Advancements in transcriptomic technologies enable global comparative studies allowing identifica... more Advancements in transcriptomic technologies enable global comparative studies allowing identification of genes with tissue-enriched expression. In this study we profile 280 brain-enriched proteins in cerebrospinal fluid (CSF) from patients with Alzheimer's disease (AD), Parkinson's disease (PD) and dementia with Lewy bodies (DLB). In total, 441 human samples of ventricular CSF collected post mortem and lumbar CSF collected ante mortem were analyzed using 376 antibodies in a suspension bead array setup, utilizing a direct labelling approach. Among several proteins displaying differentiated profiles between sample groups, we focus here on two synaptic proteins, neuromodulin (GAP43) and neurogranin (NRGN). They were 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 were also observed for patients for whom the AD diagnosi...
Analysis of the pattern of proteins or messengerRNAs (mRNAs) in histological tissue sections is a... more Analysis of the pattern of proteins or messengerRNAs (mRNAs) in histological tissue sections is a cornerstone in biomedical research and diagnostics. This typically involves the visualization of a few proteins or expressed genes at a time. We have devised a strategy, which we call "spatial transcriptomics," that allows visualization and quantitative analysis of the transcriptome with spatial resolution in individual tissue sections. By positioning histological sections on arrayed reverse transcription primers with unique positional barcodes, we demonstrate high-quality RNA-sequencing data with maintained two-dimensional positional information from the mouse brain and human breast cancer. Spatial transcriptomics provides quantitative gene expression data and visualization of the distribution of mRNAs within tissue sections and enables novel types of bioinformatics analyses, valuable in research and diagnostics.
Proceedings of the National Academy of Sciences of the United States of America, Jan 16, 2007
Autoimmune polyendocrine syndrome type 1 (APS1) is a rare autosomal recessive disorder caused by ... more Autoimmune polyendocrine syndrome type 1 (APS1) is a rare autosomal recessive disorder caused by mutations in the autoimmune regulator (AIRE) gene. High titer autoantibodies (Aabs) toward intracellular enzymes are a hallmark for APS1 and serve as diagnostic markers and predictors for disease manifestations. In this study, we aimed to identify pituitary autoantigens in patients with APS1. A pituitary cDNA expression library was screened with APS1 sera and a tudor domain containing protein 6 (TDRD6) cDNA clone was isolated. Positive immunoreactivity against in vitro translated TDRD6 fragments was shown in 42/86 (49%) APS1 patients but not in patients with other autoimmune diseases or in healthy controls. By using immunohistochemistry, sera from 3/6 APS1 patients with growth hormone (GH) deficiency showed immunostaining of a small number of guinea pig anterior pituitary cells, and 40-50% of these cells were GH-positive. No such immunostaining was seen with sera from healthy controls. The APS1 Aab-positive, GH-negative cells may represent a novel subpopulation of anterior pituitary cells. In addition, 4/6 patient sera showed staining of a fiber-plexus in the pituitary intermediate lobe recognizing enzymes of monoamine and GABA synthesis. Thus, we have identified TDRD6 as a major autoantigen in APS1 patients and shown that several sera from GH-deficient patients stain specific cell populations and nerves in the pituitary gland.
The spatiotemporal expression of cannabinoid receptors and endocannabinoid-metabolising enzymes d... more The spatiotemporal expression of cannabinoid receptors and endocannabinoid-metabolising enzymes during brain development guides major developmental processes including neurogenesis, cell differentiation, cell migration, neuronal specification and synaptogenesis.Endocannabinoids (eCBs) play an important role in fine-tuning neurotransmission and have recently been shown to play an important role in brain development. The spatiotemporal expression of cannabinoid receptors and endocannabinoid-metabolising enzymes during development guides major developmental processes including neurogenesis, cell differentiation, cell migration, neuronal specification and synaptogenesis. Furthermore, pharmacological experiments and transgenic animal models have shown the impact of disrupted eCB signalling on normal brain development and revealed the danger of both cannabis abuse and exposure to cannabinoid drugs during embryonic development, childhood and adolescence. In this review, we focus on the dynamic expression of eCB components and the physiological role eCBs play during brain development.
Drug metabolism and disposition: the biological fate of chemicals, Jan 10, 2015
Since its development, tariquidar (XR9576, TQR) has been widely regarded as one of the more poten... more Since its development, tariquidar (XR9576, TQR) has been widely regarded as one of the more potent inhibitors of P-glycoprotein (P-gp), an efflux transporter of the ATP-binding cassette (ABC) transporter family. A third-generation inhibitor, TQR exhibits high affinity for P-gp, although it is also a substrate of another ABC transporter, breast cancer resistance protein (BCRP). Recently, several studies have questioned the mechanism by which TQR interfaces with P-gp, suggesting that TQR is a substrate for P-gp instead of a noncompetitive inhibitor. We investigated TQR and its interaction with human and mouse P-gp to determine if TQR is a substrate of P-gp in vitro. To address these questions, we used multiple in vitro transporter assays including cytotoxicity, flow cytometry, accumulation, ATPase, and transwell assays. A newly generated BCRP cell line was used as a positive control that demonstrates TQR-mediated transport. Based on our results, we conclude that TQR is a potent inhibi...
The Journal of pharmacology and experimental therapeutics, Jan 6, 2015
Imaging ATP-binding cassette (ABC) transporter activity in vivo with positron emission tomography... more Imaging ATP-binding cassette (ABC) transporter activity in vivo with positron emission tomography requires both a substrate and a transporter inhibitor. However, for ABCG2, there is no inhibitor proven to be specific to that transporter alone at the blood-brain barrier. Ko143, a non-toxic analog of fungal toxin fumitremorgin C, is a potent inhibitor of ABCG2, although its specificity in mouse and human systems is unclear. This study examined the selectivity of Ko143 using human embryonic kidney cell lines transfected with ABCG2, ABCB1, or ABCC1 in several in vitro assays. The stability of Ko143 in rat plasma was measured using high performance liquid chromatography. Our results show that in addition to being a potent inhibitor of ABCG2, at higher concentrations (≥ 1 μM) Ko143 also has an effect on the transport activity of both ABCB1 and ABCC1. Furthermore, Ko143 was found to be unstable in rat plasma. These findings indicate that Ko143 lacks specificity for ABCG2 and should be take...
The mammalian brain is a complex organ composed of many specialized cells, harboring sets of both... more 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-enr...
Adequate estimation of neuroinflammatory processes following ischemic stroke is essential for bet... more Adequate estimation of neuroinflammatory processes following ischemic stroke is essential for better understanding of disease mechanisms, and for the development of treatment strategies. With the TSPO (18 kDa translocator protein) positron emission tomography (PET) radioligand [(11)C]PBR28, we monitored longitudinally the inflammatory response post-transient cerebral ischemia in rats, using a recently developed rat stroke model that produces isolated focal cortical infarcts with clinical relevance in size and pathophysiology. Six Sprague-Dawley rats were subjected to 90 min transient endovascular occlusion of the M2 segment of the middle cerebral artery (M2CAO). Animals were imaged with a nanoScan(®) PET/MRI system at 1, 4, 7 and 14 days after M2CAO with a bolus injection of [(11)C]PBR28. In the infarct region, we found a significantly increased uptake of [(11)C]PBR28 on day 4, 7 and 14 compared to day 1 as well as compared to the contralateral cortex. No significant increase was de...
Advancements in transcriptomic technologies enable global comparative studies allowing identifica... more Advancements in transcriptomic technologies enable global comparative studies allowing identification of genes with tissue-enriched expression. In this study we profile 280 brain-enriched proteins in cerebrospinal fluid (CSF) from patients with Alzheimer's disease (AD), Parkinson's disease (PD) and dementia with Lewy bodies (DLB). In total, 441 human samples of ventricular CSF collected post mortem and lumbar CSF collected ante mortem were analyzed using 376 antibodies in a suspension bead array setup, utilizing a direct labelling approach. Among several proteins displaying differentiated profiles between sample groups, we focus here on two synaptic proteins, neuromodulin (GAP43) and neurogranin (NRGN). They were 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 were also observed for patients for whom the AD diagnosi...
Analysis of the pattern of proteins or messengerRNAs (mRNAs) in histological tissue sections is a... more Analysis of the pattern of proteins or messengerRNAs (mRNAs) in histological tissue sections is a cornerstone in biomedical research and diagnostics. This typically involves the visualization of a few proteins or expressed genes at a time. We have devised a strategy, which we call "spatial transcriptomics," that allows visualization and quantitative analysis of the transcriptome with spatial resolution in individual tissue sections. By positioning histological sections on arrayed reverse transcription primers with unique positional barcodes, we demonstrate high-quality RNA-sequencing data with maintained two-dimensional positional information from the mouse brain and human breast cancer. Spatial transcriptomics provides quantitative gene expression data and visualization of the distribution of mRNAs within tissue sections and enables novel types of bioinformatics analyses, valuable in research and diagnostics.
Proceedings of the National Academy of Sciences of the United States of America, Jan 16, 2007
Autoimmune polyendocrine syndrome type 1 (APS1) is a rare autosomal recessive disorder caused by ... more Autoimmune polyendocrine syndrome type 1 (APS1) is a rare autosomal recessive disorder caused by mutations in the autoimmune regulator (AIRE) gene. High titer autoantibodies (Aabs) toward intracellular enzymes are a hallmark for APS1 and serve as diagnostic markers and predictors for disease manifestations. In this study, we aimed to identify pituitary autoantigens in patients with APS1. A pituitary cDNA expression library was screened with APS1 sera and a tudor domain containing protein 6 (TDRD6) cDNA clone was isolated. Positive immunoreactivity against in vitro translated TDRD6 fragments was shown in 42/86 (49%) APS1 patients but not in patients with other autoimmune diseases or in healthy controls. By using immunohistochemistry, sera from 3/6 APS1 patients with growth hormone (GH) deficiency showed immunostaining of a small number of guinea pig anterior pituitary cells, and 40-50% of these cells were GH-positive. No such immunostaining was seen with sera from healthy controls. The APS1 Aab-positive, GH-negative cells may represent a novel subpopulation of anterior pituitary cells. In addition, 4/6 patient sera showed staining of a fiber-plexus in the pituitary intermediate lobe recognizing enzymes of monoamine and GABA synthesis. Thus, we have identified TDRD6 as a major autoantigen in APS1 patients and shown that several sera from GH-deficient patients stain specific cell populations and nerves in the pituitary gland.
The spatiotemporal expression of cannabinoid receptors and endocannabinoid-metabolising enzymes d... more The spatiotemporal expression of cannabinoid receptors and endocannabinoid-metabolising enzymes during brain development guides major developmental processes including neurogenesis, cell differentiation, cell migration, neuronal specification and synaptogenesis.Endocannabinoids (eCBs) play an important role in fine-tuning neurotransmission and have recently been shown to play an important role in brain development. The spatiotemporal expression of cannabinoid receptors and endocannabinoid-metabolising enzymes during development guides major developmental processes including neurogenesis, cell differentiation, cell migration, neuronal specification and synaptogenesis. Furthermore, pharmacological experiments and transgenic animal models have shown the impact of disrupted eCB signalling on normal brain development and revealed the danger of both cannabis abuse and exposure to cannabinoid drugs during embryonic development, childhood and adolescence. In this review, we focus on the dynamic expression of eCB components and the physiological role eCBs play during brain development.
Drug metabolism and disposition: the biological fate of chemicals, Jan 10, 2015
Since its development, tariquidar (XR9576, TQR) has been widely regarded as one of the more poten... more Since its development, tariquidar (XR9576, TQR) has been widely regarded as one of the more potent inhibitors of P-glycoprotein (P-gp), an efflux transporter of the ATP-binding cassette (ABC) transporter family. A third-generation inhibitor, TQR exhibits high affinity for P-gp, although it is also a substrate of another ABC transporter, breast cancer resistance protein (BCRP). Recently, several studies have questioned the mechanism by which TQR interfaces with P-gp, suggesting that TQR is a substrate for P-gp instead of a noncompetitive inhibitor. We investigated TQR and its interaction with human and mouse P-gp to determine if TQR is a substrate of P-gp in vitro. To address these questions, we used multiple in vitro transporter assays including cytotoxicity, flow cytometry, accumulation, ATPase, and transwell assays. A newly generated BCRP cell line was used as a positive control that demonstrates TQR-mediated transport. Based on our results, we conclude that TQR is a potent inhibi...
The Journal of pharmacology and experimental therapeutics, Jan 6, 2015
Imaging ATP-binding cassette (ABC) transporter activity in vivo with positron emission tomography... more Imaging ATP-binding cassette (ABC) transporter activity in vivo with positron emission tomography requires both a substrate and a transporter inhibitor. However, for ABCG2, there is no inhibitor proven to be specific to that transporter alone at the blood-brain barrier. Ko143, a non-toxic analog of fungal toxin fumitremorgin C, is a potent inhibitor of ABCG2, although its specificity in mouse and human systems is unclear. This study examined the selectivity of Ko143 using human embryonic kidney cell lines transfected with ABCG2, ABCB1, or ABCC1 in several in vitro assays. The stability of Ko143 in rat plasma was measured using high performance liquid chromatography. Our results show that in addition to being a potent inhibitor of ABCG2, at higher concentrations (≥ 1 μM) Ko143 also has an effect on the transport activity of both ABCB1 and ABCC1. Furthermore, Ko143 was found to be unstable in rat plasma. These findings indicate that Ko143 lacks specificity for ABCG2 and should be take...
The mammalian brain is a complex organ composed of many specialized cells, harboring sets of both... more 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-enr...
Adequate estimation of neuroinflammatory processes following ischemic stroke is essential for bet... more Adequate estimation of neuroinflammatory processes following ischemic stroke is essential for better understanding of disease mechanisms, and for the development of treatment strategies. With the TSPO (18 kDa translocator protein) positron emission tomography (PET) radioligand [(11)C]PBR28, we monitored longitudinally the inflammatory response post-transient cerebral ischemia in rats, using a recently developed rat stroke model that produces isolated focal cortical infarcts with clinical relevance in size and pathophysiology. Six Sprague-Dawley rats were subjected to 90 min transient endovascular occlusion of the M2 segment of the middle cerebral artery (M2CAO). Animals were imaged with a nanoScan(®) PET/MRI system at 1, 4, 7 and 14 days after M2CAO with a bolus injection of [(11)C]PBR28. In the infarct region, we found a significantly increased uptake of [(11)C]PBR28 on day 4, 7 and 14 compared to day 1 as well as compared to the contralateral cortex. No significant increase was de...
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Papers by Jan Mulder