THE S100 PROTEINS AS MEDIATORS OF INFLAMMATION ARE INCREASED IN MYELOPROLIFERATIVE NEOPLASM AND DOWNREGULATED BY JAK2 INHIBITION
(Abstract release date: 05/19/16)
EHA Library. Mitrovic O. 06/09/16; 134912; PB2012
Dr. Olivera Mitrovic
Contributions
Contributions
Abstract
Abstract: PB2012
Type: Publication Only
Background
Chronic inflammation is associated with sustained myeloproliferation, while S100 proteins have been shown to regulate cell proliferation, differentiation and inflammation in pathological conditions. S100A8, A9 and A12 produced by cells of myeloid origin were mediators of inflammation, while S100A4 regulated cell proliferation in neoplasia. S100A4 and A12 are produced via activation of the JAK-STAT pathway, constitutively activated in myeloproliferative neoplasm (MPN).
Aims
This study analyzed the inflammation linked S100 proteins in MPNs: polycythemia vera (PV, n=16), essential thrombocythemia (ET, n=16), primary myelofibrosis (PMF, n=20), according to JAK2V617F and CALR mutation status, and healthy controls (n=8), and their regulation in JAK2V617F mutated human erythroleukemia cell line (HEL) during MPN simulated therapy.
Methods
S100A4, A9 and A12 mRNA levels in granulocytes and HEL cells are measured by real time PCR, while protein expression of S100A4, A8, A9 and A12 are examined in granulocytes and plasma of MPN using immunoblotting and immunoassay, respectively. In addition, S100A9 is determined by immunohistochemistry in bone marrow. Also, levels of S100A4, A9 and A12 are measured by real time PCR and immunoblotting in HEL cells incubated 48 hours with hydroxyurea and specific JAK2 inhibitor 1,2,3,4,5,6-hexabromocyclohexane (HBC). Mutations of JAK2V617F and CALR exon 9 are analyzed by DNA sequencing and allelic PCR.
Results
S100A12 mRNA level is significantly increased in JAK2V617F heterozygous ET patients, but downregulated in JAK2V617F heterozygous PV patients and PMF without mutation (p<0.05). However at protein level, S100A12 is significantly increased only in granulocytes of ET patients without mutation (p<0.05) and plasma of PV and PMF without JAK2V617F mutation (70 ng/ml, p<0.01). Besides non significant modification at mRNA levels, S100A4 and S100A9 protein expression demonstrated a common significant increase in granulocytes (p<0.01), followed by increased quantity of S100A9-positive cells in bone marrow and S100A8/A9 proteins in plasma of MPN patients (51 and 28 ng/ml, p<0.01). Presence of CALR mutation augmented S100A8/A9 levels in plasma and granulocytes of ET and PMF patients. After 48 hour, hydroxyurea and specific JAK2 inhibitor HBC significantly decreased S100A4 mRNA levels in HEL cells (p<0.01). Further on, protein expression of S100A4 and S100A9 are also significantly downregulated by hydroxyurea and JAK2 inhibitor HBC in HEL cells (p<0.05).
Conclusion
S100A4, A9 and A12 mRNA and protein levels are not generally influenced by JAK2V617F mutant allele burden. S100A4 and anti-inflammatory S100A8/A9 protein levels demonstrated stable elevation in contrast to sporadic pro-inflammatory S100A12 appearance in MPNs. Moreover, JAK2 inhibition reduced S100A4 and S100A9 levels in HEL cells. S100A8/A9 could serve as a clinical biomarker and therapeutic target in MPNs, with existing S100A8/A9 blockers permitted for clinical testing.
Session topic: E-poster
Keyword(s): Inflammation, Janus Kinase inhibitor, Myeloproliferative disorder
Type: Publication Only
Background
Chronic inflammation is associated with sustained myeloproliferation, while S100 proteins have been shown to regulate cell proliferation, differentiation and inflammation in pathological conditions. S100A8, A9 and A12 produced by cells of myeloid origin were mediators of inflammation, while S100A4 regulated cell proliferation in neoplasia. S100A4 and A12 are produced via activation of the JAK-STAT pathway, constitutively activated in myeloproliferative neoplasm (MPN).
Aims
This study analyzed the inflammation linked S100 proteins in MPNs: polycythemia vera (PV, n=16), essential thrombocythemia (ET, n=16), primary myelofibrosis (PMF, n=20), according to JAK2V617F and CALR mutation status, and healthy controls (n=8), and their regulation in JAK2V617F mutated human erythroleukemia cell line (HEL) during MPN simulated therapy.
Methods
S100A4, A9 and A12 mRNA levels in granulocytes and HEL cells are measured by real time PCR, while protein expression of S100A4, A8, A9 and A12 are examined in granulocytes and plasma of MPN using immunoblotting and immunoassay, respectively. In addition, S100A9 is determined by immunohistochemistry in bone marrow. Also, levels of S100A4, A9 and A12 are measured by real time PCR and immunoblotting in HEL cells incubated 48 hours with hydroxyurea and specific JAK2 inhibitor 1,2,3,4,5,6-hexabromocyclohexane (HBC). Mutations of JAK2V617F and CALR exon 9 are analyzed by DNA sequencing and allelic PCR.
Results
S100A12 mRNA level is significantly increased in JAK2V617F heterozygous ET patients, but downregulated in JAK2V617F heterozygous PV patients and PMF without mutation (p<0.05). However at protein level, S100A12 is significantly increased only in granulocytes of ET patients without mutation (p<0.05) and plasma of PV and PMF without JAK2V617F mutation (70 ng/ml, p<0.01). Besides non significant modification at mRNA levels, S100A4 and S100A9 protein expression demonstrated a common significant increase in granulocytes (p<0.01), followed by increased quantity of S100A9-positive cells in bone marrow and S100A8/A9 proteins in plasma of MPN patients (51 and 28 ng/ml, p<0.01). Presence of CALR mutation augmented S100A8/A9 levels in plasma and granulocytes of ET and PMF patients. After 48 hour, hydroxyurea and specific JAK2 inhibitor HBC significantly decreased S100A4 mRNA levels in HEL cells (p<0.01). Further on, protein expression of S100A4 and S100A9 are also significantly downregulated by hydroxyurea and JAK2 inhibitor HBC in HEL cells (p<0.05).
Conclusion
S100A4, A9 and A12 mRNA and protein levels are not generally influenced by JAK2V617F mutant allele burden. S100A4 and anti-inflammatory S100A8/A9 protein levels demonstrated stable elevation in contrast to sporadic pro-inflammatory S100A12 appearance in MPNs. Moreover, JAK2 inhibition reduced S100A4 and S100A9 levels in HEL cells. S100A8/A9 could serve as a clinical biomarker and therapeutic target in MPNs, with existing S100A8/A9 blockers permitted for clinical testing.
Session topic: E-poster
Keyword(s): Inflammation, Janus Kinase inhibitor, Myeloproliferative disorder
Abstract: PB2012
Type: Publication Only
Background
Chronic inflammation is associated with sustained myeloproliferation, while S100 proteins have been shown to regulate cell proliferation, differentiation and inflammation in pathological conditions. S100A8, A9 and A12 produced by cells of myeloid origin were mediators of inflammation, while S100A4 regulated cell proliferation in neoplasia. S100A4 and A12 are produced via activation of the JAK-STAT pathway, constitutively activated in myeloproliferative neoplasm (MPN).
Aims
This study analyzed the inflammation linked S100 proteins in MPNs: polycythemia vera (PV, n=16), essential thrombocythemia (ET, n=16), primary myelofibrosis (PMF, n=20), according to JAK2V617F and CALR mutation status, and healthy controls (n=8), and their regulation in JAK2V617F mutated human erythroleukemia cell line (HEL) during MPN simulated therapy.
Methods
S100A4, A9 and A12 mRNA levels in granulocytes and HEL cells are measured by real time PCR, while protein expression of S100A4, A8, A9 and A12 are examined in granulocytes and plasma of MPN using immunoblotting and immunoassay, respectively. In addition, S100A9 is determined by immunohistochemistry in bone marrow. Also, levels of S100A4, A9 and A12 are measured by real time PCR and immunoblotting in HEL cells incubated 48 hours with hydroxyurea and specific JAK2 inhibitor 1,2,3,4,5,6-hexabromocyclohexane (HBC). Mutations of JAK2V617F and CALR exon 9 are analyzed by DNA sequencing and allelic PCR.
Results
S100A12 mRNA level is significantly increased in JAK2V617F heterozygous ET patients, but downregulated in JAK2V617F heterozygous PV patients and PMF without mutation (p<0.05). However at protein level, S100A12 is significantly increased only in granulocytes of ET patients without mutation (p<0.05) and plasma of PV and PMF without JAK2V617F mutation (70 ng/ml, p<0.01). Besides non significant modification at mRNA levels, S100A4 and S100A9 protein expression demonstrated a common significant increase in granulocytes (p<0.01), followed by increased quantity of S100A9-positive cells in bone marrow and S100A8/A9 proteins in plasma of MPN patients (51 and 28 ng/ml, p<0.01). Presence of CALR mutation augmented S100A8/A9 levels in plasma and granulocytes of ET and PMF patients. After 48 hour, hydroxyurea and specific JAK2 inhibitor HBC significantly decreased S100A4 mRNA levels in HEL cells (p<0.01). Further on, protein expression of S100A4 and S100A9 are also significantly downregulated by hydroxyurea and JAK2 inhibitor HBC in HEL cells (p<0.05).
Conclusion
S100A4, A9 and A12 mRNA and protein levels are not generally influenced by JAK2V617F mutant allele burden. S100A4 and anti-inflammatory S100A8/A9 protein levels demonstrated stable elevation in contrast to sporadic pro-inflammatory S100A12 appearance in MPNs. Moreover, JAK2 inhibition reduced S100A4 and S100A9 levels in HEL cells. S100A8/A9 could serve as a clinical biomarker and therapeutic target in MPNs, with existing S100A8/A9 blockers permitted for clinical testing.
Session topic: E-poster
Keyword(s): Inflammation, Janus Kinase inhibitor, Myeloproliferative disorder
Type: Publication Only
Background
Chronic inflammation is associated with sustained myeloproliferation, while S100 proteins have been shown to regulate cell proliferation, differentiation and inflammation in pathological conditions. S100A8, A9 and A12 produced by cells of myeloid origin were mediators of inflammation, while S100A4 regulated cell proliferation in neoplasia. S100A4 and A12 are produced via activation of the JAK-STAT pathway, constitutively activated in myeloproliferative neoplasm (MPN).
Aims
This study analyzed the inflammation linked S100 proteins in MPNs: polycythemia vera (PV, n=16), essential thrombocythemia (ET, n=16), primary myelofibrosis (PMF, n=20), according to JAK2V617F and CALR mutation status, and healthy controls (n=8), and their regulation in JAK2V617F mutated human erythroleukemia cell line (HEL) during MPN simulated therapy.
Methods
S100A4, A9 and A12 mRNA levels in granulocytes and HEL cells are measured by real time PCR, while protein expression of S100A4, A8, A9 and A12 are examined in granulocytes and plasma of MPN using immunoblotting and immunoassay, respectively. In addition, S100A9 is determined by immunohistochemistry in bone marrow. Also, levels of S100A4, A9 and A12 are measured by real time PCR and immunoblotting in HEL cells incubated 48 hours with hydroxyurea and specific JAK2 inhibitor 1,2,3,4,5,6-hexabromocyclohexane (HBC). Mutations of JAK2V617F and CALR exon 9 are analyzed by DNA sequencing and allelic PCR.
Results
S100A12 mRNA level is significantly increased in JAK2V617F heterozygous ET patients, but downregulated in JAK2V617F heterozygous PV patients and PMF without mutation (p<0.05). However at protein level, S100A12 is significantly increased only in granulocytes of ET patients without mutation (p<0.05) and plasma of PV and PMF without JAK2V617F mutation (70 ng/ml, p<0.01). Besides non significant modification at mRNA levels, S100A4 and S100A9 protein expression demonstrated a common significant increase in granulocytes (p<0.01), followed by increased quantity of S100A9-positive cells in bone marrow and S100A8/A9 proteins in plasma of MPN patients (51 and 28 ng/ml, p<0.01). Presence of CALR mutation augmented S100A8/A9 levels in plasma and granulocytes of ET and PMF patients. After 48 hour, hydroxyurea and specific JAK2 inhibitor HBC significantly decreased S100A4 mRNA levels in HEL cells (p<0.01). Further on, protein expression of S100A4 and S100A9 are also significantly downregulated by hydroxyurea and JAK2 inhibitor HBC in HEL cells (p<0.05).
Conclusion
S100A4, A9 and A12 mRNA and protein levels are not generally influenced by JAK2V617F mutant allele burden. S100A4 and anti-inflammatory S100A8/A9 protein levels demonstrated stable elevation in contrast to sporadic pro-inflammatory S100A12 appearance in MPNs. Moreover, JAK2 inhibition reduced S100A4 and S100A9 levels in HEL cells. S100A8/A9 could serve as a clinical biomarker and therapeutic target in MPNs, with existing S100A8/A9 blockers permitted for clinical testing.
Session topic: E-poster
Keyword(s): Inflammation, Janus Kinase inhibitor, Myeloproliferative disorder
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