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Farizatto K.L.G.,NC Biotechnology Center | Farizatto K.L.G.,University of Sao Paulo | Ikonne U.S.,NC Biotechnology Center | Ikonne U.S.,A.T. Still University | And 3 more authors.
PLoS ONE | Year: 2017

Impaired protein clearance likely increases the risk of protein accumulation disorders including Alzheimer’s disease (AD). Protein degradation through the proteasome pathway decreases with age and in AD brains, and the Aβ42 peptide has been shown to impair proteasome function in cultured cells and in a cell-free model. Here, Aβ42 was studied in brain tissue to measure changes in protein clearance pathways and related secondary pathology. Oligomerized Aβ42 (0.5–1.5 μM) reduced proteasome activity by 62% in hippocampal slice cultures over a 4-6-day period, corresponding with increased tau phosphorylation and reduced synaptophysin levels. Interestingly, the decrease in proteasome activity was associated with a delayed inverse effect, >2-fold increase, regarding lysosomal cathepsin B (CatB) activity. The CatB enhancement did not correspond with the Aβ42-mediated phos-pho-tau alterations since the latter occurred prior to the CatB response. Hippocampal slices treated with the proteasome inhibitor lactacystin also exhibited an inverse effect on CatB activity with respect to diminished proteasome function. Lactacystin caused earlier CatB enhancement than Aβ42, and no correspondence was evident between up-regulated CatB levels and the delayed synaptic pathology indicated by the loss of pre- and postsynaptic markers. Contrasting the inverse effects on the proteasomal and lysosomal pathways by Aβ42 and lactacystin, such were not found when CatB activity was up-regulated two-fold with Z-Phe-Ala-diazomethylketone (PADK). Instead of an inverse decline, proteasome function was increased marginally in PADK-treated hippocampal slices. Unexpectedly, the proteasomal augmentation was significantly pronounced in Aβ42-compromised slices, while absent in lactacystin-treated tissue, resulting in >2-fold improvement for nearly complete recovery of proteasome function by the CatB-enhancing compound. The PADK treatment also reduced Aβ42-mediated tau phosphorylation and synaptic marker declines, corresponding with the positive modulation of both proteasome activity and the lysosomal CatB enzyme. These findings indicate that proteasomal stress contributes to AD-type pathogenesis and that governing such pathology occurs through crosstalk between the two protein clearance pathways. © 2017 Farizatto et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


Lozoya O.A.,North Carolina State University | Lozoya O.A.,NC Biotechnology Center | Wauthier E.,North Carolina State University | Wauthier E.,NC Biotechnology Center | And 9 more authors.
Biomaterials | Year: 2011

Human livers have maturational lineages of cells within liver acini, beginning periportally in stem cell niches, the canals of Hering, and ending in polyploid hepatocytes pericentrally and cholangiocytes in bile ducts. Hepatic stem cells (hHpSCs) in vivo are partnered with mesenchymal precursors to endothelia (angioblasts) and stellate cells, and reside in regulated microenvironments, stem cell niches, containing hyaluronans (HA). The in vivo hHpSC niche is modeled in vitro by growing hHpSC in two-dimensional (2D) cultures on plastic. We investigated effects of 3D microenvironments, mimicking the liver's stem cell niche, on these hHpSCs by embedding them in HA-based hydrogels prepared with Kubota's Medium (KM), a serum-free medium tailored for endodermal stem/progenitors. The KM-HA hydrogels mimicked the niches, matched diffusivity of culture medium, exhibited shear thinning and perfect elasticity under mechanical loading, and had predictable stiffness depending on their chemistry. KM-HA hydrogels, which supported cell attachment, survival and expansion of hHpSC colonies, induced transition of hHpSC colonies towards stable heterogeneous populations of hepatic progenitors depending on KM-HA hydrogel stiffness, as shown by both their gene and protein expression profile. These acquired phenotypes did not show morphological evidence of fibrotic responses. In conclusion, this study shows that the mechanical properties of the microenvironment can regulate differentiation in endodermal stem cell populations. © 2011 Elsevier Ltd.


Sun D.,Brookhaven National Laboratory | Stadler A.L.,Brookhaven National Laboratory | Gurevich M.,Brookhaven National Laboratory | Palma E.,Brookhaven National Laboratory | And 3 more authors.
Nanoscale | Year: 2012

Heterogeneous nanoclusters with trimeric and core-shell architectures containing nanoparticles of different size and composition have been fabricated via site-specific PNA-"invasion" of DNA double helix. This novel strategy facilitates the incorporation of double-stranded DNA into the nanoparticle assembly design. This journal is © 2012 The Royal Society of Chemistry.


Zhang Y.,Brookhaven National Laboratory | Lu F.,Brookhaven National Laboratory | Van Der Lelie D.,Brookhaven National Laboratory | Van Der Lelie D.,NC Biotechnology Center | Gang O.,Brookhaven National Laboratory
Physical Review Letters | Year: 2011

The phase behavior of 3D assemblies of nanocubes in a ligand-rich solution upon solvent evaporation was experimentally investigated using small-angle x-ray scattering and electron microscopy. We observed a continuous transformation of assemblies between simple cubic and rhombohedral phases, where a variable angle of rhombohedral structure is determined by ligand thickness. We established a quantitative relationship between the particle shape evolution from cubes to quasispheres and the lattice distortion during the transformation, with a pathway exhibiting the highest known packing. © 2011 American Physical Society.


Zhang Y.,Brookhaven National Laboratory | Lu F.,Brookhaven National Laboratory | Yager K.G.,Brookhaven National Laboratory | Van Der Lelie D.,NC Biotechnology Center | Gang O.,Brookhaven National Laboratory
Nature Nanotechnology | Year: 2013

Nanoparticles coated with DNA molecules can be programmed to self-assemble into three-dimensional superlattices. Such superlattices can be made from nanoparticles with different functionalities and could potentially exploit the synergetic properties of the nanoscale components. However, the approach has so far been used primarily with single-component systems. Here, we report a general strategy for the creation of heterogeneous nanoparticle superlattices using DNA and carboxylic-based conjugation. We show that nanoparticles with all major types of functionality - plasmonic (gold), magnetic (Fe 2 O 3), catalytic (palladium) and luminescent (CdSe/Te@ZnS and CdSe@ZnS) - can be incorporated into binary systems in a rational manner. We also examine the effect of nanoparticle characteristics (including size, shape, number of DNA per particle and DNA flexibility) on the phase behaviour of the heterosystems, and demonstrate that the assembled materials can have novel optical and field-responsive properties.


Wu R.R.,VA Health System | Wu R.R.,Duke University | Orlando L.A.,Duke University | Himmel T.L.,Duke University | And 7 more authors.
BMC Family Practice | Year: 2013

Background: Family health history (FHH) is the single strongest predictor of disease risk and yet is significantly underutilized in primary care. We developed a patient facing FHH collection tool, MeTree©, that uses risk stratification to generate clinical decision support for breast cancer, colorectal cancer, ovarian cancer, hereditary cancer syndromes, and thrombosis. Here we present data on the experience of patients and providers after integration of MeTree© into 2 primary care practices. Methods. This was a Type 2 hybrid controlled implementation-effectiveness study in 3 community-based primary care clinics in Greensboro, NC. All non-adopted adult English speaking patients with upcoming routine appointments were invited. Patients were recruited from December 2009 to the present and followed for one year. Ease of integration of MeTree© into clinical practice at the two intervention clinics was evaluated through patient surveys after their appointment and at 3 months post-visit, and physician surveys 3 months after tool integration. Results: Total enrollment =1,184. Average time to complete MeTree© = 27 minutes. Patients found MeTree©: easy to use (93%), easy to understand (97%), useful (98%), raised awareness of disease risk (85%), and changed how they think about their health (86%). Of the 26% (N = 311) asking for assistance to complete the tool, age (65 sd 9.4 vs. 57 sd 11.8, p-value < 0.00) and large pedigree size (24.4 sd 9.81 vs. 22.2 sd 8.30, p-value < 0.00) were the only significant factors; 77% of those requiring assistance were over the age of 60. Providers (N = 14) found MeTree©: improved their practice (86%), improved their understanding of FHH (64%), made practice easier (79%), and worthy of recommending to their peers (93%). Conclusions: Our study shows that MeTree © has broad acceptance and support from both patients and providers and can be implemented without disruption to workflow. © 2013 Wu et al.; licensee BioMed Central Ltd.


Carpino G.,Foro Italico University of Rome | Cardinale V.,University of Rome La Sapienza | Onori P.,University of L'Aquila | Franchitto A.,University of Rome La Sapienza | And 11 more authors.
Journal of Anatomy | Year: 2012

Stem/progenitors have been identified intrahepatically in the canals of Hering and extrahepatically in glands of the biliary tree. Glands of the biliary tree (peribiliary glands) are tubulo-alveolar glands with mucinous and serous acini, located deep within intrahepatic and extrahepatic bile ducts. We have shown that biliary tree stem/progenitors (BTSCs) are multipotent, giving rise in vitro and in vivo to hepatocytes, cholangiocytes or pancreatic islets. Cells with the phenotype of BTSCs are located at the bottom of the peribiliary glands near the fibromuscular layer. They are phenotypically heterogeneous, expressing transcription factors as well as surface and cytoplasmic markers for stem/progenitors of liver (e.g. SOX9/17), pancreas (e.g. PDX1) and endoderm (e.g. SOX17, EpCAM, NCAM, CXCR4, Lgr5, OCT4) but not for mature markers (e.g. albumin, secretin receptor or insulin). Subpopulations co-expressing liver and pancreatic markers (e.g. PDX1 +/SOX17 +) are EpCAM +/-, and are assumed to be the most primitive of the BTSC subpopulations. Their descendants undergo a maturational lineage process from the interior to the surface of ducts and vary in the mature cells generated: pancreatic cells in hepatopancreatic ducts, liver cells in large intrahepatic bile ducts, and bile duct cells along most of the biliary tree. We hypothesize that there is ongoing organogenesis throughout life, with BTSCs giving rise to hepatic stem cells in the canals of Hering and to committed progenitors within the pancreas. The BTSCs are likely to be central to normal tissue turnover and injury repair and to be key elements in the pathophysiology of liver, pancreas and biliary tree diseases, including oncogenesis. © 2011 The Authors. Journal of Anatomy © 2011 Anatomical Society.


Wu R.R.,Health Services Research and Development | Wu R.R.,Duke University | Himmel T.L.,Duke University | Buchanan A.H.,Duke University | And 6 more authors.
BMC Family Practice | Year: 2014

Background: Studies have shown that the quality of family health history (FHH) collection in primary care is inadequate to assess disease risk. To use FHH for risk assessment, collected data must have adequate detail. To address this issue, we developed a patient facing FHH assessment tool, MeTree. In this paper we report the content and quality of the FHH collected using MeTree. Methods. Design: A hybrid implementation-effectiveness study. Patients were recruited from 2009 to 2012. Setting: Two community primary care clinics in Greensboro, NC. Participants: All non-adopted adult English speaking patients with upcoming appointments were invited to participate. Intervention: Education about and collection of FHH with entry into MeTree. Measures: We report the proportion of pedigrees that were high-quality. High-quality pedigrees are defined as having all the following criteria: (1) three generations of relatives, (2) relatives' lineage, (3) relatives' gender, (4) an up-to-date FHH, (5) pertinent negatives noted, (6) age of disease onset in affected relatives, and for deceased relatives, (7) the age and (8) cause of death (Prim Care 31:479-495, 2004.). Results: Enrollment: 1,184. Participant demographics: age range 18-92 (mean 58.8, SD 11.79), 56% male, and 75% white. The median pedigree size was 21 (range 8-71) and the FHH entered into MeTree resulted in a database of 27,406 individuals. FHHs collected by MeTree were found to be high quality in 99.8% (N = 1,182/1,184) as compared to <4% at baseline. An average of 1.9 relatives per pedigree (range 0-50, SD 4.14) had no data reported. For pedigrees where at least one relative has no data (N = 497/1,184), 4.97 relatives per pedigree (range 1-50, SD 5.44) had no data. Talking with family members before using MeTree significantly decreased the proportion of relatives with no data reported (4.98% if you talked to your relative vs. 10.85% if you did not, p-value < 0.001.). Conclusion: Using MeTree improves the quantity and quality of the FHH data that is collected and talking with relatives prior to the collection of FHH significantly improves the quantity and quality of the data provided. This allows more patients to be accurately risk stratified and offered appropriate preventive care guided by their risk level. Trial number. NCT01372553. © 2014 Wu et al.; licensee BioMed Central Ltd.


PubMed | University of Rome La Sapienza, NC Biotechnology Center and Foro Italico University of Rome
Type: | Journal: Stem cells international | Year: 2016

Niches containing stem/progenitor cells are present in different anatomical locations along the human biliary tree and within liver acini. The most primitive stem/progenitors, biliary tree stem/progenitor cells (BTSCs), reside within peribiliary glands located throughout large extrahepatic and intrahepatic bile ducts. BTSCs are multipotent and can differentiate towards hepatic and pancreatic cell fates. These niches matrix chemistry and other characteristics are undefined. Canals of Hering (bile ductules) are found periportally and contain hepatic stem/progenitor cells (HpSCs), participating in the renewal of small intrahepatic bile ducts and being precursors to hepatocytes and cholangiocytes. The niches also contain precursors to hepatic stellate cells and endothelia, macrophages, and have a matrix chemistry rich in hyaluronans, minimally sulfated proteoglycans, fetal collagens, and laminin. The microenvironment furnishes key signals driving HpSC activation and differentiation. Newly discovered third niches are pericentral within hepatic acini, contain Axin2+ unipotent hepatocytic progenitors linked on their lateral borders to endothelia forming the central vein, and contribute to normal turnover of mature hepatocytes. Their relationship to the other stem/progenitors is undefined. Stem/progenitor niches have important implications in regenerative medicine for the liver and biliary tree and in pathogenic processes leading to diseases of these tissues.


PubMed | University of Rome La Sapienza, NC Biotechnology Center and Foro Italico University of Rome
Type: Journal Article | Journal: Journal of anatomy | Year: 2016

Pancreatic duct glands (PDGs) are tubule-alveolar glands associated with the pancreatic duct system and can be considered the anatomical counterpart of peribiliary glands (PBGs) found within the biliary tree. Recently, we demonstrated that endodermal precursor niches exist fetally and postnatally and are composed functionally of stem cells and progenitors within PBGs and of committed progenitors within PDGs. Here we have characterized more extensively the anatomy of human PDGs as novel niches containing cells with multiple phenotypes of committed progenitors. Human pancreata (n=15) were obtained from cadaveric adult donors. Specimens were processed for histology, immunohistochemistry and immunofluorescence. PDGs were found in the walls of larger pancreatic ducts (diameters >300m) and constituted nearly 4% of the duct wall area. All of the cells identified were negative for nuclear expression of Oct4, a pluripotency gene, and so are presumably committed progenitors and not stem cells. In the main pancreatic duct and in large interlobular ducts, Sox9(+) cells represented 5-30% of the cells within PDGs and were located primarily at the bottom of PDGs, whereas rare and scattered Sox9(+) cells were present within the surface epithelium. The expression of PCNA, a marker of cell proliferation, paralleled the distribution of Sox9 expression. Sox9(+) PDG cells proved to be Pdx1(+) /Ngn3(+/-) /Oct4A(-) . Nearly 10% of PDG cells were positive for insulin or glucagon. Intercalated ducts contained Sox9(+) /Pdx1(+) /Ngn3(+) cells, a phenotype that is presumptive of committed endocrine progenitors. Some intercalated ducts appeared in continuity with clusters of insulin-positive cells organized in small pancreatic islet-like structures. In summary, PDGs represent niches of a population of Sox9(+) cells exhibiting a pattern of phenotypic traits implicating a radial axis of maturation from the bottoms of the PDGs to the surface of pancreatic ducts. Our results complete the anatomical background that links biliary and pancreatic tracts and could have important implications for the common patho-physiology of biliary tract and pancreas.

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