OBJECTIVETo explore the expression of tyrosine phosphatase containing C-src homology SH-2 (SHP-1 and SHP-2) in benign prostate hyperplasia.

METHODSWith En Vision two-step method, the expression of SHP-1 and SHP-2 was detected in 10 cases of normal prostate tissue, 30 cases of BPH, 20 cases of PIN, 20 cases of high differential Pca and 20 cases of low differential Pca.

RESULTThe expression of SHP-2 in normal group was mainly distributed in the cytoplasm of secretive cells and basal cells, and a little part in the nucleu. In BPH it was distributed equally in the plasm and nucleu. In PIN, high differential Pca and low differential Pca, SHP-2 expressed mainly in nucleu. The average dyeing index of SHP-2 in each group is 0.4, 1.7, 2.1, 2.2 and 2.6. SHP-1 positive expression in normal prostate, BPH, PIN and high differential Pca showed differentiating layer staining in the cytoplasm of secretive cells and basal cells, while not in low differential Pca. The average dyeing index of SHP-1 in each group is 1.8, 1.8, 1.5, 1.2 and 0.4.

CONCLUSIONThere are transformation in signal transduction relation with SHP-1 and SHP-2 in the progress of prostate cell proliferation, differentiation and malignant. The abnormal activation and distribution of SHP-2 might induce prostate reconstruction and hyperplasia, even carcinoma.

Adult ; Aged ; Cell Nucleus ; enzymology ; Cytoplasm ; enzymology ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Prostatic Hyperplasia ; enzymology ; pathology ; Protein Tyrosine Phosphatase, Non-Receptor Type 11 ; metabolism ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; metabolism ; Protein Tyrosine Phosphatases ; metabolism ; SH2 Domain-Containing Protein Tyrosine Phosphatases ; metabolism ; src-Family Kinases ; metabolism

Diabetes mellitus is caused by deficiency of insulin secretion from the pancreatic islet beta cells and/or insulin resistance in liver, muscle and adipocytes, resulting in glucose intolerance and hyperglycemia. Several protein tyrosine phosphatases, such as PTP1B (PTPN1), TCPTP (PTPN2), LYP (PTPN22), PTPIA-2, PTPMEG2 (PTPN9) or OSTPTP are involved in insulin signaling pathway, insulin secretion and autoreactive attack to pancreatic beta cells. Genetic mutation or overexpression of these phosphotases has been found to cause or increase the risk of diabetes mellitus. Some population with high risk for type 2 diabetes has overexpressed PTP1B, a prototypical tyrosine phosphatase which down-regulates insulin and leptin signal transduction. Animal PTP1B knockout model and PTP1B specific inhibitor cellular studies indicate PTP1B may serve as a therapeutic target for type 2 diabetes. TCPTP shares more than 70% sequence identity with PTP1B in their catalytic domain. TCPTP dephosphorylates tyrosine phosphorylated substrates overlapping with PTP1B but also has its own distinct dephosphorylation sites and functions. Recent research indicates TCPTP may have role in type 1 diabetes via dysregultaion of cytokine-mediated immune responses or pancreatic beta cell apoptosis. The tyrosine phosphatase LYP, which down-regulates LCK activity in T cell response, can become mutated as R620W which is highly correlated to type 1 diabetes. LYP R620W may be a gain of function mutation which suppresses TCR signaling. Patients bearing the R620W mutant have impaired T cell responses and increased populations of (CD45RO+CD45RA-) CD4+ T cells. A detailed elucidation of mechanism of R620W in type 1 diabetes and specific LYP inhibitor development will help characterize LYP R620W as a therapeutic target. A receptor tyrosine phosphatase, PTPIA-2/beta is a major autoantigen of type 1 diabetes. A diagnosis kit identifying PTPIA-2/beta autoantibodies is valuable in early detection and prevention of type 1 diabetes. In addition, other phosphatase like OSTPTP and PTPMEG2 are involved in type 2 diabetes via regulation of insulin production, beta cell growth or insulin signaling. Research into understanding the mechanism of these tyrosine phosphatases in diabetes, such as their precise functions in the regulation of insulin secretion, the insulin response and the immune response will strengthen our knowledge of diabetes pathophysiology which may result in new diagnostic and therapeutic strategies for diabetes.
Animals ; Diabetes Mellitus ; enzymology ; Diabetes Mellitus, Type 1 ; enzymology ; Diabetes Mellitus, Type 2 ; enzymology ; Humans ; Protein Tyrosine Phosphatase, Non-Receptor Type 1 ; genetics ; metabolism ; Protein Tyrosine Phosphatase, Non-Receptor Type 2 ; genetics ; metabolism ; Protein Tyrosine Phosphatase, Non-Receptor Type 22 ; genetics ; metabolism ; Protein Tyrosine Phosphatases, Non-Receptor ; classification ; genetics ; metabolism

OBJECTIVETo investigate the effects of protein tyrosine phosphatase-SHP2 and dual-specificity MAPK phosphatase-MKP5 on the activation of MAPKs and cell invasion induced by P2Y purinergic receptor in human prostate cancer cell lines with different metastatic potentials.

METHODSThe wide type (-wt) SHP2, mutant type (-cs) SHP2 and wide type (-wt) MKP5 cDNA expression vectors were constructed and stably transfected into 1E8 cells (highly metastatic) and/or 2B4 cells (non-metastatic). The tyrosine phosphorylation of SHP2 was examined by immunoprecipitation. The activation of ERK1/2 and p38 induced by P2Y receptor agonist ATP was analyzed by Western blot with phospho-specific antibodies against the dually phosphorylated, active forms of ERK1/2 and p38. The in-vitro invasive ability through Matrigel was measured by boyden-chamber assay.

RESULTSATP induced significant SHP2 phosphorylation, which was stronger and lasted longer in 1E8 than in 2B4. SHP2-wt enhanced the ERK1/2 activation induced by ATP in 2B4 cells, while SHP2-cs delayed and decreased this effect in 1E8 cells. Both SHP2-wt and SHP2-cs had no obvious influence on p38 activation. ATP stimulated cell invasion of both 1E8 and 2B4, while transfection of SHP2-wt into 2B4 cells further increased the invasive-stimulating ability of ATP (18.7% increase compared with ATP treatment alone). Transfection of SHP2-cs into 1E8 cells, however, antagonized the invasive-stimulating ability of ATP (40.9% decrease compared with ATP treated group). Up-regulation of MKP5-wt inhibited phosphorylation of p38 by ATP and reduced cell invasion stimulated by ATP (22.4% and 28.7% decrease compared with ATP treated group of 1E8 and 2B4, respectively).

CONCLUSIONSBoth SHP2 and MKP5 play some roles in P2Y receptor-mediated activation of MEK/ERK, p38 signaling pathways and prostate cancer invasion. SHP2 positively regulates ERK activation and prostate cancer invasion, whereas MKP5 inhibits the invasion by suppressing p38 activation.

Adenosine Triphosphate ; pharmacology ; Cell Line, Tumor ; DNA, Complementary ; genetics ; Dual-Specificity Phosphatases ; Genetic Vectors ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; Male ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Mitogen-Activated Protein Kinase Phosphatases ; Neoplasm Invasiveness ; Phosphorylation ; Prostatic Neoplasms ; metabolism ; pathology ; Protein Tyrosine Phosphatase, Non-Receptor Type 11 ; Protein Tyrosine Phosphatases ; genetics ; metabolism ; Receptors, Purinergic P2 ; physiology ; Signal Transduction ; Transfection ; p38 Mitogen-Activated Protein Kinases ; metabolism

BACKGROUNDFetal insulin hypothesis was proposed that the association between low birth weight and type 2 diabetes is principally genetically mediated. The aim of this study was to investigate whether common variants in genes CDKAL1, HHEX, ADCY5, SRR, PTPRD that predisposed to type 2 diabetes were also associated with reduced birthweight in Chinese Han population.

METHODSTwelve single nucleotide polymorphisms (rs7756992/rs10946398 in CDKAL1, rs1111875 in HHEX, rs391300 in SRR, rs17584499 in PTPRD, rs1170806/rs9883204/rs4678017/rs9881942/rs7641344/rs6777397/rs6226243 in ADCY5) were genotyped in 1174 unrelated individuals born in Peking Union Medical College Hospital from 1921 to 1954 by TaqMan allelic discrimination assays, of which 645 had normal glucose tolerance, 181 had developed type 2 diabetes and 348 impaired glucose regulation. Associations of these 12 genetic variants with birthweight and glucose metabolism in later life were analyzed.

RESULTSBirthweight was inversely associated with CDKAL1-rs10946398 (β = -41 g [95% confidence interval [CI]: -80, -3], P = 0.034), common variants both associated with increased risk of impaired glucose metabolism and decreased insulin secretion index later in life. After adjusting for sex, gestational weeks, parity and maternal age, the risk allele of CDKAL1-rs7756992 was associated with reduced birthweight (β = -36 g [95% CI: -72, -0.2], P = 0.048). The risk allele in SRR showed a trend toward a reduction of birthweight (P = 0.085).

CONCLUSIONSThis study identified the association between type 2 diabetes risk variants in CDKAL1 and birthweight in Chinese Han individuals, and the carrier of risk allele within SRR had the trend of reduced birthweight. This demonstrates that there is a clear overlap between the genetics of type 2 diabetes and fetal growth, which proposes that lower birth weight and type 2 diabetes may be two phenotypes of one genotype.

Adenylyl Cyclases ; genetics ; Aged ; Alleles ; Asian Continental Ancestry Group ; genetics ; Birth Weight ; genetics ; Cyclin-Dependent Kinase 5 ; genetics ; Diabetes Mellitus, Type 2 ; genetics ; Female ; Genetic Predisposition to Disease ; genetics ; Homeodomain Proteins ; genetics ; Humans ; Infant, Low Birth Weight ; Male ; Middle Aged ; Polymorphism, Single Nucleotide ; genetics ; Receptor-Like Protein Tyrosine Phosphatases, Class 2 ; genetics ; Transcription Factors ; genetics ; tRNA Methyltransferases

Protein tyrosine phosphatases (PTPs) are key regulatory factors in inflammatory signaling pathways. Although PTPs have been extensively studied, little is known about their role in neuroinflammation. In the present study, we examined the expression of 6 different PTPs (PTP1B, TC-PTP, SHP2, MEG2, LYP, and RPTPβ) and their role in glial activation and neuroinflammation. All PTPs were expressed in brain and glia. The expression of PTP1B, SHP2, and LYP was enhanced in the inflamed brain. The expression of PTP1B, TC-PTP, and LYP was increased after treating microglia cells with lipopolysaccharide (LPS). To examine the role of PTPs in microglial activation and neuroinflammation, we used specific pharmacological inhibitors of PTPs. Inhibition of PTP1B, TC-PTP, SHP2, LYP, and RPTPβ suppressed nitric oxide production in LPS-treated microglial cells in a dose-dependent manner. Furthermore, intracerebroventricular injection of PTP1B, TC-PTP, SHP2, and RPTPβ inhibitors downregulated microglial activation in an LPS-induced neuroinflammation model. Our results indicate that multiple PTPs are involved in regulating microglial activation and neuroinflammation, with different expression patterns and specific functions. Thus, PTP inhibitors can be exploited for therapeutic modulation of microglial activation in neuroinflammatory diseases.
Brain ; Microglia ; Neuroglia ; Nitric Oxide ; Protein Tyrosine Phosphatase, Non-Receptor Type 2 ; Protein Tyrosine Phosphatases*

To obtain higher potency and specificity, a series of 7-alkoxy analogues of illudalic acid was synthesized on the base of structure-activity relationship (SAR). All of these compounds exhibited submicromolar inhibition of the enzyme when tested against human leukocyte common antigen-related phosphatase (LAR) (for example, for 15e, IC50 = 180 nmol x L(-1)). They represent the most potent small-molecule inhibitors of LAR so far. These analogues also display excellent selectivity for LAR over other protein tyrosine phosphatases (PTPs) except for the highly homologous PTPsigma. The compound 15f is of 120-fold selectivity for LAR versus PTP-1B inhibition. The development of potent enzyme-specific inhibitors is so important that they may serve both as tools to study the role of LAR and as therapeutic agents for treatment of type II diabetes.
Coumarins ; chemical synthesis ; chemistry ; pharmacology ; Enzyme Inhibitors ; chemical synthesis ; chemistry ; pharmacology ; Inhibitory Concentration 50 ; Molecular Structure ; Receptor-Like Protein Tyrosine Phosphatases, Class 2 ; antagonists & inhibitors ; Structure-Activity Relationship

The hematopoietic cell phosphatase (HCP or SHP-1), the SH2 domain contain protein tyrosine phosphatase, is a crucial negative regulator in the process of hematopoietic cell development, proliferation and receptor-mediated mitogenic signaling pathways, and its mutation is responsible for the over-expansion and inappropriate activation of myelomonocytic population in motheaten mice. The aim of the study was to evaluate the role of the HCP gene in leukemogenesis. Bone marrow and/or peripheral blood from 32 acute myeloid leukemia (AML) patients, 9 acute lymphocytic leukemia (ALL) patients, 8 leukemia cell lines and 50 normal controls were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) based on single strand conformation polymorphism (SSCP) and sequencing. RT-PCR showed that all samples expressed HCP gene, only one missense mutation at codon 225 (AAC to AGC, Asn to Ser) within N-terminal SH2 domain was found in an ALL patient. In addition, four polymorphic base substitutions were detected in codon 69, 85, 86 and 266, respectively. In conclusion, mutation of HCP gene is an infrequent genetic aberration which may only play a role in pathogenesis of a small part of leukemia, however, its significance needs to be further clarified.
Acute Disease ; Cell Line, Tumor ; Humans ; Intracellular Signaling Peptides and Proteins ; Leukemia ; enzymology ; genetics ; Mutation ; Polymorphism, Single-Stranded Conformational ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; Protein Tyrosine Phosphatases ; genetics

OBJECTIVETo investigate the role of cytoplasmic domain of integrin alpha IIb in platelet signal transduction.

METHODSBinding capacity of integrin alpha IIb(R995A) to antibody platelet activation complex-1 (PAC-1) and pp125 focal adhesion kinase (FAK) phosphorylation of cells were detected by flow cytometry, immune precipitation, and Western blotting.

RESULTSWithout activation, wild-type alpha IIb beta3 Chinese hamster ovary (CHO) cells failed to bind to PAC-1, but mutant chimera alpha IIb(R995A)beta3 CHO cells were able to bind with PAC-1. Furthermore, phosphorylation of pp125 (FAK) in wild-type alpha IIb beta3 CHO cells occured only when cells were adhered to fibrinogen, but could not be detected in bovine serum albumin suspension. However in the mutant chimera group, it could be detected in both conditions.

CONCLUSIONThe mutation in integrin alpha IIb(R995A) alters its affinity state as a receptor, thus also mediating cytoplasmic signal transduction leading to the phosphorylation of pp125 (FAK) without ligand binding.

Animals ; Blood Platelets ; metabolism ; CHO Cells ; Cell Adhesion ; Cricetinae ; Cricetulus ; Cytoplasm ; metabolism ; Dual Specificity Phosphatase 2 ; Focal Adhesion Kinase 1 ; Focal Adhesion Protein-Tyrosine Kinases ; Humans ; Phosphorylation ; Platelet Glycoprotein GPIIb-IIIa Complex ; genetics ; metabolism ; physiology ; Point Mutation ; Protein Phosphatase 2 ; Protein Tyrosine Phosphatases ; metabolism ; Protein-Tyrosine Kinases ; metabolism ; Signal Transduction ; Transfection

Cyclic ADP-ribose (cADPR) releases Ca²⁺ from ryanodine receptor (RyR)-sensitive calcium pools in various cell types. In cardiac myocytes, the physiological levels of cADPR transiently increase the amplitude and frequency of Ca²⁺ (that is, a rapid increase and decrease of calcium within one second) during the cardiac action potential. In this study, we demonstrated that cADPR levels higher than physiological levels induce a slow and gradual increase in the resting intracellular Ca²⁺ ([Ca²⁺](i)) level over 10 min by inhibiting the sarcoendoplasmic reticulum Ca²⁺ ATPase (SERCA). Higher cADPR levels mediate the tyrosine-dephosphorylation of α-actin by protein tyrosine phosphatase 1B (PTP1B) present in the endoplasmic reticulum. The tyrosine dephosphorylation of α-actin dissociates phospholamban, the key regulator of SERCA, from α-actin and results in SERCA inhibition. The disruption of the integrity of α-actin by cytochalasin B and the inhibition of α-actin tyrosine dephosphorylation by a PTP1B inhibitor block cADPR-mediated Ca²⁺ increase. Our results suggest that levels of cADPR that are relatively higher than normal physiological levels modify calcium homeostasis through the dephosphorylation of α-actin by PTB1B and the subsequent inhibition of SERCA in cardiac myocytes.
Action Potentials ; Adenosine Diphosphate* ; Adenosine Triphosphatases ; Calcium ; Cyclic ADP-Ribose ; Cytochalasin B ; Endoplasmic Reticulum ; Homeostasis ; Muscle Cells* ; Myocytes, Cardiac ; Protein Tyrosine Phosphatase, Non-Receptor Type 1* ; Protein Tyrosine Phosphatases* ; Reticulum ; Ryanodine Receptor Calcium Release Channel ; Tyrosine

OBJECTIVESTo obtain prokaryotic expressed IA-2 recombinant protein and to identify its immunological activity.

METHODSThe complimentary DNA (cDNA) coding for the intracytoplasmic part of IA-2 (IA-2ic) was amplified from human fetal brain RNA, and was subcloned into the PinPoint Xa-1 T vector to construct recombinant expression plasmid, and was then expressed in E. coli JM109 cells as a fusion protein with a biotinylated peptide sequence at the aminoterminus. The biotinylated fusion protein was then purified by affinity chromatography and was subsequently dialyzed. Finally, its immunogenicity was evaluated by enzyme linked immunosorbent assay (ELISA).

RESULTSThe purified IA-2ic fusion protein resolved on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) as a single Coomassie brilliant blue stained band with a molecular weight of 59 kDa and its immunogenicity was confirmed by ELISA.

CONCLUSIONSE. coli expressed IA-2ic fusion protein has immunological activity. It can be used for detection of IA-2 autoantibodies (IA-2A) and for further studies on type 1 diabetes in future.

Animals ; Autoantigens ; biosynthesis ; DNA, Complementary ; analysis ; Diabetes Mellitus, Type 1 ; immunology ; Escherichia coli ; genetics ; Humans ; Membrane Proteins ; biosynthesis ; genetics ; isolation & purification ; Plasmids ; Protein Tyrosine Phosphatase, Non-Receptor Type 1 ; Protein Tyrosine Phosphatases ; biosynthesis ; genetics ; isolation & purification ; Rabbits ; Receptor-Like Protein Tyrosine Phosphatases, Class 8 ; Recombinant Fusion Proteins ; biosynthesis ; immunology ; isolation & purification