Neurogranin as Cerebrospinal Fluid Biomarker for Alzheimer Disease: An Assay Comparison Study.

Background: Neurogranin in cerebrospinal fluid (CSF) correlates with cognitive decline and is a potential novel biomarker for Alzheimer disease (AD) dementia. We investigated the analytical and diagnostic performance of 3 commonly used neurogranin assays in the same cohort of patients to improve the interpretability of CSF neurogranin test results.

Methods: The neurogranin Erenna assay from Washington University, St.

Diagnostic and Prognostic Utility of the Synaptic Marker Neurogranin in Alzheimer Disease.

Importance: Synaptic loss is an early pathologic substrate of Alzheimer disease (AD). Neurogranin is a postsynaptic neuronal protein that has demonstrated utility as a cerebrospinal fluid (CSF) marker of synaptic loss in AD.

Objective: To investigate the diagnostic and prognostic utility of CSF neurogranin levels in a large, well-characterized cohort of individuals with symptomatic AD and cognitively normal controls.

Serum visinin-like protein-1 in concussed professional ice hockey players.

Primary Objective: Visinin-like protein-1 (VILIP-1) has shown potential utility as a biomarker for neuronal injury in cerebrospinal fluid. This study investigated serum VILIP-1 as a diagnostic and prognostic marker in sports-related concussion.

Methods: This multi-centre prospective cohort study involved the 12 teams of the professional ice hockey league in Sweden.

Development of an immunoassay for the kidney-specific protein myo-inositol oxygenase, a potential biomarker of acute kidney injury.

Background: Acute kidney injury (AKI) affects 45% of critically ill patients, resulting in increased morbidity and mortality. The diagnostic standard, plasma creatinine, is nonspecific and may not increase until days after injury. There is significant need for a renal-specific AKI biomarker detectable early enough that there would be a potential window for therapeutic intervention.

Xenin-25 delays gastric emptying and reduces postprandial glucose levels in humans with and without type 2 diabetes.

Xenin-25 (Xen) is a neurotensin-related peptide secreted by a subset of glucose-dependent insulinotropic polypeptide (GIP)-producing enteroendocrine cells. In animals, Xen regulates gastrointestinal function and glucose homeostasis, typically by initiating neural relays. However, little is known about Xen action in humans.

Expression of the Na+/K+-transporting ATPase gamma subunit FXYD2 in renal tumors.

Chromophobe renal cell carcinoma (RCC) is a form of renal cancer that may be confused with other eosinophilic renal tumors, including oncocytoma, type 2 papillary RCC, and clear-cell RCC with eosinophilic features. There are currently no robust markers to distinguish these neoplasms. Chromophobe RCC and renal oncocytoma are presumably derived from the distal nephron.

Xenin-25 amplifies GIP-mediated insulin secretion in humans with normal and impaired glucose tolerance but not type 2 diabetes.

Glucose-dependent insulinotropic polypeptide (GIP) potentiates glucose-stimulated insulin secretion (GSIS). This response is blunted in type 2 diabetes (T2DM). Xenin-25 is a 25-amino acid neurotensin-related peptide that amplifies GIP-mediated GSIS in hyperglycemic mice.

N-terminal pro-atrial natriuretic peptide measurement in plasma suggests covalent modification.

Background: The N-terminal fragment of cardiac-derived pro-B-type natriuretic peptide is a glycosylated polypeptide. It is unknown whether N-terminal pro-atrial natriuretic peptide (proANP) fragments are also covalently modified. We therefore evaluated the clinical performance of 2 distinctly different proANP assays on clinical outcome.

A 68 residue N-terminal fragment of pro-atrial natriuretic peptide is a monomeric intrinsically unstructured protein.

The mature pro forms of the cardiac natriuretic peptides, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), are proteolytically processed to their active hormone forms (28 and 32 residues, respectively) and N-terminal (NT)-pro fragments (68 and 76 residues, respectively). Far-ultraviolet circular dichroism (UV CD), 1D and 2D-homonuclear nuclear magnetic resonance (NMR), size exclusion-high performance liquid chromatography (SE-HPLC) and analytical ultracentrifuge sedimentation equilibrium (AUCSE) data are obtained for NT-proANP. CD data showed a large negative molar ellipticity for NT-proANP of -14,800° cm(2)/dmol at 199-200 nm.

ETRAP (efficient trapping and purification) of target protein polyclonal antibodies from GST-protein immune sera.

Recombinant GST (glutathione transferase) proteins are widely used as immunogens to generate polyclonal antibodies. Advantages of using GST proteins include: commercially available cloning vectors, vast literature for protein expression in Escherichia coli, the ease of protein purification, immunogen can be used as an ELISA standard and GST can be removed in some systems. However, there are disadvantages: GST oligomerization, inclusion body formation and target protein insolubility after GST removal.

Xenin-25 potentiates glucose-dependent insulinotropic polypeptide action via a novel cholinergic relay mechanism.

The intestinal peptides GLP-1 and GIP potentiate glucose-mediated insulin release. Agents that increase GLP-1 action are effective therapies in type 2 diabetes mellitus (T2DM). However, GIP action is blunted in T2DM, and GIP-based therapies have not been developed.

Dyskerin ablation in mouse liver inhibits rRNA processing and cell division.

Dyskerin is a component of small nucleolar ribonucleoprotein complexes and acts as a pseudouridine synthase to modify newly synthesized ribosomal, spliceosomal, and possibly other RNAs. It is encoded by the DKC1 gene, the gene mutated in X-linked dyskeratosis congenita, and is also part of the telomerase complex. The yeast ortholog, Cbf5, is an essential protein, but in mammals the effect of dyskerin ablation at the cellular level is not known.

Biomechanical stress induces novel arterial intima-enriched genes: implications for vascular adaptation to stress.

Background: The arterial vasculature is subjected to considerably greater biomechanical stress than the venous circulation. This is reflected in the difference in morphology between large arteries and veins, however little is known about the molecular differences that arise as a consequence of biomechanical stress. Previously, we identified a group of arterial intima-enriched (AIE) genes: sciellin, periplakin, SPRR3, envoplakin, galectin 7, and plakoglobin that are functionally related in that they contribute to the stress properties of stratified epithelium.

A unified sample preparation protocol for proteomic and genomic profiling of cervical swabs to identify biomarkers for cervical cancer screening.

Cervical cancer screening is ideally suited for the development of biomarkers due to the ease of tissue acquisition and the well-established histological transitions. Furthermore, cell and biologic fluid obtained from cervix samples undergo specific molecular changes that can be profiled. However, the ideal manner and techniques for preparing cervical samples remains to be determined.

Regulation of the atheroma-enriched protein, SPRR3, in vascular smooth muscle cells through cyclic strain is dependent on integrin alpha1beta1/collagen interaction.

Atherosclerotic plaques express high levels of small proline-rich repeat protein (SPRR3), a previously characterized component of the cornified cell envelope of stratified epithelia, where it is believed to play a role in cellular adaptation to biomechanical stress. We investigated the physiological signals and underlying mechanism(s) that regulate atheroma-enriched SPRR3 expression in vascular smooth muscle cells (VSMCs). We showed that SPRR3 is expressed by VSMCs in both human and mouse atheromas.

The role of human ribosomal proteins in the maturation of rRNA and ribosome production.

Production of ribosomes is a fundamental process that occurs in all dividing cells. It is a complex process consisting of the coordinated synthesis and assembly of four ribosomal RNAs (rRNA) with about 80 ribosomal proteins (r-proteins) involving more than 150 nonribosomal proteins and other factors. Diamond Blackfan anemia (DBA) is an inherited red cell aplasia caused by mutations in one of several r-proteins.

A glycosylated form of the human cardiac hormone pro B-type natriuretic peptide is an intrinsically unstructured monomeric protein.

The N-terminal fragment of pro B-type natriuretic peptide (NT-proBNP) and proBNP are used as gold standard clinical markers of myocardial dysfunction such as cardiac hypertrophy and left ventricle heart failure. The actual circulating molecular forms of these peptides have been the subject of intense investigation particularly since these analytes are measured in clinical assays. Conflicting data has been reported and no firm consensus on the exact nature of the molecular species exists.

Chemical cleavage of proteins in solution.

Described in this unit are five basic protocols that are widely used for specific and efficient chemical cleavage of proteins in solution. Cyanogen bromide (CNBr) cleaves at methionine (Met) residues; BNPS-skatole cleaves at tryptophan (Trp) residues; formic acid cleaves at aspartic acid-proline (Asp-Pro) peptide bonds; hydroxylamine cleaves at asparagine-glycine (Asn-Gly) peptide bonds, and 2-nitro-5-thiocyanobenzoic acid (NTCB) cleaves at cysteine (Cys) residues. Because the above loci are at relatively low abundance in most proteins, digestion with these agents will yield relatively long peptides.

The human cardiac hormone fragment N-terminal pro B-type natriuretic peptide is an intrinsically unstructured protein.

The cardiac hormone B-type natriuretic peptide (BNP) is synthesized as a prepro 134 residue molecule which is further proteolytically processed into a 76 residue fragment termed N-terminal proBNP (NT-proBNP) and the active portion of this hormone, a 32-residue disulfide-linked peptide (BNP-32). The active hormone regulates cardiac hemodynamic output while as yet no biological function has been attributed to NT-proBNP. Some solution properties of synthetically generated NT-proBNP in benign media are known.

Cells depleted for RPS19, a protein associated with Diamond Blackfan Anemia, show defects in 18S ribosomal RNA synthesis and small ribosomal subunit production.

The gene encoding the small subunit ribosomal protein 19 (RPS19) is mutated in about 25% of cases of the bone marrow failure syndrome Diamond Blackfan Anemia (DBA), a childhood disease characterized by failure of red cell production. In these cases DBA is inherited as an autosomal dominant trait and RPS19 haploinsufficiency is thought to cause the disease. To study the molecular pathogenesis of DBA we used siRNA to decrease the level of RPS19 in two human cell lines, HeLa cells and U-2 OS osteosarcoma cells.

Identification of novel brain biomarkers.

Background: The diagnosis of diseases leading to brain injury, such as stroke, Alzheimer disease, and Parkinson disease, can often be problematic. In this study, we pursued the discovery of biomarkers that might be specific and sensitive to brain injury.

Methods: We performed gene array analyses on a mouse model to look for biomarkers that are both preferentially and abundantly produced in the brain.

Isolation and characterization of a thermally stable recombinant anti-caffeine heavy-chain antibody fragment.

We have isolated and characterized a caffeine-specific, heavy-chain-only antibody fragment (V(HH)) from llama that is capable of being utilized to analyze caffeine in hot and cold beverages. Camelid species (llama and camel) were selected for immunization because of their potential to make heat-stable, heavy-chain-only antibodies. Llamas and camels were immunized with caffeine covalently linked to keyhole limpet hemocyanin, and recombinant antibody techniques were used to create phage displayed libraries of variable region fragments of the heavy-chain antibodies.

PEGylated murine Granulocyte-macrophage colony-stimulating factor: production, purification, and characterization.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) regulates proliferation, differentiation, and function of hematopoietic progenitor cells. Aside from expansion of hematopoietic cells, GM-CSF has shown efficacy in other diseases, including Crohn's disease. While GM-CSF being clinically used in humans, the ability to perform mechanistic studies in murine models is difficult due to the limited availability and rapid clearance of murine GM-CSF in the peripheral blood.