Laser ablation-inductively coupled plasma-mass spectrometry imaging of white and gray matter iron distribution in Alzheimer's disease frontal cortex.

Iron deposition in the brain is a feature of normal aging, though in several neurodegenerative disorders, including Alzheimer's disease, the rate of iron accumulation is more advanced than in age-matched controls. Using laser ablation-inductively coupled plasma-mass spectrometry imaging we present here a pilot study that quantitatively assessed the iron content of white and gray matter in paraffin-embedded sections from the frontal cortex of Alzheimer's and control subjects. Using the phosphorus image as a confirmed proxy for the white/gray matter boundary, we found that increased intrusion of iron into gray matter occurs in the Alzheimer's brain compared to controls, which may be indicative of either a loss of iron homeostasis in this vulnerable brain region, or provide evidence of increased inflammatory processes as a response to chronic neurodegeneration. We also observed a trend of increasing iron within the white matter of the frontal cortex, potentially indicative of disrupted iron metabolism preceding loss of myelin integrity. Considering the known potential toxicity of excessive iron in the brain, our results provide supporting evidence for the continuous development of novel magnetic resonance imaging approaches for assessing white and gray matter iron accumulation in Alzheimer's disease.

Identification of a Novel GRHL3/HOPX/Wnt/ß-Catenin Proto-oncogenic Axis in Squamous Cell Carcinoma of the Esophagus.

BACKGROUND & AIMS: Esophageal squamous cell carcinoma (ESCC) is an aggressive malignancy with a poor long-term prognosis. The molecular mechanisms underlying the initiation and progression of this tumor are largely unknown. The transcription factor GRHL3 functions as a potent tumor suppressor in SCC of skin, head, and neck. This study aims to determine whether GRHL3 also plays a role in the homeostasis of the esophageal epithelium and in the development of ESCC. METHODS: The effects of Grhl3 deletion on squamous epithelial homeostasis in embryos and adult mice were examined using immunohistochemistry, transmission electron microscopy, and real-time polymerase chain reaction. The conditionally deleted mice were subsequently used to determine susceptibility to ESCC. Whole-transcriptome sequencing (RNA-seq) was performed on ESCC in wild-type and Grhl3 deleted animals. To decipher the signaling pathways, real-time polymerase chain reaction, immunohistochemistry, analysis of chromatin immunoprecipitation sequencing, chromatin immunoprecipitation-polymerase chain reaction, and RNA seq datasets were used. Primary human samples were used to validate the findings in the mouse model. RESULTS: Loss of Grhl3 perturbs the proliferation-differentiation balance in the esophageal epithelium, thereby increasing the susceptibility to esophageal carcinogenesis in adult mice. Grhl3 imparts its tumor suppressor function by regulating the expression of HOPX. We have identified the Wnt/ß-catenin pathway as the downstream effectors of GRHL3 and HOPX through our integrated approach using patient-derived ESCC samples and mouse models. CONCLUSIONS: GRHL3 conveys its tumor suppressor function in ESCC through regulating its target gene HOPX, which limits Wnt/ß-catenin signaling. Targeted therapies to inhibit this pathway could be a potential treatment strategy for ESCC patients with reduced GRHL3 expression.

Th-1 cytotoxic cell-mediated response predominates in the tracheal mucosa following Mycoplasma synoviae infection of MS-H-vaccinated chickens.

Mycoplasma synoviae is a pathogen of poultry that causes upper respiratory tract disease. MS-H is a live attenuated temperature-sensitive vaccine that effectively control M. synoviae infection in chickens. However, the mechanisms underpinning protection have not been described previously. In this study, specific-pathogen-free chickens were vaccinated at 3 weeks of age with MS-H vaccine and challenged with field strain M. synoviae 94011/v-18d at 6 weeks of age. Tracheal mucosal inflammation was characterised by the assessment of thickness, histopathological lesions, cellular infiltrates and cytokine transcription. Tracheal lesion scores of unvaccinated-challenged (-V+C) birds were higher than that of vaccinated-challenged (+V+C) birds. +V+C birds displayed early upregulation of IL-4, consistent with a Th-2-skewed response, followed by a later increase in IFN-γ transcription, indicating transition to a Th-1-skewed response. -V+C birds displayed a concurrent early Th-2 and Th-17 response characterised by increase expression of IL-4 and IL-17A respectively, and late T regulatory response characterised by increased IL-10 transcription. +V+C chickens had more cytotoxic T cells (CD8+ T cells) at 7- and 21 days post-challenge (dpc), while -V+C chickens had higher numbers of infiltrating CD4+CD25+ at 7 and 21 dpc. Overall, these observations suggest that the immune response in +V+C chickens had an inflammation characterised by an early Th-2 skewed response followed closely by a Th-1 response and infiltration of cytotoxic T cells, while the response in -V+C chickens was an early Th-2/Th-17-skewed response closely followed by a T regulatory response.

Tracheal cellular immune response in chickens inoculated with Mycoplasma synoviae vaccine, MS-H or its parent strain 86079/7NS.

Mycoplasma synoviae causes respiratory tract disease in chickens characterised by mild to moderate lymphoplasmacytic infiltration of the tracheal mucosa. MS-H (Vaxsafe1 MS, Bioproperties Pty Ltd.) is an effective live attenuated vaccine for M. synoviae, but the immunological basis for its mechanism of protection has not been investigated, and the phenotypes of lymphocytes and associated cytokines involved in the local adaptive immune response have not been described previously. In this study, specific-pathogen-free chickens were inoculated intra-ocularly at 3 weeks of age with either M. synoviae vaccine strain MS-H or vaccine parent strain 86079/7NS (7NS), or remained uninoculated. At 2-, 7- and 21 days post-inoculation (dpi), tracheal mucosal pathology, infiltrating lymphocytes subsets and transcription levels of mRNA encoding 8 cytokines were assessed using light microscopy, indirect immunofluorescent staining and RT-qPCR, respectively. After inoculation, tracheal mucosal thickness, tracheal mucosal lesions, and numbers of infiltrating CD4+CD25- cells, B-cells, and macrophages were greater in MS-H- and 7NS-inoculated chickens compared with non-inoculated. Inoculation with 7NS induced up-regulation of IFN-γ, while vaccination with MS-H induced up-regulation of IL-17A, when compared with non-inoculated birds. Both inoculated groups had a moderate infiltrate of CD4+CD25+ T cells in the tracheal mucosa. These findings reveal that the tracheal local cellular response after MS-H inoculation is dominated by a Th-17 response, while that of 7NS-inoculated chickens is dominated by a Th-1 type response.

S-Adenosylmethionine Rescues Cognitive Deficits in the rTg4510 Animal Model by Stabilizing Protein Phosphatase 2A and Reducing Phosphorylated Tau.

BACKGROUND: Alterations in the methionine cycle and abnormal tau phosphorylation are implicated in many neurodegenerative diseases, including Alzheimer's disease and frontotemporal dementia. rTg4510 mice express mutant human P301L tau and are a model of tau hyperphosphorylation. The cognitive deficit seen in these animals correlates with a burden of hyperphosphorylated tau and is a model to test therapies aimed at lowering phosphorylated tau. OBJECTIVE: This study aimed to increase protein phosphatase 2A activity through supplementation of S-adenosylmethionine and analyze the effect on spatial memory and tau in treated animals. METHODS: 6-month-old rTg4510 mice were treated with 100 mg/kg S-adenosylmethionine by oral gavage for 3 weeks. Spatial recognition memory was tested in the Y-maze. Alterations to phosphorylated tau and protein phosphatase 2A were explored using immunohistochemistry, western blot, and enzyme-linked immunosorbent assays. RESULTS: Treatment with S-adenosylmethionine increased the Y-maze novel arm exploration time and increased both the expression and activity of protein phosphatase 2A. Furthermore, treatment reduced the number of AT8 positive neurons and reduced the expression of phosphorylated tau (Ser202/Thr205). S-adenosylmethionine contributes to multiple pathways in neuronal homeostasis and neurodegeneration. CONCLUSION: This study shows that supplementation with S-adenosylmethionine stabilizes the heterotrimeric form of PP2A resulting in an increase the enzymatic activity, a reduced level of pathological tau, and improved cognition.

Plant poisoning leads to alpha-synucleinopathy and neuromelanopathy in kangaroos.

The pathogenesis of synucleinopathies, common neuropathological lesions normally associated with some human neurodegenerative disorders such as Parkinson's disease, dementia with Lewy bodies and multiple system atrophy, remains poorly understood. In animals, ingestion of the tryptamine-alkaloid-rich phalaris pastures plants causes a disorder called Phalaris staggers, a neurological syndrome reported in kangaroos. The aim of the study was to characterise the clinical and neuropathological changes associated with spontaneous cases of Phalaris staggers in kangaroos. Gross, histological, ultrastructural and Immunohistochemical studies were performed to demonstrate neuronal accumulation of neuromelanin and aggregated α-synuclein. ELISA and mass spectrometry were used to detect serum-borne α-synuclein and tryptamine alkaloids respectively. We report that neurons in the central and enteric nervous systems of affected kangaroos display extensive accumulation of neuromelanin in the perikaryon without affecting neuronal morphology. Ultrastructural studies confirmed the typical structure of neuromelanin. While we demonstrated strong staining of α-synuclein, restricted to neurons, intracytoplasmic Lewy bodies inclusions were not observed. α-synuclein aggregates levels were shown to be lower in sera of the affected kangaroos compared to unaffected herd mate kangaroos. Finally, mass spectrometry failed to detect the alkaloid toxins in the sera derived from the affected kangaroos. Our preliminary findings warrant further investigation of Phalaris staggers in kangaroos, potentially a valuable large animal model for environmentally-acquired toxic synucleinopathy.

Role of serotonin-1A receptors in the action of antipsychotic drugs: comparison of prepulse inhibition studies in mice and rats and relevance for human pharmacology.

This study aimed to explore strain and species differences in the involvement of 5-HT1A receptors in the action of antipsychotic drugs, using prepulse inhibition (PPI), a model of sensory processing which is deficient in schizophrenia patients. We used automated startle boxes to compare the effect of the 5-HT1A receptor agonist, (+/-)-8-hydroxy-dipropyl-amino-tetralin (8-OH-DPAT), on PPI in three mouse strains. Balb/c mice were then pretreated with antipsychotics, treated with 8-OH-DPAT or saline, and tested for PPI. 8-OH-DPAT treatment dose dependently increased PPI in Balb/c mice, but had less effect in 129Sv and C57Bl/6 mice. In Balb/c mice, the effect of 8-OH-DPAT was blocked by the typical antipsychotic and dopamine D2 receptor antagonist, haloperidol and the third generation antipsychotic, aripiprazole, which has activity at both 5-HT1A and dopamine D2 receptors. The atypical antipsychotics, clozapine, olanzapine and risperidone, had lesser effects. Similar to our earlier studies in rats, the present PPI results suggest that 5-HT1A receptors are involved in the action of some antipsychotic drugs in mice. Despite strain and species differences in the magnitude and direction of the effect of 8-OH-DPAT, downstream dopamine D2 receptor activation seems to be an important mediator. These comparative results allow a theoretical framework of receptor interactions, which may guide further studies on the involvement of 5-HT1A receptors in schizophrenia.