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The creatine-phosphocreatine cycle serves as a crucial temporary energy buffering system in the brain, regulated by brain creatine kinase (CKB), in maintaining Adenosine triphosphate (ATP) levels. Alzheimer's disease (AD) has been linked to increased CKB oxidation and loss of its regulatory function, although specific pathological processes and affected cell types remain unclear. In our study, cerebral cortex samples from individuals with AD, dementia with Lewy bodies (DLB), and age-matched controls were analyzed using antibody-based methods to quantify CKB levels and assess alterations associated with disease processes. Two independently validated antibodies exclusively labeled astrocytes in the human cerebral cortex. Combining immunofluorescence (IF) and mass spectrometry (MS), we explored CKB availability in AD and DLB cases. IF and Western blot analysis demonstrated a loss of CKB immunoreactivity correlated with increased plaque load, severity of tau pathology, and Lewy body pathology. However, transcriptomics data and targeted MS demonstrated unaltered total CKB levels, suggesting posttranslational modifications (PTMs) affecting antibody binding. This aligns with altered efficiency at proteolytic cleavage sites indicated in the targeted MS experiment. These findings highlight that the proper function of astrocytes, understudied in the brain compared with neurons, is highly affected by PTMs. Reduction in ATP levels within astrocytes can disrupt ATP-dependent processes, such as the glutamate-glutamine cycle. As CKB and the creatine-phosphocreatine cycle are important in securing constant ATP availability, PTMs in CKB, and astrocyte dysfunction may disturb homeostasis, driving excitotoxicity in the AD brain. CKB and its activity could be promising biomarkers for monitoring early-stage energy deficits in AD. Brain creatine kinase (CKB) mainly expressed in strocytes in the human brain. A loss of CKB immunoreactivity correlated with increased plaque load. CKB antibod, Funding Agencies|Vetenskapsrdet

Human prefrontal cortex (hPFC) is a complex brain region involved in cognitive and emotional processes and several psychiatric disorders. Here, we present an overview of the distribution of the peptidergic systems in 17 subregions of hPFC and three reference cortices obtained by microdissection and based on RNA sequencing and RNA-scope methods integrated with published single-cell transcriptomics data. We detected expression of 60 neuropeptides and 60 neuropeptide receptors in at least one of the hPFC subregions. The results reveal that the peptidergic landscape in PFC consists of closely located and functionally different subregions with unique peptide/transmitter- related profiles. Neuropeptide-rich PFC subregions were identified, encompassing regions from anterior cingulate cortex/orbitofrontal gyrus. Furthermore, marked differences in gene expression exist between different PFC regions (>5-fold; cocaine and amphetamine-regulated transcript peptide) as well as between PFC regions and reference regions, for example, for somatostatin and several receptors. We suggest that the present approach allows definition of, still hypothetical, microcircuits exemplified by glutamatergic neurons expressing a peptide cotransmitter either as an agonist (hypocretin/orexin) or antagonist (galanin). Specific neuropeptide receptors have been identified as possible targets for neuronal afferents and, interestingly, peripheral blood-borne peptide hormones (leptin, adiponectin, gastric inhibitory peptide, glucagon-like peptides, and peptide YY). Together with other recent publications, our results support the view that neuropeptide systems may play an important role in hPFC and underpin the concept that neuropeptide signaling helps stabilize circuit connectivity and fine-tune/modulate PFC functions executed during health and disease., QC 20221223

Human prefrontal cortex (hPFC) is a complex brain region involved in cognitive and emotional processes and several psychiatric disorders. Here, we present an overview of the distribution of the peptidergic systems in 17 subregions of hPFC and three reference cortices obtained by microdissection and based on RNA sequencing and RNAscope methods integrated with published single-cell transcriptomics data. We detected expression of 60 neuropeptides and 60 neuropeptide receptors in at least one of the hPFC subregions. The results reveal that the peptidergic landscape in PFC consists of closely located and functionally different subregions with unique peptide/transmitter–related profiles. Neuropeptide-rich PFC subregions were identified, encompassing regions from anterior cingulate cortex/orbitofrontal gyrus. Furthermore, marked differences in gene expression exist between different PFC regions (>5-fold; cocaine and amphetamine–regulated transcript peptide) as well as between PFC regions and reference regions, for example, for somatostatin and several receptors. We suggest that the present approach allows definition of, still hypothetical, microcircuits exemplified by glutamatergic neurons expressing a peptide cotransmitter either as an agonist (hypocretin/orexin) or antagonist (galanin). Specific neuropeptide receptors have been identified as possible targets for neuronal afferents and, interestingly, peripheral blood-borne peptide hormones (leptin, adiponectin, gastric inhibitory peptide, glucagon-like peptides, and peptide YY). Together with other recent publications, our results support the view that neuropeptide systems may play an important role in hPFC and underpin the concept that neuropeptide signaling helps stabilize circuit connectivity and fine-tune/modulate PFC functions executed during health and disease.

Simple Summary: Pituitary neuroendocrine tumours originate from the endocrine cells of the anterior pituitary gland and may develop from any of the cell lineages responsible for producing the different pituitary hormones. The details related to tumour differentiation and hormone production in these tumours are not fully understood. The aim of our study was to investigate an uncharacterised pituitary enriched protein, transforming growth factor beta-receptor 3 like (TGFBR3L). The TGFBR3L protein is highly expressed in the pituitary compared to other organs. We found the protein to be gonadotroph-specific, i.e., detected in the cells that express follicle-stimulating and luteinizing hormones (FSH/LH). The gonadotroph-specific nature of TGFBR3L, a correlation to both FSH and LH as well as an inverse correlation to membranous E-cadherin and oestrogen receptor beta suggests a role in gonadotroph cell development and function and, possibly, tumour progression. Here, we report the investigation of transforming growth factor beta-receptor 3 like (TGFBR3L), an uncharacterised pituitary specific membrane protein, in non-neoplastic anterior pituitary gland and pituitary neuroendocrine tumours. A polyclonal antibody produced within the Human Protein Atlas project (HPA074356) was used for TGFBR3L staining and combined with SF1 and FSH for a 3-plex fluorescent protocol, providing more details about the cell lineage specificity of TGFBR3L expression. A cohort of 230 pituitary neuroendocrine tumours were analysed. In a subgroup of previously characterised gonadotroph tumours, correlation with expression of FSH/LH, E-cadherin, oestrogen (ER) and somatostatin receptors (SSTR) was explored. TGFBR3L showed membranous immunolabeling and was found to be gonadotroph cell lineage-specific, verified by co-expression with SF1 and FSH/LH staining in both tumour and non-neoplastic anterior pituitary tissues. TGFBR3L immunoreactivity was observed in gonadotroph tumours only and demonstrated int, QC 20210202

Here, we report the investigation of transforming growth factor beta-receptor 3 like (TGFBR3L), an uncharacterised pituitary specific membrane protein, in non-neoplastic anterior pituitary gland and pituitary neuroendocrine tumours. A polyclonal antibody produced within the Human Protein Atlas project (HPA074356) was used for TGFBR3L staining and combined with SF1 and FSH for a 3-plex fluorescent protocol, providing more details about the cell lineage specificity of TGFBR3L expression. A cohort of 230 pituitary neuroendocrine tumours were analysed. In a subgroup of previously characterised gonadotroph tumours, correlation with expression of FSH/LH, E-cadherin, oestrogen (ER) and somatostatin receptors (SSTR) was explored. TGFBR3L showed membranous immunolabeling and was found to be gonadotroph cell lineage-specific, verified by co-expression with SF1 and FSH/LH staining in both tumour and non-neoplastic anterior pituitary tissues. TGFBR3L immunoreactivity was observed in gonadotroph tumours only and demonstrated intra-tumour heterogeneity with a perivascular location. TGFBR3L immunostaining correlated positively to both FSH (R = 0.290) and LH (R = 0.390) immunostaining, and SSTR3 (R = 0.315). TGFBR3L correlated inversely to membranous E-cadherin staining (R = -0.351) and oestrogen receptor β mRNA (R = -0.274). In conclusion, TGFBR3L is a novel pituitary gland specific protein, located in the membrane of gonadotroph cells in non-neoplastic anterior pituitary gland and in a subset of gonadotroph pituitary tumours.

The brain, with its diverse physiology and intricate cellular organization, is the most complex organ of the mammalian body. To expand our basic understanding of the neurobiology of the brain and its diseases, we performed a comprehensive molecular dissection of 10 major brain regions and multiple subregions using a variety of transcriptomics methods and antibody-based mapping. This analysis was carried out in the human, pig, and mouse brain to allow the identification of regional expression profiles, as well as to study similarities and differences in expression levels between the three species. The resulting data have been made available in an open-access Brain Atlas resource, part of the Human Protein Atlas, to allow exploration and comparison of the expression of individual protein-coding genes in various parts of the mammalian brain.

Background: A better definition of biomarkers and biological processes related to local recurrence and disease progression is highly warranted for ductal breast carcinoma in situ (DCIS). Stromal-epithelial interactions are likely of major importance for the biological, clinical, and pathological distinctions between high- and low-risk DCIS cases. Methods: Stromal platelet derived growth factor receptor (PDGFR) was immunohistochemically assessed in two DCIS patient cohorts (n = 458 and n = 80). Cox proportional hazards models were used to calculate the hazard ratios of recurrence. The molecular mechanisms regulating stromal PDGFR expression were investigated in experimental in vitro co-culture systems of DCIS cells and fibroblasts and analyzed using immunoblot and quantitative real-time PCR. Knock-out of JAG1 in DCIS cells and NOTCH2 in fibroblasts was obtained through CRISPR/Cas9. Experimental data were validated by mammary fat pad injection of DCIS and DCIS-JAG1 knock-out cells (10 mice per group). All statistical tests were two-sided. Results: PDGFR alpha((low))/PDGFR beta((high)) fibroblasts were associated with increased risk for recurrence in DCIS (univariate hazard ratio = 1.59, 95% confidence interval [CI] = 1.02 to 2.46; P = .04 Wald test; multivariable hazard ratio = 1.78, 95% CI = 1.07 to 2.97; P = .03). Tissue culture and mouse model studies indicated that this fibroblast phenotype is induced by DCIS cells in a cell contact-dependent manner. Epithelial Jagged1 and fibroblast Notch2 were identified through loss-of-function studies as key juxtacrine signaling components driving the formation of the poor prognosis-associated fibroblast phenotype. Conclusions: A PDGFR alpha((low))/PDGFR beta((high)) fibroblast subset was identified as a marker for high-risk DCIS. The Jagged-1/Notch2/PDGFR stroma-epithelial pathway was described as a novel signaling mechanism regulating this poor prognosis-associated fibroblast subset. In general terms, the study highlights ep

Introduction: Due to the rise in the number of patients with dementia the imperative for finding new diagnostic and treatment options becomes ever more pressing. While significant progress has been made in PET imaging of A beta aggregates both in vitro and in vivo, options for imaging tau protein aggregates selectively are still limited. Based on the work previously published by researchers from the Tohoku University, Japan, that resulted in the development of [F-18]THK-5351, we have undertaken an effort to develop a carbon-11 version of the identical structure - [C-11]THK-5351. In parallel, THK-5351 was also labeled with tritium ([H-3]THK-5351) for use in in vitro autoradiography (ARG). Methods: The carbon -11 labeling was performed starting with di-protected enantiomeric pure precursor-tertbutyl 5-(6-( (2S)-3-fluoro-2-(tetrahydro-2H-Pyran-2-yloxy)proPoxy)quinolin-2-yl)pyridin-2-yl carbamate, which was reacted with [C-11]MeI, using DMF as the solvent and NaH as base, followed by deprotection with trifluoroacetic acid/water mixture, resulting in enantiomerically pure carbon-11 radioligand,[C-11]THK-5351 (S)-1-fluoro-3-(2-(6-([C-11]methylamino)pyridin-3-yl)quinolin-6-yloxy)propan-2-ol. Tritium labeling and purification of [H-3]THK-5351 were undertaken using similar approach, resulting in [H-3]THK-5351 with RCP >99.8% and specific radioactivity of 13 GB q/mu mol. Results: [C-11]THK-5351 was produced in good yield (1900 +/- 355 MBq), specific radioactivity (SRA) (361 +/- 119 GBq/mu rnol at EOS + 20 min) and radiochemical purity (RCP) (>99.8%), with enantiomeric purity of 98.7%. [H-3]TM-5351 was evaluated for ARG of tau binding in post-mortem human brain tissue using cortical sections from one AD patient and one control subject. [H-3]THK-5351 binding density was higher in the AD patient compared to the control subject, the binding was displaced by unlabeled THK-5351 confirming specific [H-3] THK-5351 binding. (C) 2016 Elsevier Inc. All rights reserved.

Purpose: This study is part of a larger effort aiming to expand the knowledge of brain-enriched proteins in human cerebrospinal fluid (CSF) and to provide novel insight into the relation between such proteins and different neurodegenerative diseases. Experimental design: Here 280 brain-enriched proteins in CSF from patients with Alzheimer's disease (AD), Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are profiled. In total, 441 human samples of ventricular CSF collected post mortem and lumbar CSF collected ante mortem are analyzed using 376 antibodies in a suspension bead array setup, utilizing a direct labelling approach. Results: Among several proteins displaying differentiated profiles between sample groups, we focus here on two synaptic proteins, neuromodulin (GAP43) and neurogranin (NRGN). They are both found at elevated levels in CSF from AD patients in two independent cohorts, providing disease-associated profiles in addition to verifying and strengthening previously observed patterns. Increased levels are also observed for patients for whom the AD diagnosis was not established at the time of sampling. Conclusions and clinical relevance: These findings indicate that analyzing the brain-enriched proteins in CSF is of particular interest to increase the understanding of the CSF proteome and its relation to neurodegenerative disorders. In addition, this study lends support to the notion that measurements of these synaptic proteins could potentially be of great relevance in future diagnostic tests for AD.

The mammalian brain is a complex organ composed of many specialized cells, harboring sets of both common, widely distributed, as well as specialized and discretely localized proteins. Here we focus on the human brain, utilizing transcriptomics and public available Human Protein Atlas (HPA) data to analyze brain-enriched (frontal cortex) polyadenylated messenger RNA and long non-coding RNA and generate a genome-wide draft of global and cellular expression patterns of the brain. Based on transcriptomics analysis of altogether 27 tissues, we have estimated that approximately 3% (n=571) of all protein coding genes and 13% (n=87) of the long non-coding genes expressed in the human brain are enriched, having at least five times higher expression levels in brain as compared to any of the other analyzed peripheral tissues. Based on gene ontology analysis and detailed annotation using antibody-based tissue micro array analysis of the corresponding proteins, we found the majority of brain-enriched protein coding genes to be expressed in astrocytes, oligodendrocytes or in neurons with molecular properties linked to synaptic transmission and brain development. Detailed analysis of the transcripts and the genetic landscape of brainenriched coding and non-coding genes revealed brain-enriched splice variants. Several clusters of neighboring brain-enriched genes were also identified, suggesting regulation of gene expression on the chromatin level. This multi-angle approach uncovered the brainenriched transcriptome and linked genes to cell types and functions, providing novel insights into the molecular foundation of this highly specialized organ., QC 20150713