MDCK-Adaptive Mutation of A169S Changes Glycosylation Pattern of Hemagglutinin and Enhances MDCK-Based H7N9 Vaccine Virus Production without Loss of Antigenicity and Immunogenicity.

The adaptation of egg-derived H7N9 candidate vaccine virus (CVV) in the mammalian cell line is an approach to developing a high-growth virus strain for the mass production of vaccine manufacturing. The adaptive mutations that occur in hemagglutinin (HA) are critical to the activity and potency of the vaccine virus. Previously, we identified a new mutation of A169S in the HA protein of an MDCK-adapted H7N9 vaccine virus (A/Anhui/2013, RG268); however, whether and how this mutation affects vaccine potency remain to be investigated. In this study, we serially passaged RG268 in MDCK cells and found that the HA titer and the TCID50 of the passaged virus RG268-M5 were 4-fold (HA units/50 µL) and 3.5-fold (log10 TCID50/mL) higher than those of the original CVV. By inspecting tandem MS spectra, we identified a new glycosylation site at N167 near the receptor binding site of the HA protein of RG268-M5. Flow cytometry results revealed that RG268-M5 could efficiently infect MDCK cells and initiate viral protein replication as well as that of RG268. Though the new glycosylation site is in the antigenic epitope of viral HA protein, the HI assay result indicated that the antigenicity of RG268-M5 was similar to RG268. Additionally, immunizing mice with RG268-M5 mixed aluminum hydroxide could induce potent antibody responses against the homologous and heterologous H7N9 viruses in vitro whereas the titers were comparable with those from the RG268 group. These results provide in-depth structural information regarding the effects of site-specific glycosylation on virus properties, which have implications for novel avian influenza vaccine development.

A TLR9 agonist synergistically enhances protective immunity induced by an Alum-adjuvanted H7N9 inactivated whole-virion vaccine.

Antigen sparing is an important strategy for pandemic vaccine development because of the limitation of worldwide vaccine production during disease outbreaks. However, several clinical studies have demonstrated that the current aluminum (Alum)-adjuvanted influenza vaccines fail to sufficiently enhance immune responses to meet licensing criteria. Here, we used pandemic H7N9 as a model virus to demonstrate that a 10-fold lower amount of vaccine antigen combined with Alum and TLR9 agonist can provide stronger protective effects than using Alum as the sole adjuvant. We found that the Alum/CpG 1018 combination adjuvant could induce more robust virus-specific humoral immune responses, including higher total IgG production, hemagglutination-inhibiting antibody activity, and neutralizing antibody titres, than the Alum-adjuvanted formulation. Moreover, this combination adjuvant shifted the immune response toward a Th1-biased immune response. Importantly, the Alum/CpG 1018-formulated vaccine could confer better protective immunity against H7N9 challenge than that adjuvanted with Alum alone. Notably, the addition of CpG 1018 to the Alum-adjuvanted H7N9 whole-virion vaccine exhibited an antigen-sparing effect without compromising vaccine efficacy. These findings have significant implications for improving Alum-adjuvanted influenza vaccines using the approved adjuvant CpG 1018 for pandemic preparedness.

Correction: DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss.

[This corrects the article DOI: 10.1371/journal.pntd.0009374.].

A DNA vaccine candidate delivered by an electroacupuncture machine provides protective immunity against SARS-CoV-2 infection.

A major challenge in the use of DNA vaccines is efficient DNA delivery in vivo. Establishing a safe and efficient electric transfer method is the key to developing rapid DNA vaccines against emerging infectious diseases. To overcome the complexity of designing new electric transfer machines for DNA delivery, a clinically approved electric transfer machine could be considered as an alternative. Here, we report an electroacupuncture machine-based method for DNA vaccine delivery after intramuscular injection of the COVID-19 DNA vaccine. The S gene of SARS-CoV-2 in the pVAX1 plasmid (pSARS2-S) was used as an antigen in this study. We optimized the clinically used electroacupuncture machine settings for efficient induction of the neutralizing antibody titer after intramuscular injection of pSARS2-S in mice. We found that pSARS2-S immunization at 40 Vpp for 3-5 s could induce high neutralizing antibody titers and Th1-biased immune responses. IFN-γ/TNF-α-secreting CD4+ and CD8+ T cells were also observed in the DNA vaccination group but not in the recombinant protein vaccination group. T-cell epitope mapping shows that the major reactive epitopes were located in the N-terminal domain (a.a. 261-285) and receptor-binding domain (a.a. 352-363). Importantly, pSARS2-S immunization in hamsters could induce protective immunity against SARS-CoV-2 challenge in vivo. In the preclinical toxicology study, blood biochemistry, hematology, and DNA persistence analysis reveal that the DNA delivery method is safe. Furthermore, the raised antisera could also cross-neutralize different variants of concern. These findings suggest that DNA vaccination using an electroacupuncture machine is feasible for use in humans in the future.

Graptopetalum paraguayense Extract Ameliorates Proteotoxicity in Aging and Age-Related Diseases in Model Systems.

Declines in physiological functions are the predominant risk factors for age-related diseases, such as cancers and neurodegenerative diseases. Therefore, delaying the aging process is believed to be beneficial in preventing the onset of age-related diseases. Previous studies have demonstrated that Graptopetalum paraguayense (GP) extract inhibits liver cancer cell growth and reduces the pathological phenotypes of Alzheimer's disease (AD) in patient IPS-derived neurons. Here, we show that GP extract suppresses ß-amyloid pathology in SH-SYS5Y-APP695 cells and APP/PS1 mice. Moreover, AMP-activated protein kinase (AMPK) activity is enhanced by GP extract in U87 cells and APP/PS1 mice. Intriguingly, GP extract enhances autophagy in SH-SYS5Y-APP695 cells, U87 cells, and the nematode Caenorhabditis elegans, suggesting a conserved molecular mechanism by which GP extract might regulate autophagy. In agreement with its role as an autophagy activator, GP extract markedly diminishes mobility decline in polyglutamine Q35 mutants and aged wild-type N2 animals in C. elegans. Furthermore, GP extract significantly extends lifespan in C. elegans.

Assessment of adjuvantation strategy of lipid squalene nanoparticles for enhancing the immunogenicity of a SARS-CoV-2 spike subunit protein against COVID-19.

Vaccination is regarded as the most effective intervention for controlling the coronavirus disease 2019 (COVID-19) pandemic. The objective of this study is to provide comprehensive information on lipid squalene nanoparticle (SQ@NP)-adjuvanted COVID-19 vaccines regarding modulating immune response and enhancing vaccine efficacy. After being adjuvanted with SQ@NP, the SARS-CoV-2 spike (S) subunit protein was intramuscularly (i.m.) administered to mice. Serum samples investigated by ELISA and virus neutralizing assay showed that a single-dose SQ@NP-adjuvanted S-protein vaccine can induce antigen-specific IgG and protective antibodies comparable with those induced by two doses of nonadjuvanted protein vaccine. When the mice received a boosting vaccine injection, anamnestic response was observed in the groups of adjuvanted vaccine. Furthermore, the secretion of cytokines in splenocytes, such as interferon (IFN)-γ, interleukin (IL)-5 and IL-10, was significantly enhanced after adjuvantation of S-protein vaccine with SQ@NP; however, this was not the case for the vaccine adjuvanted with conventional aluminum mineral salts. Histological examination of injection sites showed that the SQ@NP-adjuvanted vaccine was considerably well tolerated following i.m. injection in mice. These results pave the way for the performance tuning of optimal vaccine formulations against COVID-19.

DNA vaccination induced protective immunity against SARS CoV-2 infection in hamsterss.

The development of efficient vaccines against COVID-19 is an emergent need for global public health. The spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a major target for the COVID-19 vaccine. To quickly respond to the outbreak of the SARS-CoV-2 pandemic, a nucleic acid-based vaccine is a novel option, beyond the traditional inactivated virus vaccine or recombinant protein vaccine. Here, we report a DNA vaccine containing the spike gene for delivery via electroporation. The spike genes of SARS-CoV and SARS-CoV-2 were codon optimized for mammalian cell expression and then cloned into mammalian cell expression vectors, called pSARS-S and pSARS2-S, respectively. Spike protein expression was confirmed by immunoblotting after transient expression in HEK293T cells. After immunization, sera were collected for antigen-specific antibody and neutralizing antibody titer analyses. We found that both pSARS-S and pSARS2-S immunization induced similar levels of antibodies against S2 of SARS-CoV-2. In contrast, only pSARS2-S immunization induced antibodies against the receptor-binding domain of SARS-CoV-2. We further found that pSARS2-S immunization, but not pSARS-S immunization, could induce very high titers of neutralizing antibodies against SARS-CoV-2. We further analyzed SARS-CoV-2 S protein-specific T cell responses and found that the immune responses were biased toward Th1. Importantly, pSARS2-S immunization in hamsters could induce protective immunity against SARS-CoV-2 challenge in vivo. These data suggest that DNA vaccination could be a promising approach for protecting against COVID-19.

Development and Evaluation of Vero Cell-Derived Master Donor Viruses for Influenza Pandemic Preparedness.

The embryonated egg-based platform currently produces the majority of seasonal influenza vaccines by employing a well-developed master donor virus (MDV, A/PR/8/34 (PR8)) to generate high-growth reassortants (HGRs) for A/H1N1 and A/H3N2 subtypes. Although the egg-based platform can supply enough seasonal influenza vaccines, it cannot meet surging demands during influenza pandemics. Therefore, multi-purpose platforms are desirable for pandemic preparedness. The Vero cell-based production platform is widely used for human vaccines and could be a potential multi-purpose platform for pandemic influenza vaccines. However, many wild-type and egg-derived influenza viruses cannot grow efficiently in Vero cells. Therefore, it is critical to develop Vero cell-derived high-growth MDVs for pandemic preparedness. In this study, we evaluated two in-house MDVs (Vero-15 and VB5) and two external MDVs (PR8 and PR8-HY) to generate Vero cell-derived HGRs for five avian influenza viruses (AIVs) with pandemic potentials (H5N1 clade 2.3.4, H5N1 clade 2.3.2.1, American-lineage H5N2, H7N9 first wave and H7N9 fifth wave). Overall, no single MDV could generate HGRs for all five AIVs, but this goal could be achieved by employing two in-house MDVs (vB5 and Vero-15). In immunization studies, mice received two doses of Vero cell-derived inactivated H5N1 and H7N9 whole virus antigens adjuvanted with alum and developed robust antibody responses.

Development of high-growth influenza H7N9 prepandemic candidate vaccine viruses in suspension MDCK cells.

BACKGROUND: Influenza vaccine manufacturers traditionally use egg-derived candidate vaccine viruses (CVVs) to produce high-yield influenza viruses for seasonal or pandemic vaccines; however, these egg-derived CVVs need an adaptation process for the virus to grow in mammalian cells. The low yields of cell-based manufacturing systems using egg-derived CVVs remain an unsolved issue. This study aimed to develop high-growth cell-derived CVVs for MDCK cell-based vaccine manufacturing platforms. METHODS: Four H7N9 CVVs were generated in characterized Vero and adherent MDCK (aMDCK) cells. Furthermore, reassortant viruses were amplified in adherent MDCK (aMDCK) cells with certification, and their growth characteristics were detected in aMDCK cells and new suspension MDCK (sMDCK) cells. Finally, the plaque-forming ability, biosafety, and immunogenicity of H7N9 reassortant viruses were evaluated. RESULTS: The HA titers of these CVVs produced in proprietary suspension MDCK (sMDCK) cells and chicken embryos were 2- to 8-fold higher than those in aMDCK cells. All H7N9 CVVs showed attenuated characteristics by trypsin-dependent plaque assay and chicken embryo lethality test. The alum-adjuvanted NHRI-RG5 (derived from the fifth wave H7N9 virus A/Guangdong/SP440/2017) vaccine had the highest immunogenicity and cross-reactivity among the four H7N9 CVVs. Finally, we found that AddaVax adjuvant improved the cross-reactivity of low pathogenic H7N9 virus against highly pathogenic H7N9 viruses. CONCLUSIONS: Our study indicates that cell-derived H7N9 CVVs possessed high growth rate in new sMDCK cells and low pathogenicity in chicken embryo, and that CVVs generated by this platform are also suitable for both cell- and egg-based prepandemic vaccine production.

Assessing the therapeutic potential of Graptopetalum paraguayense on Alzheimer's disease using patient iPSC-derived neurons.

Alzheimer's disease (AD) is the most common type of dementia and also one of the leading causes of death worldwide. However, the underlying mechanisms remain unclear, and currently there is no drug treatment that can prevent or cure AD. Here, we have applied the advantages of using induced pluripotent stem cell (iPSC)-derived neurons (iNs) from AD patients, which are able to offer human-specific drug responsiveness, in order to evaluate therapeutic candidates for AD. Using approach involving an inducible neurogenin-2 transgene, we have established a robust and reproducible protocol for differentiating human iPSCs into glutamatergic neurons. The AD-iN cultures that result have mature phenotypic and physiological properties, together with AD-like biochemical features that include extracellular ß-amyloid (Aß) accumulation and Tau protein phosphorylation. By screening using a gene set enrichment analysis (GSEA) approach, Graptopetalum paraguayense (GP) has been identified as a potential therapeutic agent for AD from among a range of Chinese herbal medicines. We found that administration of a GP extract caused a significantly reduction in the AD-associated phenotypes of the iNs, including decreased levels of extracellular Aß40 and Aß42, as well as reduced Tau protein phosphorylation at positions Ser214 and Ser396. Additionally, the effect of GP was more prominent in AD-iNs compared to non-diseased controls. These findings provide valuable information that suggests moving extracts of GP toward drug development, either for treating AD or as a health supplement to prevent AD. Furthermore, our human iN-based platform promises to be a useful strategy when it is used for AD drug discovery.

Process development for pandemic influenza VLP vaccine production using a baculovirus expression system.

BACKGROUND: Influenza viruses cause hundreds of thousands of respiratory diseases worldwide each year, and vaccination is considered the most effective approach for preventing influenza annual epidemics or pandemics. Since 1950, chicken embryonated eggs have been used as the main method for producing seasonal influenza vaccines. However, this platform has the main drawback of a lack of scale-up flexibility, and thus, egg-based vaccine manufacturers cannot supply sufficient doses within a short period for use for pandemic prevention. As a result, strategies for reducing the manufacturing time and increasing production capacity are urgently needed. Non-virion vaccine methods have been considered an alternative strategy against an influenza pandemic, and the purpose of maintaining an immunogenic capsule structure with infectious properties appears to be met by the virus-like particle (VLP) platform. RESULTS: An influenza H7N9-TW VLP production platform using insect cells, which included the expression of hemagglutinin (HA), NA, and M1 proteins, was established. To scale up H7N9-TW VLP production, several culture conditions were optimized to obtain a higher production yield. A high level of dissolved oxygen (DO) could be critical to H7N9-TW VLP production. If the DO was maintained at a high level, the HA titer obtained in the spinner flask system with ventilation was similar to that obtained in a shake flask. In this study, the HA titer in a 5-L bioreactor with a well-controlled DO level was substantially improved by 128-fold (from 4 HA units (HAU)/50 µL to 512 HAU/50 µL). CONCLUSIONS: In this study, a multigene expression platform and an effective upstream process were developed. Notably, a high H7N9-TW VLP yield was achieved using a two-step production strategy while a high DO level was maintained. The upstream process, which resulted in high VLP titers, could be further used for large-scale influenza VLP vaccine production.

The stability and immunogenicity of inactivated MDCK cell-derived influenza H7N9 viruses.

In recent years, cell-based influenza vaccines have gained a great interest over the egg-based vaccines. Several inactivated H7N9 vaccines have been evaluated in clinical trials, including whole-virion vaccines, split vaccines and subunit vaccines. Recently, we developed a new suspension MDCK (sMDCK) cell line for influenza viruses production. However, the properties of purified antigen from sMDCK cells remain unclear. In this study, the stability of influenza H7N9 vaccine bulk derived from sMDCK cells was investigated, and the data were compared with the vaccine antigen derived from our characterized adhesion MDCK (aMDCK) cells in serum-free medium. The influenza H7N9 bulks derived from sMDCK and aMDCK cells were stored at 2-8 °C for different periods of time, and a number of parameters selected to monitor the H7N9 vaccine antigen stability were evaluated at each interval (1, 3 and 12 months). The monitored parameters included virus morphology, hemagglutinin (HA) activity, HA concentration, antigenicity, and immunogenicity. The sMDCK-derived H7N9 bulk showed similar morphology to that of the aMDCK-derived H7N9 bulk, and there were no obvious changes after the extended storage periods. Furthermore, the HA titer, HA concentration, and antigenicity of sMDCK-derived H7N9 bulk were stable after 28 months of storage. Finally, the results of hemagglutination inhibition and neutralization tests showed that sMDCK- and aMDCK-derived H7N9 vaccines had comparable immunogenicity. These results indicated that sMDCK-derived H7N9 bulk has good stability compared to that of aMDCK-derived H7N9 bulk. Thus, the newly developed suspension MDCK cell line shows a great alternative for manufacturing cell-based influenza vaccines.

Neurohealth Properties of Hericium erinaceus Mycelia Enriched with Erinacines.

Hericium erinaceus, an ideal culinary-medicinal mushroom, has become a well-established candidate in promoting positive brain and nerve health-related activities by inducing the nerve growth factor from its bioactive ingredient. Among its active compounds, only erinacine A has confirmed pharmacological actions in the central nervous system in rats. Hence, this review has summarized the available information on the neurohealth properties of H. erinaceus mycelia enriched with erinacines, which may contribute to further research on the therapeutic roles of these mycelia. The safety of this mushroom has also been discussed. Although it has been difficult to extrapolate the in vivo studies to clinical situations, preclinical studies have shown that there can be improvements in ischemic stroke, Parkinson's disease, Alzheimer's disease, and depression if H. erinaceus mycelia enriched with erinacines are included in daily meals.

The Cyanthin Diterpenoid and Sesterterpene Constituents of Hericium erinaceus Mycelium Ameliorate Alzheimer's Disease-Related Pathologies in APP/PS1 Transgenic Mice.

Hericium erinaceus was used in traditional Chinese medicine for physiologically beneficial medicines. Recently, it has become a candidate in causing positive brain health-related activities. We previously reported that Hericium erinaceus mycelium ameliorates Alzheimer's disease (AD)-related pathologies. To reveal the role of the cyanthin diterpenoid and sesterterpene constituents on this effects, erinacine A and S were isolated and their effects on attenuating AD-related pathology in APPswe/PS1dE9 transgenic mice were investigated. A 30 day short-term administration of erinacine A and S were performed to explore the effect of each erinacine on AD-related pathology including amyloid ß production and degradation, plaque formation, plaque growth, glial activation and neurogenesis deterioration. Our results indicated the benefit effects of both erinacine A and S in cerebrum of APPswe/PS1dE9 mice, including: (1) attenuating cerebral plaque loading by inhibiting plaque growth; (2) diminishing the activation of glial cells; (3) raising the level of insulin degrading enzyme; and (4) promoting hippocampal neurogenesis. Moreover, erinacine A reduced the level of insoluble amyloid ß and C-terminal fragment of amyloid precursor protein which was not mediated by erinacine S. We further performed a long term administration of erinacine A and found that erinacine A recovered the impairment in the tasks including burrowing, nesting, and Morris water maze. Our data pointed out that although both erinacine A and S reduce AD pathology via reducing amyloid deposition and promoting neurogenesis, erinacine A can also inhibit amyloid ß production and is worth to be further developed for AD therapeutic use.

Xuefu Zhuyu decoction ameliorates obesity, hepatic steatosis, neuroinflammation, amyloid deposition and cognition impairment in metabolically stressed APPswe/PS1dE9 mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Metabolic syndrome and vascular dysfunction was suggested to be the risk factors for Alzheimer's disease (AD). Xuefu Zhuyu decoction (XZD) is a traditional Chinese medicine used to treat metabolic syndrome and cardiac-cerebral vascular disease. The effects of XZD on ameliorating metabolic syndrome, amyloid-related pathologies and cognitive impairment in an animal model of AD with metabolic stress was investigated. MATERIALS AND METHOD: The animal model of AD with metabolic stress was created by administrating high-fat diet and a low-dose injection of streptozotocin prior to the appearance of senile plaques in APP/PS1 transgenic mice. The diabesity-associated metabolic changes and AD-related pathological alterations were examined. RESULTS: We found that XZD reduced body weight, insulin and leptin level, HOMA-IR, hepatic triglyceride, serum Aß42 in the metabolic stressed AD animal. XZD also ameliorated oral glucose tolerant, Aß deposition, astrocyte and microglia activation in the vicinity of plaques, and nesting behavior in the metabolic stressed AD animal. CONCLUSION: The results of this study suggest that XZD is able to reduce the peripheral metabolic stress-mediated vascular hypoperfusion, neuroinflammation and AD-related pathology in APP/PS1 mice.

Neuroprotective and Antineuroinflammatory Effects of Hydroxyl-Functionalized Stilbenes and 2-Arylbenzo[b]furans.

The drugs currently used to treat Alzheimer's disease (AD) are limited in the benefits they confer, and no medication has been clearly proven to cure or delay the progression of AD. Most candidate AD drugs are meant to reduce the production, aggregation, and toxicity of amyloid ß (Aß) or to promote Aß clearance. Herein, we demonstrate the efficient synthesis of hydroxyl-functionalized stilbene and 2-arylbenzo[b]furan derivatives and report on the neuroprotective and anti-inflammatory effects of these phenolic compounds in vitro and in an animal model. Structure-activity relationships revealed that the presence of an acrylate group on 2-arylbenzo[b]furan confers neuroprotective and anti-inflammatory effects. Furthermore, compounds 11 and 37 in this study showed particular potential for development as disease-modifying anti-Alzheimer's drugs, based on their neuroprotective effects on neuron cells, their antineuroinflammatory effects on glial cells, and the ability to ameliorate nesting behavior in APP/PS1 mice. These results indicate that 2-arylbenzo[b]furans could be candidate compounds for the treatment of AD.

Erinacine S, a Rare Sesterterpene from the Mycelia of Hericium erinaceus.

A new sesterterpene, erinacine S, and one cyathane diterpene xyloside, erinacine A, were isolated from the ethanol extract of the mycelia of Hericium erinaceus. Their structures were elucidated by spectroscopic and X-ray analysis. A 30-day oral course of erinacines A and S attenuated Aß plaque burden in the brains of 5-month-old female APP/PS1 transgenic mice. Moreover, erinacines A and S significantly increased the level of insulin-degrading enzyme in cerebral cortex.

Impaired cognition and cerebral glucose regulation are associated with astrocyte activation in the parenchyma of metabolically stressed APPswe/PS1dE9 mice.

Although metabolic syndrome was suggested to be a risk factor for Alzheimer's disease (AD), the role of metabolic stress in the initiation of AD pathology remains unclear. In this study, metabolic stress was induced by a high-fat diet and low-dose injection of streptozotocin (HFSTZ) before the appearance of senile plaques in APP/PS1 transgenic mice. We found that, HFSTZ treatment exacerbated amyloid beta burden and astrocyte activation in the vicinity of plaques. Moreover, we observed an upregulation of astrocytic S100B expression in the brain parenchyma of HFSTZ-treated APP/PS1 mice concurrent with increased interleukin-6 expression in cerebral microvascular cells. To determine the impact of HFSTZ treatment on brain function, we performed [(18)F]fludeoxyglucose-positron emission tomography and analyzed nesting behavior. HFSTZ treatment impaired nest construction and cerebral glucose metabolism in several brain regions of APP/PS1 mice during the early stage of AD. These results suggest that HFSTZ-induced peripheral metabolic stress may contribute to vascular inflammation and astrocyte reactivity in the parenchyma and may impair activity of daily living skill and cerebral glucose metabolism in APP/PS1 mice.

Obesity and Hepatic Steatosis Are Associated with Elevated Serum Amyloid Beta in Metabolically Stressed APPswe/PS1dE9 Mice.

Diabesity-associated metabolic stresses modulate the development of Alzheimer's disease (AD). For further insights into the underlying mechanisms, we examine whether the genetic background of APPswe/PS1dE9 at the prodromal stage of AD affects peripheral metabolism in the context of diabesity. We characterized APPswe/PS1dE9 transgenic mice treated with a combination of high-fat diet with streptozotocin (HFSTZ) in the early stage of AD. HFSTZ-treated APPswe/PS1dE9 transgenic mice exhibited worse metabolic stresses related to diabesity, while serum ß-amyloid levels were elevated and hepatic steatosis became apparent. Importantly, two-way analysis of variance shows a significant interaction between HFSTZ and genetic background of AD, indicating that APPswe/PS1dE9 transgenic mice are more vulnerable to HFSTZ treatment. In addition, body weight gain, high hepatic triglyceride, and hyperglycemia were positively associated with serum ß-amyloid, as validated by Pearson's correlation analysis. Our data suggests that the interplay between genetic background of AD and HFSTZ-induced metabolic stresses contributes to the development of obesity and hepatic steatosis. Alleviating metabolic stresses including dysglycemia, obesity, and hepatic steatosis could be critical to prevent peripheral ß-amyloid accumulation at the early stage of AD.

Lithospermic acid attenuates 1-methyl-4-phenylpyridine-induced neurotoxicity by blocking neuronal apoptotic and neuroinflammatory pathways.

BACKGROUND: Parkinson's disease is the second most common neurodegenerative disorders after Alzheimer's disease. The main cause of the disease is the massive degeneration of dopaminergic neurons in the substantia nigra. Neuronal apoptosis and neuroinflammation are thought to be the key contributors to the neuronal degeneration. RESULTS: Both CATH.a cells and ICR mice were treated with 1-methyl-4-phenylpyridin (MPP(+)) to induce neurotoxicity in vitro and in vivo. Western blotting and immunohistochemistry were also used to analyse neurotoxicity, neuroinflammation and aberrant neurogenesis in vivo. The experiment in CATH.a cells showed that the treatment of MPP(+) impaired intake of cell membrane and activated caspase system, suggesting that the neurotoxic mechanisms of MPP(+) might include both necrosis and apoptosis. Pretreatment of lithospermic acid might prevent these toxicities. Lithospermic acid possesses specific inhibitory effect on caspase 3. In mitochondria, MPP(+) caused mitochondrial depolarization and induced endoplasmic reticulum stress via increasing expression of chaperone protein, GRP-78. All the effects mentioned above were reduced by lithospermic acid. In animal model, the immunohistochemistry of mice brain sections revealed that MPP(+) decreased the amount of dopaminergic neurons, enhanced microglia activation, promoted astrogliosis in both substantia nigra and hippocampus, and MPP(+) provoked the aberrant neurogenesis in hippocampus. Lithospermic acid significantly attenuates all of these effects induced by MPP(+). CONCLUSIONS: Lithospermic acid is a potential candidate drug for the novel therapeutic intervention on Parkinson's disease.