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Ulf-håkan Stenman - One of the best experts on this subject based on the ideXlab platform.
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MAPK inhibitors induce serine peptidase inhibitor Kazal type 1 (SPINK1) secretion in BRAF V600E-mutant colorectal adenocarcinoma.
Molecular oncology, 2017Co-Authors: Kati Räsänen, Hannu Koistinen, Ulf-håkan Stenman, Caj Haglund, Kien Xuan Dang, Harri Mustonen, Susanna Lintula, Jakob StenmanAbstract:The mitogen-activated protein kinase (MAPK) pathway plays a central role in colorectal cancers (CRC). In particular, BRAF V600E-mutant tumors, which represent around 10% of CRCs, are refractory to current therapies. Overexpression and secretion of serine peptidase inhibitor Kazal type 1 (SPINK1) are observed in around 50% of CRCs, and its serum level can be used as a biomarker for poor prognosis. Utilizing a recently developed extendable blocking probe assay, we analyzed the BRAF mutation status in a CRC patient cohort (N = 571) using tissue-derived RNA as the starting material. From the same RNA samples, we measured the relative SPINK1 expression levels using a quantitative real-time PCR method. Expression of mutant BRAF V600E correlated with poor prognosis, as did low expression of SPINK1 mRNA. Further, BRAF V600E correlated negatively with SPINK1 levels. In order to investigate the effect of MAPK pathway-targeted therapies on SPINK1 secretion, we conducted in vitro studies using both wild-type and V600E CRC cell lines. BRAF inhibitor vemurafenib, and subsequent MAPK pathway inhibitors trametinib and SCH772984, significantly increased SPINK1 secretion in V600E CRC cell lines Colo205 and HT-29 with a concomitant decrease in Trypsin-1 and -2 secretion. Notably, no SPINK1 increase or Trypsin-1 decrease was observed in BRAF wild-type CRC cell line Caco-2 in response to MAPK pathway inhibitors. In further mechanistic studies, we observed that only trametinib was able to diminish completely both MEK and ERK phosphorylation in the V600E CRC cells. Furthermore, the key regulator of integrated stress response, activating transcription factor 4 (ATF-4), was downregulated both at mRNA and at protein level in response to trametinib treatment. In conclusion, these data suggest that sustained inhibition of not only MAPK pathway activation, but also ATF-4 and Trypsin, might be beneficial in the therapy of BRAF V600E-mutant CRC and that SPINK1 levels may serve as an indicator of therapy response.
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Abstract A42: Interleukin-6 induces secretion of SPINK1 and Trypsin in colorectal cancer
Cell-Cell Signaling in the Tumor Microenvironment, 2016Co-Authors: Kati Räsänen, Outi Itkonen, Ulf-håkan Stenman, Elina Lehtinen, Kristiina Nokelainen, Laura Hautala, Hannu KoistinenAbstract:Inflammation is known to promote colorectal cancer (CRC) tumorigenesis, but the underlying molecular mechanisms are still being uncovered. Proinflammatory cytokine interleukin-6 (IL-6) is primarily secreted by the cells of the tumor microenvironment (TME), mainly by inflammatory cells but also by activated fibroblasts. IL-6 stimulates growth and survival signaling in CRC. Inflammatory signals regulate also the production and activity of proteases and their inhibitors. Serine protease inhibitor Kazal type1 (SPINK1, aka tumor-associated Trypsin inhibitor, TATI) is expressed in several tissues and over-expression of SPINK1 predicts an unfavorable outcome in many cancers, including colon cancer. The SPINK1 gene contains an IL-6 responsive element and in addition to being a protease inhibitor, it also acts as an acute phase reactant and a growth factor. Expression of serine proteases Trypsin-1 and -2, the main targets of SPINK1, also correlate with malignancy and metastatic potential. The aim of this study was to assess the paracrine signaling between fibroblast-derived IL-6 and cancer cell-derived SPINK1, Trypsin-1 and -2. The following expression analyses were used: quantitative PCR, immunohistochemistry, immunofluorometric assays and Western blotting. The results show that CRC cells Colo205 and HT-29 express SPINK1 and secrete it into the culture medium. IL-6 dose-dependently increased the mRNA expression and protein levels of SPINK1. Conditioned media from fibroblasts had the same effect, and conversely CRC media increased IL-6 secretion in the fibroblasts, indicating paracrine signaling between these cell types. In CRC tissues cancer cells were positive for SPINK1, while IL-6 was found in fibroblasts surrounding the cancer cells, confirming the in vitro results. In Colo205 cells the baseline levels of Trypsin-1 and -2 and the respective genes PRSS1 and PRSS2 were much higher compared to HT-29 cells. In Colo205 cells IL-6 led to concomitant increase in the secretion of Trypsin-1 and -2, whereas in HT-29 cells these remained constantly low. Mechanistically, addition of IL-6 led to activation of the canonical Stat3 pathway, as indicated by Stat3 phosphorylation. Stat3 inhibitor reduced both SPINK1 and Trypsin-1 and -2 levels, demonstrating that Stat3 is the transcription factor driving their expression. Taken together, our results show a connection between TME-derived inflammatory response and increased SPINK1 and Trypsin-1 and -2 levels. Elevated serum SPINK1 has been shown to be an independent prognostic factor in colon cancer. As SPINK1 acts as a growth factor in some cancers, it has also been suggested as a therapeutic target. Therefore assessment of SPINK1 expression and function has several potentially important clinical applications in colorectal and other cancers. The study also strengthens the role of tumor microenvironment in tumor progression and emphasizes the importance of studying tumor-stroma crosstalk of proteolytic processing. Citation Format: Kati Rasanen, Elina Lehtinen, Kristiina Nokelainen, Laura Hautala, Outi Itkonen, Ulf-Hakan Stenman, Hannu Koistinen. Interleukin-6 induces secretion of SPINK1 and Trypsin in colorectal cancer. [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr A42.
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Interleukin-6 increases expression of serine protease inhibitor Kazal type 1 through STAT3 in colorectal adenocarcinoma.
Molecular carcinogenesis, 2015Co-Authors: Kati Räsänen, Outi Itkonen, Ulf-håkan Stenman, Elina Lehtinen, Kristiina Nokelainen, Laura Hautala, Teijo Kuopio, Hannu KoistinenAbstract:Inflammation promotes colorectal cancer (CRC) tumorigenesis, but the underlying molecular mechanisms are still being uncovered. Proinflammatory cytokine interleukin-6 (IL-6) stimulates survival signaling in CRC; inflammatory signals also regulate production and activity of proteases and their inhibitors. Over-expression of serine protease inhibitor Kazal type 1 (SPINK1) predicts an unfavorable outcome in colon cancer. The SPINK1 gene contains an IL-6 responsive element, suggesting it could act as an acute phase reactant. We assessed the connection between IL-6 and SPINK1, and the function and mechanism of this signaling. Our results show that Colo205 and HT-29 cells express and secrete SPINK1, and both fibroblast-derived and recombinant IL-6 further increased the SPINK1 levels. Concurrently CRC cells augmented the IL-6 production in fibroblasts. In CRC tissues cancer cells were positive for SPINK1, whereas IL-6 was found in stromal cells. In Colo205 cells IL-6 also stimulated the secretion of Trypsin-1 and -2, the key targets of SPINK1 protease inhibition, whereas in HT-29 cells Trypsin-1 and -2 levels remained constantly low. Functionally, both IL-6 and SPINK1 increased the motility of the CRC cells. Mechanistically, IL-6 activated the canonical STAT3 pathway and inhibition of STAT3 phosphorylation decreased the levels of SPINK1, Trypsin-1 and -2. Taken together, our results indicate a novel link between inflammatory signals originating from the tumor microenvironment and increased SPINK1 levels. This finding has potential therapeutic implications for targeted therapy, as it confirms that SPINK1 acts as an acute phase reactant and that it participates in the paracrine crosstalk with the tumor microenvironment of colon cancer. © 2015 Wiley Periodicals, Inc.
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Collagen degradation by tumor-associated Trypsins.
Archives of Biochemistry and Biophysics, 2013Co-Authors: Lynn S. Mirigian, Hannu Koistinen, Outi Itkonen, Elena Makareeva, Timo Sorsa, Ulf-håkan Stenman, Tuula Salo, Sergey LeikinAbstract:In normal soft tissues, collagen is degraded primarily by collagenases from the matrix metalloproteinase family. Yet, collagenase-like activity of tumor-associated isoforms of other enzymes might be involved in cancer invasion as well. In the present study, we systematically examined collagen degradation by non-sulfated isoforms of Trypsins, which were proposed to possess such an activity. We found that non-sulfated Trypsin-1, -2, and -3 were able to cleave non-helical and unfolded regions of collagen chains but not the intact triple helix, similar to sulfated Trypsins produced by the pancreas. Trypsin-2 sulfation did not affect the cleavage rate either. An apparent triple helix cleavage by tumor-associated Trypsin-2 reported earlier likely occurred after triple helix unfolding during sample denaturation for gel electrophoresis. Nevertheless, tumor-associated Trypsins might be important for releasing collagen from fibers through telopeptide cleavage as well as for degrading unfolded collagen chains, e.g. after initial cleavage and destabilization of triple helices by collagenases.
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Urinary matrix metalloproteinase -8, -9, -14 and their regulators (TRY-1, TRY-2, TATI) in patients with diabetic nephropathy
Annals of medicine, 2008Co-Authors: Anneli Lauhio, Timo Sorsa, Ulf-håkan Stenman, Ravi Srinivas, Mathias Stenman, Taina Tervahartiala, Carola Grönhagen-riska, Eero HonkanenAbstract:Matrix metalloproteinase-9 (MMP-9) has been shown to be involved in the development of diabetic nephropathy (DNP). We studied the levels, molecular forms, and degree of activation of urinary MMP-8, -9, -14, Trypsin-1 and -2, as well as tumor-associated Trypsin inhibitor (TATI) of DNP patients and healthy controls.Urinary samples were analyzed for MMPs by Western blotting and gelatin zymography and for Trypsin-1, -2, and TATI by time-resolved immunofluorometric assays.Total MMP-8 immunoreactivity, the proportion of active MMP-9, and gelatinolytic activity in urine were significantly higher in DNP patients than in controls. In urine of DNP patients the proportion of active polymorphonuclear neutrophil (PMN)-type (but not fibroblast-type) MMP-8 was increased. MMP-8 and MMP-9 were found to form high molecular weight complexes in DNP urine. Total immunoreactivity of soluble urinary MMP-14 and the levels of Trypsin (TRY)-1 and TRY-2, but not of TATI, were also significantly increased in DNP. Zymography, Western...
Hannu Koistinen - One of the best experts on this subject based on the ideXlab platform.
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MAPK inhibitors induce serine peptidase inhibitor Kazal type 1 (SPINK1) secretion in BRAF V600E-mutant colorectal adenocarcinoma.
Molecular oncology, 2017Co-Authors: Kati Räsänen, Hannu Koistinen, Ulf-håkan Stenman, Caj Haglund, Kien Xuan Dang, Harri Mustonen, Susanna Lintula, Jakob StenmanAbstract:The mitogen-activated protein kinase (MAPK) pathway plays a central role in colorectal cancers (CRC). In particular, BRAF V600E-mutant tumors, which represent around 10% of CRCs, are refractory to current therapies. Overexpression and secretion of serine peptidase inhibitor Kazal type 1 (SPINK1) are observed in around 50% of CRCs, and its serum level can be used as a biomarker for poor prognosis. Utilizing a recently developed extendable blocking probe assay, we analyzed the BRAF mutation status in a CRC patient cohort (N = 571) using tissue-derived RNA as the starting material. From the same RNA samples, we measured the relative SPINK1 expression levels using a quantitative real-time PCR method. Expression of mutant BRAF V600E correlated with poor prognosis, as did low expression of SPINK1 mRNA. Further, BRAF V600E correlated negatively with SPINK1 levels. In order to investigate the effect of MAPK pathway-targeted therapies on SPINK1 secretion, we conducted in vitro studies using both wild-type and V600E CRC cell lines. BRAF inhibitor vemurafenib, and subsequent MAPK pathway inhibitors trametinib and SCH772984, significantly increased SPINK1 secretion in V600E CRC cell lines Colo205 and HT-29 with a concomitant decrease in Trypsin-1 and -2 secretion. Notably, no SPINK1 increase or Trypsin-1 decrease was observed in BRAF wild-type CRC cell line Caco-2 in response to MAPK pathway inhibitors. In further mechanistic studies, we observed that only trametinib was able to diminish completely both MEK and ERK phosphorylation in the V600E CRC cells. Furthermore, the key regulator of integrated stress response, activating transcription factor 4 (ATF-4), was downregulated both at mRNA and at protein level in response to trametinib treatment. In conclusion, these data suggest that sustained inhibition of not only MAPK pathway activation, but also ATF-4 and Trypsin, might be beneficial in the therapy of BRAF V600E-mutant CRC and that SPINK1 levels may serve as an indicator of therapy response.
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Abstract A42: Interleukin-6 induces secretion of SPINK1 and Trypsin in colorectal cancer
Cell-Cell Signaling in the Tumor Microenvironment, 2016Co-Authors: Kati Räsänen, Outi Itkonen, Ulf-håkan Stenman, Elina Lehtinen, Kristiina Nokelainen, Laura Hautala, Hannu KoistinenAbstract:Inflammation is known to promote colorectal cancer (CRC) tumorigenesis, but the underlying molecular mechanisms are still being uncovered. Proinflammatory cytokine interleukin-6 (IL-6) is primarily secreted by the cells of the tumor microenvironment (TME), mainly by inflammatory cells but also by activated fibroblasts. IL-6 stimulates growth and survival signaling in CRC. Inflammatory signals regulate also the production and activity of proteases and their inhibitors. Serine protease inhibitor Kazal type1 (SPINK1, aka tumor-associated Trypsin inhibitor, TATI) is expressed in several tissues and over-expression of SPINK1 predicts an unfavorable outcome in many cancers, including colon cancer. The SPINK1 gene contains an IL-6 responsive element and in addition to being a protease inhibitor, it also acts as an acute phase reactant and a growth factor. Expression of serine proteases Trypsin-1 and -2, the main targets of SPINK1, also correlate with malignancy and metastatic potential. The aim of this study was to assess the paracrine signaling between fibroblast-derived IL-6 and cancer cell-derived SPINK1, Trypsin-1 and -2. The following expression analyses were used: quantitative PCR, immunohistochemistry, immunofluorometric assays and Western blotting. The results show that CRC cells Colo205 and HT-29 express SPINK1 and secrete it into the culture medium. IL-6 dose-dependently increased the mRNA expression and protein levels of SPINK1. Conditioned media from fibroblasts had the same effect, and conversely CRC media increased IL-6 secretion in the fibroblasts, indicating paracrine signaling between these cell types. In CRC tissues cancer cells were positive for SPINK1, while IL-6 was found in fibroblasts surrounding the cancer cells, confirming the in vitro results. In Colo205 cells the baseline levels of Trypsin-1 and -2 and the respective genes PRSS1 and PRSS2 were much higher compared to HT-29 cells. In Colo205 cells IL-6 led to concomitant increase in the secretion of Trypsin-1 and -2, whereas in HT-29 cells these remained constantly low. Mechanistically, addition of IL-6 led to activation of the canonical Stat3 pathway, as indicated by Stat3 phosphorylation. Stat3 inhibitor reduced both SPINK1 and Trypsin-1 and -2 levels, demonstrating that Stat3 is the transcription factor driving their expression. Taken together, our results show a connection between TME-derived inflammatory response and increased SPINK1 and Trypsin-1 and -2 levels. Elevated serum SPINK1 has been shown to be an independent prognostic factor in colon cancer. As SPINK1 acts as a growth factor in some cancers, it has also been suggested as a therapeutic target. Therefore assessment of SPINK1 expression and function has several potentially important clinical applications in colorectal and other cancers. The study also strengthens the role of tumor microenvironment in tumor progression and emphasizes the importance of studying tumor-stroma crosstalk of proteolytic processing. Citation Format: Kati Rasanen, Elina Lehtinen, Kristiina Nokelainen, Laura Hautala, Outi Itkonen, Ulf-Hakan Stenman, Hannu Koistinen. Interleukin-6 induces secretion of SPINK1 and Trypsin in colorectal cancer. [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr A42.
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Interleukin-6 increases expression of serine protease inhibitor Kazal type 1 through STAT3 in colorectal adenocarcinoma.
Molecular carcinogenesis, 2015Co-Authors: Kati Räsänen, Outi Itkonen, Ulf-håkan Stenman, Elina Lehtinen, Kristiina Nokelainen, Laura Hautala, Teijo Kuopio, Hannu KoistinenAbstract:Inflammation promotes colorectal cancer (CRC) tumorigenesis, but the underlying molecular mechanisms are still being uncovered. Proinflammatory cytokine interleukin-6 (IL-6) stimulates survival signaling in CRC; inflammatory signals also regulate production and activity of proteases and their inhibitors. Over-expression of serine protease inhibitor Kazal type 1 (SPINK1) predicts an unfavorable outcome in colon cancer. The SPINK1 gene contains an IL-6 responsive element, suggesting it could act as an acute phase reactant. We assessed the connection between IL-6 and SPINK1, and the function and mechanism of this signaling. Our results show that Colo205 and HT-29 cells express and secrete SPINK1, and both fibroblast-derived and recombinant IL-6 further increased the SPINK1 levels. Concurrently CRC cells augmented the IL-6 production in fibroblasts. In CRC tissues cancer cells were positive for SPINK1, whereas IL-6 was found in stromal cells. In Colo205 cells IL-6 also stimulated the secretion of Trypsin-1 and -2, the key targets of SPINK1 protease inhibition, whereas in HT-29 cells Trypsin-1 and -2 levels remained constantly low. Functionally, both IL-6 and SPINK1 increased the motility of the CRC cells. Mechanistically, IL-6 activated the canonical STAT3 pathway and inhibition of STAT3 phosphorylation decreased the levels of SPINK1, Trypsin-1 and -2. Taken together, our results indicate a novel link between inflammatory signals originating from the tumor microenvironment and increased SPINK1 levels. This finding has potential therapeutic implications for targeted therapy, as it confirms that SPINK1 acts as an acute phase reactant and that it participates in the paracrine crosstalk with the tumor microenvironment of colon cancer. © 2015 Wiley Periodicals, Inc.
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Collagen degradation by tumor-associated Trypsins.
Archives of Biochemistry and Biophysics, 2013Co-Authors: Lynn S. Mirigian, Hannu Koistinen, Outi Itkonen, Elena Makareeva, Timo Sorsa, Ulf-håkan Stenman, Tuula Salo, Sergey LeikinAbstract:In normal soft tissues, collagen is degraded primarily by collagenases from the matrix metalloproteinase family. Yet, collagenase-like activity of tumor-associated isoforms of other enzymes might be involved in cancer invasion as well. In the present study, we systematically examined collagen degradation by non-sulfated isoforms of Trypsins, which were proposed to possess such an activity. We found that non-sulfated Trypsin-1, -2, and -3 were able to cleave non-helical and unfolded regions of collagen chains but not the intact triple helix, similar to sulfated Trypsins produced by the pancreas. Trypsin-2 sulfation did not affect the cleavage rate either. An apparent triple helix cleavage by tumor-associated Trypsin-2 reported earlier likely occurred after triple helix unfolding during sample denaturation for gel electrophoresis. Nevertheless, tumor-associated Trypsins might be important for releasing collagen from fibers through telopeptide cleavage as well as for degrading unfolded collagen chains, e.g. after initial cleavage and destabilization of triple helices by collagenases.
Esko Kemppainen - One of the best experts on this subject based on the ideXlab platform.
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Sequential changes in pancreatic markers in acute pancreatitis.
Scandinavian journal of gastroenterology, 2003Co-Authors: Marko Lempinen, Pauli Puolakkainen, U H Stenman, A. Hietaranta, Reijo Haapiainen, Esko KemppainenAbstract:Background: Trypsinogen activation within acinar cells plays a crucial role in the pathogenesis of acute pancreatitis (AP). Our aim was to characterize temporal changes of Trypsinogen-1, Trypsinogen-2, complexes of Trypsin-1- ! 1 -antiTrypsin (T1-AAT) and Trypsin-2- ! 1 -antiTrypsin (T2-AAT), Trypsinogen activation peptide (TAP) and pancreatic secretory Trypsin inhibitor (PSTI) in patients with AP. Methods: The study comprised 64 consecutive patients with AP (19 with severe disease) and 32 controls. The concentrations of Trypsinogen-1 and -2, PSTI, T1-AAT and T2-AAT were determined by time-resolved immunofluorometric assays (IFMA), and TAP was measured using a competitive enzyme immunoassay from serum and urine. Results: The concentrations of Trypsinogen-1 and -2 in serum reflected similar patterns, but excretion of Trypsinogen-1 into urine was markedly lower than that of Trypsinogen-2, the concentration of which correlated strongly with disease severity. The concentrations of T1-AAT were no higher ...
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The Ratio of Trypsin-2-α1-AntiTrypsin to Trypsinogen-1 Discriminates Biliary and Alcohol-induced Acute Pancreatitis
Clinical chemistry, 2001Co-Authors: Jan M. Andersén, J. Hedström, Esko Kemppainen, Patrik Finne, Pauli Puolakkainen, Ulf-håkan StenmanAbstract:Background: Rapid determination of the etiology of acute pancreatitis (AP) enables institution of appropriate treatment. We evaluated the ability of Trypsinogen-1, Trypsinogen-2, Trypsin-1-α1-antiTrypsin (AAT), and Trypsin-2-AAT in serum to identify the etiology of AP. Methods: The study consisted of 67 consecutive patients with AP admitted to Helsinki University Central Hospital. Forty-two had alcohol-induced AP, 16 had biliary AP, and 9 had unexplained etiology. Serum samples were drawn within 12 h after admission. Trypsinogen-1, Trypsinogen-2, Trypsin-1-AAT, and Trypsin-2-AAT were determined by time-resolved immunofluorometric assays. Logistic regression was used to estimate the ability of the serum analytes to discriminate between alcohol-induced and biliary AP. The validity of the tests was evaluated by ROC curve analysis. Results: Patients with alcohol-induced AP had higher median values of Trypsin-1-AAT ( P = 0.065), Trypsinogen-2 ( P = 0.034), and Trypsin-2-AAT ( P
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Time-resolved Immunofluorometric Assay of Trypsin-1 Complexed with α1-AntiTrypsin in Serum: Increased Immunoreactivity in Patients with Biliary Tract Cancer
Clinical chemistry, 1999Co-Authors: J. Hedström, Caj Haglund, Esko Kemppainen, Maarit Leinimaa, Jari Leinonen, Ulf-håkan StenmanAbstract:Background: Increased serum concentrations of Trypsin immunoreactivity occur in patients with biliary tract cancer. To characterize this Trypsin, we developed a sensitive time-resolved immunofluorometric assay for Trypsin-1 complexed with α1-antiTrypsin (AAT) and studied the concentrations of this complex in sera from healthy individuals (n = 130) and patients with benign biliary disease (n = 32), biliary tract cancer (n = 17), pancreatic cancer (n = 27), and hepatocellular cancer (n = 12). Methods: We used a Trypsin-1-specific monoclonal antibody on the solid phase and a europium-labeled polyclonal antibody to AAT as tracer. The detection limit was 0.42 μg/L. The validity of the Trypsin-1-AAT test for detection of biliary tract cancer was compared with Trypsin-2-AAT and CA19-9. Results: Increased concentrations of Trypsin-1-AAT (>33 μg/L) were found in 76% of patients with biliary tract cancer, and the concentrations were significantly higher than in those with benign biliary disease ( P
J. Hedström - One of the best experts on this subject based on the ideXlab platform.
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The Ratio of Trypsin-2-α1-AntiTrypsin to Trypsinogen-1 Discriminates Biliary and Alcohol-induced Acute Pancreatitis
Clinical chemistry, 2001Co-Authors: Jan M. Andersén, J. Hedström, Esko Kemppainen, Patrik Finne, Pauli Puolakkainen, Ulf-håkan StenmanAbstract:Background: Rapid determination of the etiology of acute pancreatitis (AP) enables institution of appropriate treatment. We evaluated the ability of Trypsinogen-1, Trypsinogen-2, Trypsin-1-α1-antiTrypsin (AAT), and Trypsin-2-AAT in serum to identify the etiology of AP. Methods: The study consisted of 67 consecutive patients with AP admitted to Helsinki University Central Hospital. Forty-two had alcohol-induced AP, 16 had biliary AP, and 9 had unexplained etiology. Serum samples were drawn within 12 h after admission. Trypsinogen-1, Trypsinogen-2, Trypsin-1-AAT, and Trypsin-2-AAT were determined by time-resolved immunofluorometric assays. Logistic regression was used to estimate the ability of the serum analytes to discriminate between alcohol-induced and biliary AP. The validity of the tests was evaluated by ROC curve analysis. Results: Patients with alcohol-induced AP had higher median values of Trypsin-1-AAT ( P = 0.065), Trypsinogen-2 ( P = 0.034), and Trypsin-2-AAT ( P
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Time-resolved Immunofluorometric Assay of Trypsin-1 Complexed with α1-AntiTrypsin in Serum: Increased Immunoreactivity in Patients with Biliary Tract Cancer
Clinical chemistry, 1999Co-Authors: J. Hedström, Caj Haglund, Esko Kemppainen, Maarit Leinimaa, Jari Leinonen, Ulf-håkan StenmanAbstract:Background: Increased serum concentrations of Trypsin immunoreactivity occur in patients with biliary tract cancer. To characterize this Trypsin, we developed a sensitive time-resolved immunofluorometric assay for Trypsin-1 complexed with α1-antiTrypsin (AAT) and studied the concentrations of this complex in sera from healthy individuals (n = 130) and patients with benign biliary disease (n = 32), biliary tract cancer (n = 17), pancreatic cancer (n = 27), and hepatocellular cancer (n = 12). Methods: We used a Trypsin-1-specific monoclonal antibody on the solid phase and a europium-labeled polyclonal antibody to AAT as tracer. The detection limit was 0.42 μg/L. The validity of the Trypsin-1-AAT test for detection of biliary tract cancer was compared with Trypsin-2-AAT and CA19-9. Results: Increased concentrations of Trypsin-1-AAT (>33 μg/L) were found in 76% of patients with biliary tract cancer, and the concentrations were significantly higher than in those with benign biliary disease ( P
Juan-carlos Fontecilla-camps - One of the best experts on this subject based on the ideXlab platform.
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Crystal structure of human Trypsin 1: unexpected phosphorylation of Tyr151.
Journal of molecular biology, 1996Co-Authors: Christine Gaboriaud, Laurence Serre, Odette Guy-crotte, Eric Forest, Juan-carlos Fontecilla-campsAbstract:Abstract The X-ray structure of human Trypsin 1 has been determined in the presence of diisopropyl-phosphofluoridate by the molecular replacement method and refined at a resolution of 2.2 A to anR-factor of 18%. Crystals belong to the space groupP4, with two independent molecules in the asymmetric unit packing as crystallographic tetramers. This study was performed in order to seek possible structural peculiarities of human Trypsin 1, suggested by some striking differences in its biochemical behavior as compared to other Trypsins of mammalian species. Its fold is, in fact, very similar to those of the bovine, rat and porcine Trypsins, with root-mean-square differences in the 0.4 to 0.6 A range for all 223 Cαpositions. The most unexpected feature of the human Trypsin 1 structure is in the phosphorylated state of tyrosine residue 151 in the present X-ray study. This feature was confirmed by mass spectrometry on the same inhibited sample and also on the native enzyme. This phosphorylation strengthens the outstanding clustering of highly negative or highly positive electrostatic surface potentials. The peculiar inhibitory behaviour of pancreatic secretory Trypsin inhibitors of the Kazal type on this enzyme is discussed as a possible consequence of these properties. A charged surface loop has also been interpreted as an epitope site recognised by a monoclonal antibody specific to human Trypsin 1.
