Jan Johansson - Academia.edu (original) (raw)

Papers by Jan Johansson

Research paper thumbnail of Choline up-regulates BDNF and down-regulates TrkB neurotrophin receptor in rat cortical cell culture

Neuroreport, Jan 17, 2009

In this study, possible involvements of choline and nicotinic acetylcholine receptors (nAChRs) in... more In this study, possible involvements of choline and nicotinic acetylcholine receptors (nAChRs) in neurotrophic-related neuronal plasticity were investigated. Primary cell cultures from rat cerebral cortex were exposed for 72 h to the alpha7 nAChR selective agonist choline and protein expression levels of the neurotrophin receptors p75, TrkA, TrkB and TrkC were examined. The results revealed a choline-induced attenuation of the TrkB expression, whereas the other neurotrophin receptors were not affected. Further analysis of choline-exposed cell cultures showed an increased protein level of the TrkB ligand brain-derived neurotrophic factor (BDNF). This increase was obtained in cell cultures where the alpha7 nAChR subunit was detected, but not in younger cell cultures where this subunit could not be detected. It is speculated that a choline-induced change of alpha7 nAChRs activity may have resulted in the observed increase of BDNF level and down-regulation of the TrkB receptor.

Research paper thumbnail of Functional Evaluation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) in human monocytes

Journal of Cystic Fibrosis, 2009

NP is a useful model to study pathogenesis of chronic airway inflammation. NP could be either of ... more NP is a useful model to study pathogenesis of chronic airway inflammation. NP could be either of unknown origin (primary NP) or secondary to congenital impairment of mucociliary clearance (CF, or primary ciliary dyskinesia, PCD). The aim of the present study was to identify the differentially expressed proteins using human nasal epithelial cell (HNEC) primary cultures from NP, and two successive proteomic approaches (2D gels following mass spectrometry (MS) and semi-quantitative iTRAQ MS/MS analysis). Using 2D approach, 51 proteins were identified (pH range 5−8) with 7 being differentially expressed between primary NP (n = 4) and secondary NP (2 CF, 2 PCD). Using iTRAQ approach, 583 proteins were identified with 21 and 25 of them being differentially expressed between primary NP (n = 1) vs control mucosa (n = 1) and primary NP (n = 1) vs CF NP (n = 1), respectively. These proteins include inflammatory (annexin 1 and 2), oxidative stress (peroxiredoxin 1, GSTp1), structural (keratin 18, alpha actinin) and chaperone proteins (several HSPs). Multiple proteins belonging to Unfolding Protein Response (UPR) proteins (e.g. PPIB, HSP70, GRP78) were also identified. Focusing on the UPR, we found that UPR decreases as function of time in HNEC cultures from p_NP (n = 3). Cytokine secretion, measured in same HNEC, decreases in parallel to UPR response (n = 3). All together, these results suggest that UPR is related to inflammatory microenvironment and play a role in pathogenesis of primary and CF NP.

Research paper thumbnail of Detection of CFTR protein in human leukocytes by flow cytometry

Cytometry Part A, 2014

Leukocytes have previously been shown to express detectable levels of the protein cystic fibrosis... more Leukocytes have previously been shown to express detectable levels of the protein cystic fibrosis transmembrane conductance regulator (CFTR). This study aims to evaluate the application of flow cytometric (FC) analysis to detect CFTR expression, and changes thereof, in these cells. Aliquots (200 lL) of peripheral whole blood from 12 healthy control volunteers (CTRLs), 12 carriers of a CFTR mutation (CFC), and 40 patients with cystic fibrosis (CF) carrying various combinations of CFTR mutations were incubated with specific fluorescent probes recognizing CFTR protein expressed on the plasma membrane of leukocytes. FC was applied to analyze CFTR expression in monocytes, lymphocytes, and polymorphonuclear (PMN) cells. CFTR protein was detected in monocytes and lymphocytes, whereas inconclusive results were obtained from the analysis of PMN cells. Mean fluorescence intensity (MFI) ratio value and %CFTRpositive cells above a selected threshold were the two parameters selected to quantify CFTR expression in cells. Lowest variability and the highest reproducibility were obtained when analyzing monocytes. ANOVA results indicated that both parameters were able to discriminate monocytes of healthy controls and CF individuals according to CFTR mutation classes with high accuracy. Significantly increased MFI ratio values were recorded in CFTR-defective cells that were also able to improve CFTR function after ex vivo treatment with PTC124 (Ataluren), an investigative drug designed to permit the ribosome to read through nonsense CFTR mutations. The method described is minimally invasive and may be used in the monitoring of responses to drugs whose efficacy can depend on increased CFTR protein expression levels.

Research paper thumbnail of Electrophysiological evaluation of cystic fibrosis conductance transmembrane regulator (CFTR) expression in human monocytes

Biochimica et biophysica acta, 2014

Cystic fibrosis is caused by mutations of CFTR gene, a protein kinase A-activated anion channel, ... more Cystic fibrosis is caused by mutations of CFTR gene, a protein kinase A-activated anion channel, and is associated to a persistent and excessive chronic lung inflammation, suggesting functional alterations of immune cells. Leukocytes express detectable levels of CFTR but the molecule has not been fully characterized in these cells. Freshly isolated monocytes from healthy individuals and CF patients were assessed by protein expression, single cell electrophysiological and membrane depolarization assays. We recorded chloride currents by patch clamp in healthy monocytes, after the administration of a CFTR stimulus. Currents were sensitive to a specific blocker of the CFTR channel, CFTRinh-172 and were absent in CF monocytes. Next, we evaluated the effects of ex vivo exposure of monocytes from cystic fibrosis patients carrying the F508del mutation to a chemical corrector, Vertex-325. We found an increase in CFTR expression by confocal microscopy and a recovery of CFTR function by both p...

Research paper thumbnail of Impaired CFTR function in mild cystic fibrosis associated with the S977F/T5TG12complex allele in trans with F508del mutation

Journal of Cystic Fibrosis, 2013

The S977F mutation (c.2930C&a... more The S977F mutation (c.2930C>T) in the CFTR gene (CFTR/ABCC7) is extremely rare. We describe the case of an adult patient carrying the complex allele S977F/T5TG12 in trans with the F508del mutation. Mild respiratory manifestations arose in adulthood associated with azoospermia, acute pancreatitis, minor hemoptysis and Cl(-) levels ranging from 40 to 42 mEq/L. Diagnosis was confirmed by repeated NPD measurements, genetic DHPLC analysis and a recently described functional assay measuring cAMP-dependent cell depolarization in peripheral blood monocytes. NPD measurements, DHPLC and monocyte functional assay (CF index=-18). Results were consistent with a CF phenotype. The combined application of DHPLC and NPD analysis in the algorithm for CF diagnosis appears useful for the management of similar cases. In addition, the novel monocyte functional assay might contribute to improve our diagnostic capability, counseling and better treatment of these challenging clinical cases.

Research paper thumbnail of Functional Evaluation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) in human monocytes

Journal of Cystic Fibrosis, 2009

NP is a useful model to study pathogenesis of chronic airway inflammation. NP could be either of ... more NP is a useful model to study pathogenesis of chronic airway inflammation. NP could be either of unknown origin (primary NP) or secondary to congenital impairment of mucociliary clearance (CF, or primary ciliary dyskinesia, PCD). The aim of the present study was to identify the differentially expressed proteins using human nasal epithelial cell (HNEC) primary cultures from NP, and two successive proteomic approaches (2D gels following mass spectrometry (MS) and semi-quantitative iTRAQ MS/MS analysis). Using 2D approach, 51 proteins were identified (pH range 5−8) with 7 being differentially expressed between primary NP (n = 4) and secondary NP (2 CF, 2 PCD). Using iTRAQ approach, 583 proteins were identified with 21 and 25 of them being differentially expressed between primary NP (n = 1) vs control mucosa (n = 1) and primary NP (n = 1) vs CF NP (n = 1), respectively. These proteins include inflammatory (annexin 1 and 2), oxidative stress (peroxiredoxin 1, GSTp1), structural (keratin 18, alpha actinin) and chaperone proteins (several HSPs). Multiple proteins belonging to Unfolding Protein Response (UPR) proteins (e.g. PPIB, HSP70, GRP78) were also identified. Focusing on the UPR, we found that UPR decreases as function of time in HNEC cultures from p_NP (n = 3). Cytokine secretion, measured in same HNEC, decreases in parallel to UPR response (n = 3). All together, these results suggest that UPR is related to inflammatory microenvironment and play a role in pathogenesis of primary and CF NP.

Research paper thumbnail of Detection of CFTR protein in human leukocytes by flow cytometry

Cytometry Part A, 2014

Leukocytes have previously been shown to express detectable levels of the protein cystic fibrosis... more Leukocytes have previously been shown to express detectable levels of the protein cystic fibrosis transmembrane conductance regulator (CFTR). This study aims to evaluate the application of flow cytometric (FC) analysis to detect CFTR expression, and changes thereof, in these cells. Aliquots (200 lL) of peripheral whole blood from 12 healthy control volunteers (CTRLs), 12 carriers of a CFTR mutation (CFC), and 40 patients with cystic fibrosis (CF) carrying various combinations of CFTR mutations were incubated with specific fluorescent probes recognizing CFTR protein expressed on the plasma membrane of leukocytes. FC was applied to analyze CFTR expression in monocytes, lymphocytes, and polymorphonuclear (PMN) cells. CFTR protein was detected in monocytes and lymphocytes, whereas inconclusive results were obtained from the analysis of PMN cells. Mean fluorescence intensity (MFI) ratio value and %CFTRpositive cells above a selected threshold were the two parameters selected to quantify CFTR expression in cells. Lowest variability and the highest reproducibility were obtained when analyzing monocytes. ANOVA results indicated that both parameters were able to discriminate monocytes of healthy controls and CF individuals according to CFTR mutation classes with high accuracy. Significantly increased MFI ratio values were recorded in CFTR-defective cells that were also able to improve CFTR function after ex vivo treatment with PTC124 (Ataluren), an investigative drug designed to permit the ribosome to read through nonsense CFTR mutations. The method described is minimally invasive and may be used in the monitoring of responses to drugs whose efficacy can depend on increased CFTR protein expression levels.

Research paper thumbnail of Challenging the diagnosis of Cystic Fibrosis in a patient carrying the 186-8T/C allelic variant in the CF Transmembrane Conductance Regulator gene

BMC Pulmonary Medicine, 2014

Background: This report describe for the first time a clinical case with a CFTR allelic variant 1... more Background: This report describe for the first time a clinical case with a CFTR allelic variant 186-8T/C (c.54-8 T/C) in intron 1 of CFTR and underline the importance of applying a combination of genetic and functional tests to establish or exclude a diagnosis of Cystic Fibrosis. In this case the diagnostic algorithm proposed for CF has been successfully applied at our Center and previous CF diagnosis assigned in a different Center was not confirmed.

Research paper thumbnail of Defective CFTR Expression and Function Are Detectable in Blood Monocytes: Development of a New Blood Test for Cystic Fibrosis

PLoS ONE, 2011

Background: Evaluation of cystic fibrosis transmembrane conductance regulator (CFTR) functional a... more Background: Evaluation of cystic fibrosis transmembrane conductance regulator (CFTR) functional activity to assess new therapies and define diagnosis of cystic fibrosis (CF) is cumbersome. It is known that leukocytes express detectable levels of CFTR but the molecule has not been characterized in these cells. In this study we aim at setting up and validating a blood test to evaluate CFTR expression and function in leukocytes.

Research paper thumbnail of Choline up-regulates BDNF and down-regulates TrkB neurotrophin receptor in rat cortical cell culture

Neuroreport, Jan 17, 2009

In this study, possible involvements of choline and nicotinic acetylcholine receptors (nAChRs) in... more In this study, possible involvements of choline and nicotinic acetylcholine receptors (nAChRs) in neurotrophic-related neuronal plasticity were investigated. Primary cell cultures from rat cerebral cortex were exposed for 72 h to the alpha7 nAChR selective agonist choline and protein expression levels of the neurotrophin receptors p75, TrkA, TrkB and TrkC were examined. The results revealed a choline-induced attenuation of the TrkB expression, whereas the other neurotrophin receptors were not affected. Further analysis of choline-exposed cell cultures showed an increased protein level of the TrkB ligand brain-derived neurotrophic factor (BDNF). This increase was obtained in cell cultures where the alpha7 nAChR subunit was detected, but not in younger cell cultures where this subunit could not be detected. It is speculated that a choline-induced change of alpha7 nAChRs activity may have resulted in the observed increase of BDNF level and down-regulation of the TrkB receptor.

Research paper thumbnail of Functional Evaluation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) in human monocytes

Journal of Cystic Fibrosis, 2009

NP is a useful model to study pathogenesis of chronic airway inflammation. NP could be either of ... more NP is a useful model to study pathogenesis of chronic airway inflammation. NP could be either of unknown origin (primary NP) or secondary to congenital impairment of mucociliary clearance (CF, or primary ciliary dyskinesia, PCD). The aim of the present study was to identify the differentially expressed proteins using human nasal epithelial cell (HNEC) primary cultures from NP, and two successive proteomic approaches (2D gels following mass spectrometry (MS) and semi-quantitative iTRAQ MS/MS analysis). Using 2D approach, 51 proteins were identified (pH range 5−8) with 7 being differentially expressed between primary NP (n = 4) and secondary NP (2 CF, 2 PCD). Using iTRAQ approach, 583 proteins were identified with 21 and 25 of them being differentially expressed between primary NP (n = 1) vs control mucosa (n = 1) and primary NP (n = 1) vs CF NP (n = 1), respectively. These proteins include inflammatory (annexin 1 and 2), oxidative stress (peroxiredoxin 1, GSTp1), structural (keratin 18, alpha actinin) and chaperone proteins (several HSPs). Multiple proteins belonging to Unfolding Protein Response (UPR) proteins (e.g. PPIB, HSP70, GRP78) were also identified. Focusing on the UPR, we found that UPR decreases as function of time in HNEC cultures from p_NP (n = 3). Cytokine secretion, measured in same HNEC, decreases in parallel to UPR response (n = 3). All together, these results suggest that UPR is related to inflammatory microenvironment and play a role in pathogenesis of primary and CF NP.

Research paper thumbnail of Detection of CFTR protein in human leukocytes by flow cytometry

Cytometry Part A, 2014

Leukocytes have previously been shown to express detectable levels of the protein cystic fibrosis... more Leukocytes have previously been shown to express detectable levels of the protein cystic fibrosis transmembrane conductance regulator (CFTR). This study aims to evaluate the application of flow cytometric (FC) analysis to detect CFTR expression, and changes thereof, in these cells. Aliquots (200 lL) of peripheral whole blood from 12 healthy control volunteers (CTRLs), 12 carriers of a CFTR mutation (CFC), and 40 patients with cystic fibrosis (CF) carrying various combinations of CFTR mutations were incubated with specific fluorescent probes recognizing CFTR protein expressed on the plasma membrane of leukocytes. FC was applied to analyze CFTR expression in monocytes, lymphocytes, and polymorphonuclear (PMN) cells. CFTR protein was detected in monocytes and lymphocytes, whereas inconclusive results were obtained from the analysis of PMN cells. Mean fluorescence intensity (MFI) ratio value and %CFTRpositive cells above a selected threshold were the two parameters selected to quantify CFTR expression in cells. Lowest variability and the highest reproducibility were obtained when analyzing monocytes. ANOVA results indicated that both parameters were able to discriminate monocytes of healthy controls and CF individuals according to CFTR mutation classes with high accuracy. Significantly increased MFI ratio values were recorded in CFTR-defective cells that were also able to improve CFTR function after ex vivo treatment with PTC124 (Ataluren), an investigative drug designed to permit the ribosome to read through nonsense CFTR mutations. The method described is minimally invasive and may be used in the monitoring of responses to drugs whose efficacy can depend on increased CFTR protein expression levels.

Research paper thumbnail of Electrophysiological evaluation of cystic fibrosis conductance transmembrane regulator (CFTR) expression in human monocytes

Biochimica et biophysica acta, 2014

Cystic fibrosis is caused by mutations of CFTR gene, a protein kinase A-activated anion channel, ... more Cystic fibrosis is caused by mutations of CFTR gene, a protein kinase A-activated anion channel, and is associated to a persistent and excessive chronic lung inflammation, suggesting functional alterations of immune cells. Leukocytes express detectable levels of CFTR but the molecule has not been fully characterized in these cells. Freshly isolated monocytes from healthy individuals and CF patients were assessed by protein expression, single cell electrophysiological and membrane depolarization assays. We recorded chloride currents by patch clamp in healthy monocytes, after the administration of a CFTR stimulus. Currents were sensitive to a specific blocker of the CFTR channel, CFTRinh-172 and were absent in CF monocytes. Next, we evaluated the effects of ex vivo exposure of monocytes from cystic fibrosis patients carrying the F508del mutation to a chemical corrector, Vertex-325. We found an increase in CFTR expression by confocal microscopy and a recovery of CFTR function by both p...

Research paper thumbnail of Impaired CFTR function in mild cystic fibrosis associated with the S977F/T5TG12complex allele in trans with F508del mutation

Journal of Cystic Fibrosis, 2013

The S977F mutation (c.2930C&a... more The S977F mutation (c.2930C>T) in the CFTR gene (CFTR/ABCC7) is extremely rare. We describe the case of an adult patient carrying the complex allele S977F/T5TG12 in trans with the F508del mutation. Mild respiratory manifestations arose in adulthood associated with azoospermia, acute pancreatitis, minor hemoptysis and Cl(-) levels ranging from 40 to 42 mEq/L. Diagnosis was confirmed by repeated NPD measurements, genetic DHPLC analysis and a recently described functional assay measuring cAMP-dependent cell depolarization in peripheral blood monocytes. NPD measurements, DHPLC and monocyte functional assay (CF index=-18). Results were consistent with a CF phenotype. The combined application of DHPLC and NPD analysis in the algorithm for CF diagnosis appears useful for the management of similar cases. In addition, the novel monocyte functional assay might contribute to improve our diagnostic capability, counseling and better treatment of these challenging clinical cases.

Research paper thumbnail of Functional Evaluation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) in human monocytes

Journal of Cystic Fibrosis, 2009

NP is a useful model to study pathogenesis of chronic airway inflammation. NP could be either of ... more NP is a useful model to study pathogenesis of chronic airway inflammation. NP could be either of unknown origin (primary NP) or secondary to congenital impairment of mucociliary clearance (CF, or primary ciliary dyskinesia, PCD). The aim of the present study was to identify the differentially expressed proteins using human nasal epithelial cell (HNEC) primary cultures from NP, and two successive proteomic approaches (2D gels following mass spectrometry (MS) and semi-quantitative iTRAQ MS/MS analysis). Using 2D approach, 51 proteins were identified (pH range 5−8) with 7 being differentially expressed between primary NP (n = 4) and secondary NP (2 CF, 2 PCD). Using iTRAQ approach, 583 proteins were identified with 21 and 25 of them being differentially expressed between primary NP (n = 1) vs control mucosa (n = 1) and primary NP (n = 1) vs CF NP (n = 1), respectively. These proteins include inflammatory (annexin 1 and 2), oxidative stress (peroxiredoxin 1, GSTp1), structural (keratin 18, alpha actinin) and chaperone proteins (several HSPs). Multiple proteins belonging to Unfolding Protein Response (UPR) proteins (e.g. PPIB, HSP70, GRP78) were also identified. Focusing on the UPR, we found that UPR decreases as function of time in HNEC cultures from p_NP (n = 3). Cytokine secretion, measured in same HNEC, decreases in parallel to UPR response (n = 3). All together, these results suggest that UPR is related to inflammatory microenvironment and play a role in pathogenesis of primary and CF NP.

Research paper thumbnail of Detection of CFTR protein in human leukocytes by flow cytometry

Cytometry Part A, 2014

Leukocytes have previously been shown to express detectable levels of the protein cystic fibrosis... more Leukocytes have previously been shown to express detectable levels of the protein cystic fibrosis transmembrane conductance regulator (CFTR). This study aims to evaluate the application of flow cytometric (FC) analysis to detect CFTR expression, and changes thereof, in these cells. Aliquots (200 lL) of peripheral whole blood from 12 healthy control volunteers (CTRLs), 12 carriers of a CFTR mutation (CFC), and 40 patients with cystic fibrosis (CF) carrying various combinations of CFTR mutations were incubated with specific fluorescent probes recognizing CFTR protein expressed on the plasma membrane of leukocytes. FC was applied to analyze CFTR expression in monocytes, lymphocytes, and polymorphonuclear (PMN) cells. CFTR protein was detected in monocytes and lymphocytes, whereas inconclusive results were obtained from the analysis of PMN cells. Mean fluorescence intensity (MFI) ratio value and %CFTRpositive cells above a selected threshold were the two parameters selected to quantify CFTR expression in cells. Lowest variability and the highest reproducibility were obtained when analyzing monocytes. ANOVA results indicated that both parameters were able to discriminate monocytes of healthy controls and CF individuals according to CFTR mutation classes with high accuracy. Significantly increased MFI ratio values were recorded in CFTR-defective cells that were also able to improve CFTR function after ex vivo treatment with PTC124 (Ataluren), an investigative drug designed to permit the ribosome to read through nonsense CFTR mutations. The method described is minimally invasive and may be used in the monitoring of responses to drugs whose efficacy can depend on increased CFTR protein expression levels.

Research paper thumbnail of Challenging the diagnosis of Cystic Fibrosis in a patient carrying the 186-8T/C allelic variant in the CF Transmembrane Conductance Regulator gene

BMC Pulmonary Medicine, 2014

Background: This report describe for the first time a clinical case with a CFTR allelic variant 1... more Background: This report describe for the first time a clinical case with a CFTR allelic variant 186-8T/C (c.54-8 T/C) in intron 1 of CFTR and underline the importance of applying a combination of genetic and functional tests to establish or exclude a diagnosis of Cystic Fibrosis. In this case the diagnostic algorithm proposed for CF has been successfully applied at our Center and previous CF diagnosis assigned in a different Center was not confirmed.

Research paper thumbnail of Defective CFTR Expression and Function Are Detectable in Blood Monocytes: Development of a New Blood Test for Cystic Fibrosis

PLoS ONE, 2011

Background: Evaluation of cystic fibrosis transmembrane conductance regulator (CFTR) functional a... more Background: Evaluation of cystic fibrosis transmembrane conductance regulator (CFTR) functional activity to assess new therapies and define diagnosis of cystic fibrosis (CF) is cumbersome. It is known that leukocytes express detectable levels of CFTR but the molecule has not been characterized in these cells. In this study we aim at setting up and validating a blood test to evaluate CFTR expression and function in leukocytes.