on the lysosomes, mitochondria, Golgi apparatus, and endoplasmic reticulum. After irradiation, pu-18 can generate ROS to destroy the mitochondrial membrane potential (MMP) and eventually induce apoptosis in the 2D 4T1 cells. Light-activated pu-18 could also induce the destruction of the 3D 4T1 cell spheroids. The in vivo study was conducted by using a subcutaneous 4T1 breast cancer animal model. The results demonstrated that pu-18 could remain in the tumor for more than 4 days by direct intra-tumoral injection. The PDT treatment was performed every 2 days for a total of 3 times. The results showed that PDT treatment could significantly inhibit tumor growth in vivo, indicating a good photodynamic efficacy of pu-18 in the mouse breast cancer model, without influencing weight and major organ function. The survival pattern results showed that PDT treatment could largely extend the survival time of mice with breast cancer. The preliminary conclusion is that photodynamic treatment using pu-18 is effective at preventing the growth of triple negative breast cancer cells both in vitro and in vivo. A combination of light irradiation and pu-18 could therefore be a worthwhile approach for the treatment of triple negative breast cancer.

Alzheimer’s disease (AD) is a common neurodegenerative disease characterized by progressive memory loss. Magnolol (MN), the main active ingredient of Magnolia officinalis, possesses anti-AD effects in several experimental models of AD. In this study, we aimed to explore whether MN could ameliorate the cognitive deficits in TgCRND8 transgenic mice and to elucidate its molecular mechanisms. Male TgCRND8 mice were orally administered with MN (20 and 40 mg/kg) daily for 4 consecutive months, followed by assessing the spatial learning and memory functions using the open-field, radial arm maze, and novel object recognition tests. The results demonstrated that MN (20 and 40 mg/kg) could markedly ameliorate the cognitive deficits in TgCRND8 mice. In addition, MN significantly increased the expression of postsynaptic density protein 93 (PSD93), PSD-95, synapsin-1, synaptotagmin-1, synaptophysin (SYN), and interleukin-10 (IL-10), while markedly reduced the protein levels of tumor necrosis factor alpha (TNF-α), IL-6, IL-1β, Aβ40, and Aβ42, and modulated the amyloid precursor protein (APP) processing and phosphorylation. Immunofluorescence showed that MN significantly suppressed the activation of microglia (Iba-1) and astrocytes (GFAP) in the hippocampus and cerebral cortex of TgCRND8 mice. Mechanistic studies revealed that MN could significantly increase the ratios of p-GSK-3β (Ser9)/GSK-3β, p-Akt (Ser473)/Akt, and p-NF-κB p65/NF-κB p65. These findings indicate that MN exerted cognitive deficits improving effects via suppressing neuroinflammation, amyloid pathology, and synaptic dysfunction through regulating the PI3K/Akt/GSK-3β and NF-κB pathways, suggesting that MN is a promising naturally occurring polyphenol worthy of further developing into a therapeutic agent for AD treatment.

// Qiuju Yuan 1, 2 , Jian Yang 3 , Wutian Wu 3, 4, 5, 6 and Zhi-Xiu Lin 1, 2 1 School of Chinese Medicine, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong SAR, China 2 Brain Research Centre, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong SAR, China 3 School of Biomedical Sciences, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China 4 State Key Laboratory of Brain and Cognitive Sciences, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China 5 Research Center of Reproduction, Development and Growth, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China 6 GHM Institute of CNS regeneration, Jinan University, Guangzhou, China Correspondence to: Zhi-Xiu Lin, email: linzx@cuhk.edu.hk Wutian Wu, email: wtwu@hkucc.hku.hk Keywords: axonal transport, dystrophic axon swellings, corticospinal tract, motor deficits Received: February 03, 2017 Accepted: May 08, 2017 Published: June 09, 2017 ABSTRACT There have been an increasing number of reports of non-cognitive symptoms in Alzheimer’s disease (AD). Some symptoms are associated with the loss of motor functions, e.g. gait disturbances, disturbed activity level and balance. Consistent with clinical findings, several AD mouse models harboring amyloid pathology develop motor impairment. Although the factors that contribute to the motor deficits have not yet been determined, it has been suggested that axonopathy is one of the key factors that may contribute to this particular feature of the disease. Our previous study found that TgCRND8 mice exhibited profound motor deficits as early as 3 months old. In this study, we explored the possible factors that may be related to motor deficits in TgCRND8 mice. Results from silver, neurofilament and amyloid precursor protein (APP) staining revealed no axonopathy occurred in the brain and spinal cord of TgCRND8 mice at the age of 3 months. Anterograde labeling of corticospinal tract of spinal cord and electronic microscopy (EM) analysis showed that no axonopathy occurred in TgCRND8 mice at the age of 3 months. According to these results, it could be concluded that no axonal alterations were evident in the TgCRND8 mice when motor deficits was overt. Thus, axonopathy may play a less prominent role in motor deficits in AD. These results suggest that mechanisms by which motor function undergo impairment in AD need to be further studied.

Cerebral amyloid angiopathy (CAA) refers to pathological changes occurring in cerebral blood vessels caused by deposition of beta amyloid (Aβ) protein. However, the mechanisms involved in the origin of Aβ for the formation of CAA and its link to parenchymal amyloid depositions remained to be unraveled. Here, we found CAA and parenchymal plaques distributed separately instead of mingling with each other in the spinal cord of TgCRND8 mice. Parenchymal plaques predominantly located in the dorsal horn whereas CAA distributed in the ventral horn. We further found that the ratio of Aβ40/Aβ42 was significantly higher in the ventral than that in the dorsal by ELISA assay, suggesting that origin of Aβ forming parenchymal plaques may be different from that of CAA in the spinal cord. This hypothesis was further demonstrated by the surgical methods which indicated eliminating parenchymal plaques did not alter CAA in the affected spinal cord. We also examined the ratio of Aβ40/Aβ42 in the cerebral spinal fluid (CSF) in order to identify the origin of the CAA formation, and found the Aβ40/Aβ42 ratio was similar to that of CAA formation in the ventral horn. We further demonstrated that CSF tracer distributed along ventral horn vessels, in exactly the same pattern as Aβ deposition in CAA in ventral part of spinal cord. These findings verified the concept that CSF influx may act as a constant source for delivering Aβ, and contribute to the growth of paraarterial deposits in CAA. Taken together, the results of the present study highlight the important role of the Aβ40/Aβ42 ratio in determining vascular versus parenchymal amyloid deposition. Unlike parenchymal plaques, Aβ of CAA comes from CSF; thus, manipulation of CSF Aβ could represent a novel strategy to treat CAA.

Isorhynchophylline (IRN) has been demonstrated to have distinct anti-Alzheimer’s disease (AD) activity in several animal models of AD. In this study, we aimed at evaluating the preventive effect of IRN on the cognitive deficits and amyloid pathology in TgCRND8 mice. Male TgCRND8 mice were administered with IRN (20 or 40 mg/kg) by oral gavage daily for 4 months, followed by assessing the spatial learning and memory functions with the Radial Arm Maze (RAM) test. Brain tissues were determined immunohistochemically or biochemically for changes in amyloid pathology, tau hyperphosphorylation and neuroinflammation. Our results revealed that IRN (40 mg/kg) significantly ameliorated cognitive deficits in TgCRND8 mice. In addition, IRN (40 mg/kg) markedly reduced the levels of Aβ40, Aβ42 and tumor necrosis factor (TNF-α), interleukin 6 (IL-6) and IL-1β, and modulated the amyloid precursor protein (APP) processing and phosphorylation by altering the protein expressions of β-site APP cleaving enzyme-1 (BACE-1), phosphorylated APP (Thr668), presenilin-1 (PS-1) and anterior pharynx-defective-1 (APH-1), as well as insulin degrading enzyme (IDE), a major Aβ-degrading enzyme. IRN was also found to inhibit the phosphorylation of tau at the sites of Thr205 and Ser396. Immunofluorescence showed that IRN reduced the Aβ deposition, and suppressed the activation of microglia (Iba-1) and astrocytes (GFAP) in the cerebral cortex and hippocampus of TgCRND8 mice. Furthermore, IRN was able to attenuate the ratios of p-c-Jun/c-Jun and p-JNK/JNK in the brains of TgCRND8 mice. IRN also showed marked inhibitory effect on JNK signaling pathway in the Aβ-treated rat primary hippocampus neurons. We conclude that IRN improves cognitive impairment in TgCRND8 transgenic mice via reducing Aβ generation and deposition, tau hyperphosphorylation and neuroinflammation through inhibiting the activation of JNK signaling pathway, and has good potential for further development into pharmacological treatment for AD.

Several genetic studies have identified a rare variant of triggering receptor expressed on myeloid cells 2 (TREM2) as a risk factor for Alzheimer’s disease (AD). However, findings on the effects of TREM2 on Aβ deposition are quite inconsistent in animal studies, requiring further investigation. In this study, we investigated whether elevation of TREM2 mitigates Aβ pathology in TgCRND8 mice. We found that peripheral nerve injury resulted in a robust elevation of TREM2 exclusively in reactive microglia in the ipsilateral spinal cord of aged TgCRND8 mice at the age of 20 months. TREM2 expression appeared on day 1 post-injury and the upregulation was maintained for at least 28 days. Compared to the contralateral side, neither amyloid beta plaque load nor soluble Aβ40 and Aβ42 levels were attenuated upon TREM2 induction. We further showed direct evidence that TREM2 elevation in reactive microglia did not affect amyloid-β pathology in plaque-bearing TgCRND8 mice by applying anti-TREM2 neutralizing antibody to selectively block TREM2. Our results question the ability of TREM2 to ameliorate established Aβ pathology, discouraging future development of disease-modifying pharmacological treatments targeting TREM2 in the late stage of AD.

Increasing evidence suggests that activation of satellite glia cells (SGCs) in sensory ganglia play important roles in the development of neuropathic pain. The present study aimed to investigate the involvement of SGC activation in a novel model of motor nerve injury induced pain hypersensitivity. The neuropathic pain model was established by cervical 8 ventral root avulsion (C8VA). Glial fibrillary acidic protein (GFAP) was used as a marker of SGC activation. Unilateral C8VA resulted in mechanical allodynia, but not thermal hyperalgesia in bilateral paws. Expectedly, SGCs were robustly activated on as early as 1 day and persisted for at least 7 days in the ipsilateral and contralateral dorsal root ganglia (DRG) of C6, C7 and C8 after C8VA. Double immunofluorescence showed that almost all the activated SGCs enveloped neurofilament 200 (NF200) positive myelinated neurons in DRG. Local application of fluorocitrate (FC), a glial metabolism inhibitor, significantly decreased the number of activated SGCs and alleviated bilateral mechanical allodynia. These results suggest that SGC activation contributed to ipsilateral and mirror-image pain hypersensitivity after C8VA. Inhibition of SGC activation represented a promising therapeutic strategy for the management of neuropathic pain following brachial plexus root avulsion.

Dorsal root injury commonly results in irreversible loss of sensory functions because of the limited intrinsic regenerative capacity of adult sensory axons and the growth-inhibitory environment at the dorsal root entry zone (DREZ) between the dorsal root and the spinal cord. Chondroitin sulfate proteoglycans (CSPGs) are the dominant suppressors of axonal regeneration, acting via neuronal receptors including protein tyrosine phosphatase-σ (PTPσ). ISP (Intracellular Sigma Peptide) is a small peptide mimetic of the PTPσ wedge region that has been developed to target PTPσ and relieve CSPG inhibition. Extracellular regulated kinases (Erks) and cAMP response element binding protein (CREB) are signaling molecules downstream of CSPGs and PTPσ; they are expressed in neurons and essential for axon growth. In this study, we observed that ISP administration could promote sensory function restoration in adult rats after dorsal spinal root crush injury. Our results show that systemic ISP administration would not only significantly increase sensory axon regeneration and functional recovery, but also activate Erk and CREB signaling pathway. Furthermore, ISP has also been verified to increase dorsal root ganglion axonal remyelination in vitro. These results suggest that modulation of PTPσ by ISP represents a promising therapeutic strategy for sensory neuronal injuries.

High energy cosmic ray electrons plus positrons (CREs), which lose energy quickly during their propagation, provide an ideal probe of Galactic high-energy processes and may enable the observation of phenomena such as dark-matter particle annihilation or decay. The CRE spectrum has been directly measured up to $\sim 2$ TeV in previous balloon- or space-borne experiments, and indirectly up to $\sim 5$ TeV by ground-based Cherenkov $\gamma$-ray telescope arrays. Evidence for a spectral break in the TeV energy range has been provided by indirect measurements of H.E.S.S., although the results were qualified by sizeable systematic uncertainties. Here we report a direct measurement of CREs in the energy range $25~{\rm GeV}-4.6~{\rm TeV}$ by the DArk Matter Particle Explorer (DAMPE) with unprecedentedly high energy resolution and low background. The majority of the spectrum can be properly fitted by a smoothly broken power-law model rather than a single power-law model. The direct detection of a spectral break at $E \sim0.9$ TeV confirms the evidence found by H.E.S.S., clarifies the behavior of the CRE spectrum at energies above 1 TeV and sheds light on the physical origin of the sub-TeV CREs., Comment: 18 pages, 6 figures, Nature in press, doi:10.1038/nature24475

// Wen Wen Xu 1, 2 , Bin Li 1, 2, 3 , Alfred KY Lam 4 , Sai Wah Tsao 1, 3 , Simon YK Law 3, 5 , Kwok Wah Chan 2, 3, 6 , Qiu Ju Yuan 1 , Annie LM Cheung 1, 2, 3 1 Department of Anatomy, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong SAR, China 2 The University of Hong Kong-Shenzhen Institute of Research and Innovation (HKU-SIRI), China 3 Centre for Cancer Research, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong SAR, China 4 Department of Pathology, Griffith Medical School and Griffith Health Institute, Gold Coast Campus, Gold Coast, QLD 4222, Australia 5 Department of Surgery, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong SAR, China 6 Department of Pathology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong SAR, China Correspondence to: Annie LM Cheung, e-mail: lmcheung@hku.hk Keywords: Tumor angiogenesis, Bone marrow-derived cells, Tumor microenvironment, VEGF receptors, Antibody therapy Received: September 18, 2014 Accepted: November 19, 2014 Published: December 17, 2014 ABSTRACT Increasing appreciation of tumor heterogeneity and the tumor-host interaction has stimulated interest in developing novel therapies that target both tumor cells and tumor microenvironment. Bone marrow derived cells (BMDCs) constitute important components of the tumor microenvironment. In this study, we aim to investigate the significance of VEGFR1- and VEGFR2-expressing non-tumor cells, including BMDCs, in esophageal cancer (EC) progression and in VEGFR1/VEGFR2-targeted therapies. Here we report that VEGFR1 or VEGFR2 blockade can significantly attenuate VEGF-induced Src and Erk signaling, as well as the proliferation and migration of VEGFR1 + and VEGFR2 + bone marrow cells and their pro-invasive effect on cancer cells. Importantly, our in vivo data show for the first time that systemic blockade of VEGFR1 + or VEGFR2 + non-tumor cells with neutralizing antibodies is sufficient to significantly suppress esophageal tumor growth, angiogenesis and metastasis in mice. Moreover, our tissue microarray study of human EC clinical specimens showed the clinicopathological significance of VEGFR1 and VEGFR2 in EC, which suggest that anti-VEGFR1/VEGFR2 therapies may be particularly beneficial for patients with aggressive EC. In conclusion, this study demonstrates the important contributions of VEGFR1 + and VEGFR2 + non-tumor cells in esophageal cancer progression, and substantiates the validity of these receptors as therapeutic targets for this deadly disease.

Object The purpose of this study was to examine whether choline acetyltransferase (ChAT) staining can be used for assessing the rate of motor neuron regeneration at an early phase of axon outgrowth. Methods The authors developed a new sciatic nerve crush model in adult mice. In this model, in addition to performing a sciatic nerve crush injury, the authors excised the ipsilateral lumbar L3–6 dorsal root ganglion (DRG), which resulted in degeneration of the sensory fibers entering into the sciatic nerve. Crushed nerve sections obtained at Day 3 or Day 7 postinjury were analyzed by means of immunostaining. Results The immunostaining showed that ChAT, a motor axon–specific antigen, was totally co-localized with growth-associated protein 43 (GAP-43), which is expressed in regenerating nerves and transported into growth cones. Conclusions Our results suggest that measuring the length of motor axon outgrowth by ChAT immunostaining is reliable. ChAT staining provides a more convenient method for evaluating the rate of motor axon outgrowth in a mixed nerve.

In this study, the synthesis of 5-hydroxymethylfurfural (5-HMF) from sucrose was carried out in ionic liquids 1-ethyl-3-methylimidazolium bromide ([Emim]Br) catalyzed by amino acids, and tyrosine displays the best activity. Under the optimal reaction conditions, 76.0% yield of 5-HMF from sucrose was obtained at 160 °C for 4 h. The uniquely effective activity of tyrosine for sucrose conversion into 5-HMF in [Emim]Br is mainly attributed to its two types of active sites, free base NH2 and dissociated H+ sites. The former one plays a crucial role in the isomerization of glucose to fructose, and the latter one is active in the hydrolysis of sucrose into monosaccharides and dehydration of generated fructose to 5-HMF. Furthermore, the presence of acidic phenol group in tyrosine also has the synergic catalytic effect on sucrose conversion. In addition, with the use of tyrosine catalyst, other carbohydrates to form 5-HMF were also tested, and the effects of solvent, reaction temperature and reaction time on sucrose conversion into 5-HMF were investigated in detail. A possible mechanism for this catalytic process has been proposed.

Peripheral nerve injury still remains a refractory challenge for both clinical and basic researchers. A novel nanofiber conduit made of blood vessel and filled with amphiphilic hydrogel of self-assembling nanofiber scaffold (SAPNS) was implanted to repair a 10 mm nerve gap after sciatic nerve transection. Empty blood vessel conduit was implanted serving as control. Results showed that this novel nanofiber conduit enabled the peripheral axons to regenerate across and beyond the 10 mm gap. Motoneuron protection, axonal regeneration and remyelination were significantly enhanced with SAPNS scaffold treatments. The target reinnervation and functional recovery induced by the regenerative nerve conduit suggest that SAPNS-based conduit is highly promising application in the treatment of peripheral nerve defect. From the Clinical Editor In this paper by Zhan et al, a novel self-assembling nanofiber scaffold is reported to promote regeneration of peripheral nerves in a sciatic nerve injury model. The promising results and the obvious medical need raises hope for a clinical translation of this approach hopefully in the near future.

Conflicting findings exist regarding the formation of diffuse and dense-core β-amyloid (Aβ) plaques in Alzheimer’s disease (AD). In the present study, we characterized Aβ plaque types in the brain and spinal cord of TgCRND8 mice, which express a transgene incorporating both the Indiana mutation (V717F) and the Swedish mutations (K670N/M671L) in the human amyloid-β protein precursor (APP) gene. By combining immunohistochemistry and thioflavin S staining, we were able to define dense-core and diffuse plaques in neocortex of the brain and spinal cord of 9 week-, 5 month-, 10 month- and 20-month-old TgCRND8 mice. The senile plaques in the neocortex were predominantly dense-core plaques, even in the youngest mice. However, diffuse plaques were instead detected in spinal cord of the mice, regardless of age. Our results that relative predominance of dense-core plaques in the neocortex and diffuse plaques in the spinal cord of TgCNRD8 mice of all disease durations argue against the notion that diffuse plaques may represent an early stage in the evolution of dense-core plaques. Furthermore, we also found that the ratio of Aβ42/Aβ40 of the brain was much higher than that of the spinal cord by Aβ ELISA assay. Our findings strongly indicate that diffuse and dense-core plaques may form via independent processes in AD and Aβ42 is more prone to form dense-core plaques than is Aβ40.

Neural progenitor cells (NPCs) are suggested to be a valuable source of cell transplant in treatment of various neurological diseases because of their distinct attributes. They can be expanded and induced to differentiate in vitro. However, it remains uncertain whether in vitro expanded NPCs have the capacity to give rise to functional motoneurons after transplantation in vivo. Here, we showed that in vitro expanded NPCs, when transplanted into the musculocutaneous nerve, generated motoneuron-like cells that exhibited typical morphology with large cell bodies, expressed specific molecules, and extended axons to form functional connections with the target muscle. In contrast, transplanted NPCs failed to yield motoneurons in the injured ventral horn of the spinal cord. The results of the study demonstrate that NPCs have the potential to generate functional motoneurons in an appropriate environment. The distinct differentiating fate of NPCs in the musculocutaneous nerve and the injured ventral horn suggests the importance and necessity of modifying the host microenvironment in use of NPCs for cell replacement therapies for motoneuron diseases.

Post-injury nerve regeneration of the peripheral nervous system declines with age, but the mechanisms underlying the weakened axonal regeneration are not well understood. Increased synthesis and activity of the AP-1 transcription factor c-Jun have been implicated in efficient motor axonal regeneration. In the present study, we evaluated the hypothesis that the impaired regenerative capacity in the aged is associated with impaired induction of c-Jun. In non-manipulated young adult or aged mice, no c-Jun and its phosphorylated form were detected in the ventral horn of the spinal cord. Following nerve crush, significant c-Jun and phosphorylated c-Jun occurred in the injured motoneurons of young adult mice, but not in aged animals. In accord with the immunohistochemistry, Western blots also showed that sciatic nerve crush induced c-Jun and its phosphorylation expression in the ventral horn of young adult but not in aged mice. Changes in c-Jun mRNA level detected by in situ hybridization are congruent with that in c-Jun protein content, showing an increase at 5 days after crush in young adult but not aged. Moreover, compared with young adult mice, aged mice showed impaired motor axonal regeneration. These results demonstrate that the impaired motor axonal regeneration seen in aged mice is correlated with impaired c-Jun expression and phosphorylation following injury. These data provide a neurobiological explanation for the poor outcome associated with nerve repair in the aged.

Neuronal nitric oxide synthase (nNOS) is induced after axonal injury. The role of induced nNOS in injured neurons is not well established. In the present study, we investigated the co-expression of nNOS with GAP-43 in spinal motoneurons following axonal injury. The role of induced nNOS was discussed and evaluated. In normal rats, spinal motoneurons do not express nNOS or GAP-43. Following spinal root avulsion, expression of nNOS and GAP-43 were induced and colocalized in avulsed motoneurons. Reimplantation of avulsed roots resulted in a remarkable decrease of GAP-43- and nNOS-IR in the soma of the injured motoneurons. A number of GAP-43-IR regenerating motor axons were found in the reimplanted nerve. In contrast, the nNOS-IR was absent in reimplanted nerve. These results suggest that expression of GAP-43 in avulsed motoneurons is related to axonal regeneration whereas nNOS is not.

One of the major pathological features of Alzheimer's disease (AD) is the appearance of senile plaques characterized by extracellular aggregation of amyloid beta-peptide (Abeta) fibrils. Inhibition of Abeta fibril aggregation is therefore viewed as one possible method to halt the progression of AD. Salvianolic acid B (Sal B) is an active ingredient isolated from Salvia miltiorrhiza, a Chinese herbal medicine commonly used for the treatment of cardiovascular and cerebrovascular disorders. Recent findings show that Sal B prevents Abeta-induced cytotoxicity in a rat neural cell line. To understand the mechanism of Sal B-mediated neuroprotection, its effects on the inhibition of Abeta1-40 fibril formation and destabilization of the preformed Abeta1-40 fibrils were studied. The results were obtained using Thioflavin T fluorescence assay and Abeta aggregating immunoassay. We found that Sal B can inhibit fibril aggregation (IC(50): 1.54-5.37 microM) as well as destabilize preformed Abeta fibril (IC(50): 5.00-5.19 microM) in a dose- and time-dependent manner. Sal B is a better aggregation inhibitor than ferulic acid but less active than curcumin in the inhibition of Abeta1-40 aggregation. In electron microscope study, Sal B-treated Abeta1-40 fibrils are seen in various stages of shortening or wrinkling with numerous deformed aggregates of amorphous structure. Circular dichroism data indicate that Sal B dose dependently prevents the formation of beta-structured aggregates of Abeta1-40. Addition of preincubated Sal B with Abeta1-42 significantly reduces its cytotoxic effects on human neuroblastoma SH-SY5Y cells. These results suggest that Sal B has therapeutic potential in the treatment of AD, and warrant its study in animal models.

Intermediate filaments (IFs) of nestin and vimentin are expressed in immature astrocytes. In this study, we examined the re-expression of these early glial traits in rat reactive astrocytes in affected neuronal perikarya in supraoptic (SON) and paraventricular (PVN) nuclei induced by hypophysectomy. Double-labeling immunofluorescence confocal laser microscopy demonstrated that by 7 days post-lesion, both nestin and vimentin were present intensely in hypertrophied GFAP-IR reactive astrocytes in the area of hypophysectomized magnocellular neurons in SON and PVN, while nestin and vimentin are absent in the normal or sham-operated animals. As the gliotic reaction progressed, the morphology of nestin or vimentin-positive reactive astrocytes in SON but not PVN changed from stellate form at 7 days to thin and elongated shape, morphologically compatible with radial glia during development, at 14 days post-lesion. By 28 days post-lesion, while vimentin-IR persisted in reactive astrocytes in SON and PVN, nestin-IR could hardly be detected. The spatiotemporal pattern of nestin-IR and/or vimentin-IR in reactive astrocytes suggests astrocytes attempt to revert to a more primitive glia form indicated by changes in morphology and phenotype following hypophysectomy, which may contribute to neuronal trophism and plasticity in the lesioned HNS favoring neuronal maintenance and fiber outgrowth.

Root avulsion of the brachial plexus is a devastating injury that results in massive motoneuron death and paralysis of the corresponding muscle groups. Various approaches such as pharmaceutical applications and microsurgical interventions have been extensively studied to restore the loss of injured motoneurons and recover function. However, the treatment for brachial plexus injury is still a challenging problem. The challenge that lies in brachial plexus injury treatment is that motoneuron death in the lesioned segments is relatively high and little is known about how inhibitory signals or lack of appropriate guidance molecules influences regeneration. This chapter summarizes the anatomical structure of the brachial plexus, the experimental models of brachial plexus avulsion, neuronal and glial responses to avulsion injury, and cellular and molecular profiles in the avulsed spinal cord at various ages and in various species. This chapter also summarizes a number of potential strategies for the treatment of brachial plexus avulsion, including microsurgical interventions using peripheral nerve graft or ventral root reimplantation; the use of small molecules, trophic factors, or small interfering RNA to promote motoneuron survival and regeneration; the application of inhibitors of the degeneration pathway; cell-based therapies; and, last, combined therapies for brachial plexus avulsion. We believe that a combination of multiple treatments may be necessary to acquire the most beneficial effects in treating the devastating injury characterized by motoneuron loss.

This study examined the age-related changes in nitric oxide synthase immunoreactivity (NOS-IR), survival and regeneration of magnocellular neurons in the hypothalamo-neurohypophyseal system (HNS) in rats following hypophysectomy. In adult animal, hypophysectomy induced a significant increase in NOS-IR in the supraoptic (SON), paraventricular nuclei (PVN) and median eminence (ME) by 3 days post-lesion. NOS sustained an increased level until 2 weeks after hypophysectomy and then returned to normal control level. In contrast, at postnatal day 7 (PN7), no obvious increase in NOS-IR was observed in the SON, PVN and ME following the injury compared with age-matched controls. At PN14, the same injury induced an increase in NOS-IR in SON, PVN and ME but the increase was more transient with peak NOS-IR at 3 days and returning to the corresponding control level at 1 week after hypophysectomy. In contrast to a striking age-dependent alteration in NOS-IR in the SON and PVN, hypophysectomy induced substantial degeneration of arginine vasopressin (AVP) and oxytocin (OT) neurons in the SON and PVN in both immature and adult rats and there was no obvious difference in neuronal survival after the same injury among these three groups of different ages by quantitative analysis. Following hypophysectomy, a large number of fibers were observed in the contact zone of the median eminence and the adjacent lumen of the third cerebral ventricle (V3) in adult rats, whereas few fibers could be found in the lumen of the V3 in the immature rats after the same injury. Relationships between NOS induction and magnocellular neuronal survival and regeneration were discussed.

The present study investigated the immunohistochemical localization of neuronal nitric oxide synthase (nNOS) in the hypothalamoneurohypophyseal system (HNS) of the developing rats on postnatal day 1 (PN1), 7 (PN7), 14 (PN14), 21 (PN21), and the adult rats. The nNOS-positive neurons were not discernable in the supraoptic nucleus (SON), the paraventricular nucleus (PVN), and the median eminence (ME) at PN1 and PN7. A few neurons positive for nNOS were first detected at PN14. At PN21, the nNOS-positive cells in SON and PVN rapidly increased in number. The pattern of nNOS expression at this stage approached that of the adult. Moreover, the increase of nNOS expression in the SON and PVN during the postnatal period was accompanied by the maturation of arginine vasopressin (AVP) and oxytocin (OT) neurons as indicated by the number and size of OT or AVP neurons in the SON and PVN. The patterns of AVP versus OT expression also reached that of the adult by the end of the third postnatal week. The time course of the change in nNOS expression coincided with the maturation of AVP and OT neurons in the HNS and suggested that NO synthesized by conversion of NOS is involved in the modulation of activity of neurons in the SON and PVN of the HNS. © 2005 Wiley-Liss, Inc.

Alzheimer's disease (AD) is a neurodegenerative disorder with the histopathological hallmark of extracellular accumulation of amyloid-β (Aβ) peptide in brain senile plaques. Though many studies have shown the neural toxicity from various forms of Aβ peptides, the subcellular mechanisms of Aβ peptide are still not well understood, partially due to the technical challenges of isolating axons or dendrites from the cell body for localized investigation. In this study, the subcellular toxicity and localization of Aβ peptides are investigated by utilizing a microfluidic compartmentalized device, which combines physical restriction and chemotactic guidance to enable the isolation of axons and dendrites for localized pharmacological studies. It is found that Aβ peptides induced neuronal death is mostly resulted from Aβ treatment at cell body or axonal processes, but not at dendritic neurites. Simply applying Aβ to axons alone induces significant hyperactive spiking activity. Dynamic transport of Aβ aggregates is only observed between axon terminal and cell body. In addition to differential cellular uptake, more Aβ-peptide secretion is detected significantly from axons than from dendritic side. These results clearly demonstrate the existence of a localized mechanism in Aβ-induced neurotoxicity, and can potentially benefit the development of new therapeutic strategies for AD.

Brachial plexus injury often involves traumatic root avulsion resulting in permanent paralysis of the innervated muscles. The lack of sufficient regeneration from spinal motoneurons to the peripheral nerve (PN) is considered to be one of the major causes of the unsatisfactory outcome of various surgical interventions for repair of the devastating injury. The present study was undertaken to investigate potential inhibitory signals which influence axonal regeneration after root avulsion injury. The results of the study showed that root avulsion triggered GSK-3βactivation in the injured motoneurons and remaining axons in the ventral funiculus. Systemic application of a clinical dose of lithium suppressed activated GSK-3βin the lesioned spinal cord to the normal level and induced extensive axonal regeneration into replanted ventral roots. Our study suggests that GSK-3βactivity is involved in negative regulation for axonal elongation and regeneration and lithium, the specific GSK-3βinhibitor, enhances motoneuron regeneration from CNS to PNS.

c-Jun activation has been implicated not only in neuronal degeneration, but also in survival and regeneration. Here, we investigated c-Jun activation in injured motoneurons by using a nerve crush model in neonatal rats. We identified two distinct subpopulations of motoneurons: about 60% underwent degeneration following injury whereas the remaining 40% survived and induced a regeneration response at 3 weeks post injury. However, all motoneurons examined expressed phosphorylated-c-Jun-immunoreactivity (p-c-Jun-IR) at the early stage of 3 days following injury. These results suggest that active c-Jun was induced in all neonatal motoneurons following nerve crush injury, regardless of whether they were destined to degenerate or undergo successful regeneration at a later stage. Our findings therefore support the hypothesis that active c-Jun is involved in both neuronal degeneration and regeneration.

Using two different lesion models, the spinal root avulsion and the distal nerve axotomy, the present study investigated effects of known neurotrophic factors on motoneuron survival in newborn rats. Results of the present study show that 100% of motoneurons in the lesioned spinal segment die at 1 week following root avulsion, and more than 80% of them die at 2 weeks following distal nerve axotomy. Local application of GDNF can rescue 92% of motoneurons up to 1 week from degeneration due to root avulsion and almost 100% of them up to 2 weeks from degeneration due to distal nerve axotomy. Local application of BDNF fails to prevent any motoneuron death in newborn rats following root avulsion, but it can rescue about 50% of motoneurons up to 2 weeks from degeneration due to distal nerve axotomy. CNTF and IGF-1 fail to prevent any motoneuron death following either distal nerve axotomy or root avulsion. Thus, comparing all the neurotrophic factors tested in this study, GDNF is most effective in preventing death of motoneurons following axonal injury in newborn rats.

The purpose of this study was to examine whether choline acetyltransferase (ChAT) staining can be used for assessing the rate of motor neuron regeneration at an early phase of axon outgrowth.The authors developed a new sciatic nerve crush model in adult mice. In this model, in addition to performing a sciatic nerve crush injury, the authors excised the ipsilateral lumbar L3-6 dorsal root ganglion (DRG), which resulted in degeneration of the sensory fibers entering into the sciatic nerve. Crushed nerve sections obtained at Day 3 or Day 7 postinjury were analyzed by means of immunostaining.The immunostaining showed that ChAT, a motor axon-specific antigen, was totally co-localized with growth-associated protein 43 (GAP-43), which is expressed in regenerating nerves and transported into growth cones.Our results suggest that measuring the length of motor axon outgrowth by ChAT immunostaining is reliable. ChAT staining provides a more convenient method for evaluating the rate of motor axon outgrowth in a mixed nerve.

Conflicting findings exist regarding the link between functional recovery and the regrowth of spinal tracts across the lesion leading to the restoration of functional contacts. In the present study, we investigated whether functional locomotor recovery was attributable to anatomical regeneration at postnatal day 1 (PN1), PN7, PN14 and in adult rats two months after transection injury at the tenth thoracic segment of the spinal cord. The Basso, Beattie, and Bresnahan scores showed that transection led to a failure of hindlimb locomotor function in PN14 and adult rats. However, PN1 and PN7 rats showed a significant level of stepping function after complete spinal cord transection. Unexpectedly, unlike the transected PN14 and adult rats in which the spinal cord underwent limited secondary degeneration and showed a scar at the lesion site, the rats transected at PN1 and PN7 showed massive secondary degeneration both anterograde and retrograde, leaving a >5-mm gap between the two stumps. Furthermore, retrograde tracing with fluorogold (FG) also showed that FG did not cross the transection site in PN1 and PN7 rats as in PN14 and adult rats, and re-transection of the cord caused no apparent loss in locomotor performance in the rats transected at PN1. Thus, these three lines of evidence strongly indicated that the functional recovery after transection in neonatal rats is independent of regrowth of spinal tracts across the lesion site. Our results support the notion that the recovery of locomotor function in developing rats may be due to intrinsic adaptations in the spinal circuitry below the lesion that control hindlimb locomotor activity rather than the regrowth of spinal tracts across the lesion. The difference in secondary degeneration between neonatal and adult rats remains to be explored.

The transgenic TgCRND8 mouse is widely used as an animal model of Alzheimer's disease (AD) and exhibits an early onset of senile plaque pathogenesis in the brain. Here we report that TgCRND8 mice also have amyloid-β (Aβ) neuropathology in spinal cord. TgCRND8 mice began to show obvious Aβ deposition in both gray matter of dorsal horn and white matter in the central part of dorsal column of the spinal cord at 10 months of age onward. Further experiments showed that the distribution of Aβ deposition in the spinal cord corresponds to the corticospinal tract pathway and its projection regions in TgCRND8 mice. We hypothesized that neurons in the sensorimotor cortex is the source of the Aβ peptide deposited in the spinal cord of these mice. To test the hypothesis, we ablated the sensorimotor cortex to interrupt connections between the sensorimotor cortex and spinal cord. We found that Aβ burden was significantly reduced in the denervated side compared to the contralateral side. Our results suggest that the sensorimotor cortex might be the primary source of Aβ in spinal cord of TgCRND8 mice. This is consistent with the observation that the sensorimotor cortex is one region particularly vulnerable during the progression of AD. The characteristics of Aβ distribution in TgCRND8 mice suggest that there are other ways related to the formation of Aβ plaques in addition to the terminal and synaptic release of Aβ.

Our group has shown that the polysaccharides extracted from Lycium barbarum (LBP) are neuroprotective for retinal ganglion cells (RGCs) in different animal models. Protecting RGCs from secondary degeneration is a promising direction for therapy in glaucoma management. The complete optic nerve transection (CONT) model can be used to study primary degeneration of RGCs, while the partial optic nerve transection (PONT) model can be used to study secondary degeneration of RGCs because primary degeneration of RGCs and secondary degeneration can be separated in location in the same retina in this model; in other situations, these types of degeneration can be difficult to distinguish. In order to examine which kind of degeneration LBP could delay, both CONT and PONT models were used in this study. Rats were fed with LBP or vehicle daily from 7 days before surgery until sacrifice at different time-points and the surviving numbers of RGCs were evaluated. The expression of several proteins related to inflammation, oxidative stress, and the c-jun N-terminal kinase (JNK) pathways were detected with Western-blot analysis. LBP did not delay primary degeneration of RGCs after either CONT or PONT, but it did delay secondary degeneration of RGCs after PONT. We found that LBP appeared to exert these protective effects by inhibiting oxidative stress and the JNK/c-jun pathway and by transiently increasing production of insulin-like growth factor-1 (IGF-1). This study suggests that LBP can delay secondary degeneration of RGCs and this effect may be linked to inhibition of oxidative stress and the JNK/c-jun pathway in the retina. © 2013 Li et al., published_or_final_version

Conflicting findings exist regarding the link between environmental factors and development of Alzheimer's disease (AD) in a variety of transgenic mouse models of AD. In the present study, we investigated the effect of behavioral stress on the onset and progression of Abeta pathology in the brains of TgCRND8 mice, a transgenic mouse model of AD. One group of TgCRND8 mice was subjected to restraint stress starting at 1 month of age until they were 3 months old, while restraint stress in the second group started at 4 months of age until they were 6 months old. After 2 months of treatment, no differences in the soluble, formic acid extracted, or histologically detected Abeta deposition in the cortical and hippocampal levels were found between non-stressed and stressed mice. These results showed that restraint stress alone failed to aggravate amyloid pathology when initiated either before or after the age of amyloid plaque deposition in TgCRND8 mice, suggesting that if stress aggravated AD phenotype, it may not be via an amyloid-related mechanism in the TgCRND8 mice. These findings are indicative that plaque load per se may not be used as a significant criterion for evaluating the effect of stress on AD patients., published_or_final_version