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Prions are proteinaceous infectious particles that replicate by structural conversion of the host-encoded cellular prion protein (PrPC), causing fatal neurodegenerative diseases in mammals. Species-specific amino acid substitutions (AAS) arising from single nucleotide polymorphisms within the prion protein gene (Prnp) modulate prion disease pathogenesis, and, in several instances, reduce susceptibility of homo- or heterozygous AAS carriers to prion infection. However, a mechanistic understanding of their protective effects against clinical disease is missing. We generated gene-targeted mouse infection models of chronic wasting disease (CWD), a highly contagious prion disease of cervids. These mice express wild-type deer or PrPC harboring the S138N substitution homo- or heterozygously, a polymorphism found exclusively in reindeer (Rangifer tarandus spp.) and fallow deer (Dama dama). The wild-type deer PrP-expressing model recapitulated CWD pathogenesis including fecal shedding. Encoding at least one 138N allele prevented clinical CWD, accumulation of protease-resistant PrP (PrPres) and abnormal PrP deposits in the brain tissue. However, prion seeding activity was detected in spleens, brains, and feces of these mice, suggesting subclinical infection accompanied by prion shedding. 138N-PrPC was less efficiently converted to PrPres in vitro than wild-type deer (138SS) PrPC. Heterozygous coexpression of wild-type deer and 138N-PrPC resulted in dominant-negative inhibition and progressively diminished prion conversion over serial rounds of protein misfolding cyclic amplification. Our study indicates that heterozygosity at a polymorphic Prnp codon can confer the highest protection against clinical CWD and highlights the potential role of subclinical carriers in CWD transmission.

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Adrenergic stimuli are important for corneal epithelial structure and healing. The purpose of the present study was to examine the hypothesis that the lack of a single α1-adrenoceptor (α1-AR) subtype affects corneal epithelial thickness and cell proliferation. Expression levels of α1-AR mRNA were determined in mouse cornea using real-time PCR. In mice devoid of one of the three α1-AR subtypes (α1A-AR-/-, α1B-AR-/-, α1D-AR-/-) and in wild-type controls, thickness of individual corneal layers, the number of epithelial cell layers, and average epithelial cell size were determined in cryosections. Endothelial cell density and morphology were calculated in corneal explants, and epithelial cell proliferation rate was determined with immunofluorescence microscopy. Moreover, the ultrastructure of the corneal epithelium was examined by transmission electron microscopy. Messenger RNA for all three α1-AR subtypes was expressed in whole cornea and in corneal epithelium from wild-type mice with a rank order of abundance of α1A ≥ α1B > α1D. In contrast, no α1-AR mRNA was detected in the stroma, and only α1B-AR mRNA was found in the Descemet endothelial complex. Remarkably, corneal epithelial thickness and mean epithelial cell size were reduced in α1A-AR-/- mice. Our findings suggest that the α1A-AR exerts growth effects in mouse corneal epithelial cells.

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An increased number of tumor-associated macrophages (TAMs) that exhibit the M2 macrophage phenotype is related to poorer prognosis in cancer patients. MafB is a transcription factor regulating the differentiation of macrophages. However, involvement of MafB for the development of TAMs is unknown. This study was designed to investigate the role of MafB in a murine urethane-induced lung cancer model. Urethane was injected intraperitoneally into wild-type and dominant-negative MafB transgenic mice. Twenty-four weeks later, mice were sacrificed and their lungs removed for pathological analysis. The numbers and mean areas of lung cancer were evaluated. In addition, the numbers of Mac-3-positive macrophages were evaluated in each tumor. The numbers and mean areas of lung cancer induced by urethane administration were not significantly different between wild-type and dominant-negative MafB transgenic mice. The numbers of TAMs in lung cancer tissue were not significantly different between the two groups. MafB silencing using dominant-negative MafB did not influence the initiation and growth of lung cancer in mice exposed to urethane. These data suggest that MafB may not be related to the development of TAMs.

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BACKGROUND: Chemokine C-C motif ligand 1 (CCL1) accumulates C-C motif chemokine receptor 8 positive leukocytes to the inflammatory sites. Single-nucleotide polymorphisms in the chemokine CCL1 gene are associated with exacerbation of chronic obstructive lung disease. However, it is unclear whether CCL1 has immunomodulatory functions during pulmonary inflammation. This study aimed to elucidate this issue using newly generated transgenic mice that express CCL1 in the lungs (SPC-CCL1 mice). METHODS: To evaluate the phenotypes of these mice, lung section and bronchoalveolar lavage (BAL) fluid analyses were performed. We intratracheally administered lipopolysaccharide (LPS) or Mycobacterium bovis as a model of acute or chronic lung inflammation, respectively. RESULTS: No histological differences were observed between lung tissue from SPC-CCL1 Tg and wild-type mice in the resting condition and after LPS administration. In the resting condition, the total BAL cell concentration was lower in SPC-CCL1 Tg mice than in wild-type mice (P = 0.0097). Flow cytometric analyses showed that SPC-CCL1 Tg mice had fewer F4/80-positive cells than wild-type mice (P = 0.0278). After intratracheal LPS administration, CCL1 overexpression changed neither the total numbers nor population of BAL cells. After mycobacterial administration, pulmonary granuloma formation was significantly enhanced. The degree of Immunostaining for endoplasmic reticulum to nucleus signaling 1, a molecule associated with granuloma formation and endoplasmic reticulum stress, was significantly enhanced in the granuloma regions of SPC-CCL1 mice treated with Mycobacterium, compared to those of wild-type mice. CONCLUSIONS: CCL1 overexpression in the lungs did not change the acute inflammatory response induced by LPS, but enhanced granuloma formation after mycobacterial treatment, possibly through enhancing endoplasmic reticulum stress.

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Mutations in the LRRK2 gene represent the most common genetic cause of late onset Parkinson's disease. The physiological and pathological roles of LRRK2 are yet to be fully determined but evidence points towards LRRK2 mutations causing a gain in kinase function, impacting on neuronal maintenance, vesicular dynamics and neurotransmitter release. To explore the role of physiological levels of mutant LRRK2, we created knock-in (KI) mice harboring the most common LRRK2 mutation G2019S in their own genome. We have performed comprehensive dopaminergic, behavioral and neuropathological analyses in this model up to 24months of age. We find elevated kinase activity in the brain of both heterozygous and homozygous mice. Although normal at 6months, by 12months of age, basal and pharmacologically induced extracellular release of dopamine is impaired in both heterozygous and homozygous mice, corroborating previous findings in transgenic models over-expressing mutant LRRK2. Via in vivo microdialysis measurement of basal and drug-evoked extracellular release of dopamine and its metabolites, our findings indicate that exocytotic release from the vesicular pool is impaired. Furthermore, profound mitochondrial abnormalities are evident in the striatum of older homozygous G2019S KI mice, which are consistent with mitochondrial fission arrest. We anticipate that this G2019S mouse line will be a useful pre-clinical model for further evaluation of early mechanistic events in LRRK2 pathogenesis and for second-hit approaches to model disease progression.

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Alveolar macrophages (AMs) play important roles in the pathogenesis of chronic obstructive pulmonary disease (COPD). We previously demonstrated upregulation of the transcription factor MafB in AMs of mice exposed to cigarette smoke. The aim of this study was to elucidate the roles of MafB in the development of pulmonary emphysema. Porcine pancreatic elastase was administered to wild-type (WT) and dominant-negative (DN)-MafB transgenic (Tg) mice in which MafB activity was suppressed only in macrophages. We measured the mean linear intercept and conducted cell differential analysis of bronchoalveolar lavage (BAL) cells, surface marker analysis using flow cytometry, and immunohistochemical staining using antibodies to matrix metalloproteinase (MMP)-9 and MMP-12. Airspace enlargement of the lungs was suppressed significantly in elastase-treated DN-MafB Tg mice compared with treated WT mice. AMs with projected pseudopods were decreased in DN-MafB Tg mice. The number of cells intermediately positive for F4/80 and weakly or intermediately positive for CD11b, which are considered cell subsets of matured AMs, decreased in the BAL of DN-MafB Tg mice. Furthermore, MMP-9 and -12 were significantly downregulated in BAL cells of DN-MafB Tg mice. Because MMPs exacerbate emphysema, MafB may be involved in pulmonary emphysema development through altered maturation of macrophages and MMP expression.

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