PURPOSE: The dipeptidyl peptidase IV (DPPIV) gene family exhibits multiple functions and is involved in the pathogenesis of various diseases. It has attracted pharmaceutical interest in the areas of metabolic disorders as well as cancer. However, clinicopathologic significance of DPPIV family in colorectal cancer is not fully understood. MATERIALS AND METHODS: The clinical relevance of DPPIV and DPP10 expression was determined by immunohistochemical staining, and by assessing its clinicopathologic correlation in 383 colorectal cancer patients with known clinical outcomes. RESULTS: DPPIV was not expressed in normal colon mucosa, but it showed luminal expression in 52 of the 383 colorectal cancers (13.5%). DPPIV expression in tumors was associated with right-sided location of the colon (p=0.010) and more advanced tumor stage (p=0.045). DPP10 was expressed in normal colonic mucosa, but its expression varied in primary colorectal cancer tissues. Loss of DPP10 expression was found in 11 colorectal cancers (CRCs) (2.9%), and multivariate analysis showed that loss of DPP10 expression was an independent factor for poor patient prognosis (p=0.008). CONCLUSION: DPP10 may play a role in disease progression of colorectal cancer and loss of DPP10 expression in primary CRC is significantly associated with poor survival outcomes.
Colorectal Neoplasms/*metabolism/*pathology ; Dipeptidyl-Peptidases and Tripeptidyl-Peptidases/*metabolism ; Disease Progression ; Female ; Humans ; Immunohistochemistry ; Male ; Prognosis ; Tumor Markers, Biological/*metabolism

A trienzyme extraction method (use of alpha-amylase, protease and folate conjugase) for food folate assay has been used to release folate from the food matrix. In order to reduce the incubation time with three enzymes, folate values were compared between two incubation protocols; separate incubation (SI, incubated with alpha-amylase and conjugase separately for 2 hours after protease treatment) and combined incubation (CI, incubated with alpha-amylase and conjugase together for 2 hours after protease treatment) using 88 food items from 12 kinds of fast foods and processed foods. We found that folate values by CI were comparable to or higher than those by SI, indicating that CI might be a better extraction procedure to shorten the entire incubation time. We measured folate contents in 49 fast foods and 26 processed foods by microbiological assay after CI. Mean folate contents of one serving of various burgers ranged from 43.1 to 62.0 microgram. One serving of French fries, pizza, sandwich and triangled kimbab contained a mean of 53.3, 28.4, 47.4, and 25.7 microgram of folate, respectively. Folate contents of non-alcoholic beverages were very low, ranging from 1.0 to 5.2 microgram/100 g. Some of our values were comparable to the values in the folate database published in Korean Nutrition Society, however, some of the published values were 140 times higher than the measured values in this study. Folate values measured by the more recent modifications here can be used to update Korean folate database to accurately estimate dietary folate intake
alpha-Amylases ; Beverages ; Fast Foods ; Folic Acid ; gamma-Glutamyl Hydrolase

OBJECTIVETo screen the molecular markers for refractory anemia with excess blasts in transformation (RAEB) in myelodysplastic syndromes (MDS) by serum proteome profiling.

METHODSThe serum protein were isolated from patients with RAEB, acute myeloid leukemia or normal subjects by 2-dimensional electrophoresis (2-DE), and the electrophoresis gels were obtained to identify the differentially reacting protein spots. The replica gels of the differentially reacting proteins were analyzed to locate the matching protein spots, which were identified by peptide mass fingerprint based on matrix-assisted laser desorption/ionization time of-flight mass spectrometry (MALDI-TOF-MS) and database searching.

RESULTSSeven differentially expressed proteins in RAEB were found by 2-DE. Of the 7 proteins, 4 were identified by MALDI-TOF-MS to have significantly differential expression in RAEB, including dipeptidyl peptidase (DPP/CD26), polymerase (DNA directed) kappa, PRO2044 and an albumin-like protein.

CONCLUSION2-DE-based serum proteome profiling helps identify serum proteomic biomarkers related to MDS. DDP/CD26 has increased expression in the serum in RAEB subtype MDS, suggesting its possible role in advanced MDS.

Anemia, Refractory, with Excess of Blasts ; blood ; genetics ; Bone Marrow ; pathology ; DNA-Directed DNA Polymerase ; blood ; Dipeptidyl-Peptidases and Tripeptidyl-Peptidases ; blood ; Female ; Humans ; Male ; Middle Aged ; Myelodysplastic Syndromes ; blood ; classification ; genetics ; Proteomics

BACKGROUNDAmyotrophic lateral sclerosis (ALS) is a progressive degenerative disease characterized by the loss of motor neurons in the spinal cord, brainstem, and cerebral cortex, which results in muscle weakness, atrophy. Sporadic ALS (SALS) accounts for about 90% of ALS cases, but the etiology is largely unknown. Most of the researchers consider it to be a complex disease. There have been several genome-wide association (GWA) studies reporting several single nucleotide polymorphisms (SNPs) which are susceptible to ALS, but no data of Asians (including Chinese) yet. We investigate whether the polymorphism of rs10260404 in DPP6 gene is associated with SALS in Chinese Han origin to compare the ethnic differences between Chinese Han origin and other populations.

METHODSThe genomic DNA was extracted from the leukocytes of whole blood samples in 58 Chinese Han patients with SALS and 52 healthy controls. The asymmetric PCR was processed in the presence of an unlabeled probe that contained the rs10260404 locus. The product was genotyped on a light scanner using high resolution melting method and some were confirmed with sequencing.

RESULTSThe rs10260404 polymorphism was in Hardy-Weinberg equilibrium in patients and controls. The CC genotype and the C allele were similar in patients compared with healthy subjects and not associated with an increased risk of Chinese SALS patients (chi(2) = 0.29, OR = 1.26, 95% CI 0.55 - 2.87, P > 0.05).

CONCLUSIONSThe rs10260404 is not associated with ALS susceptibility in Chinese people with Han origin which may be due to ethnic differences. More study with large number of cases in Chinese population is really necessary.

Adult ; Alleles ; Amyotrophic Lateral Sclerosis ; genetics ; Asian Continental Ancestry Group ; genetics ; Dipeptidyl-Peptidases and Tripeptidyl-Peptidases ; genetics ; Female ; Genotype ; Humans ; Male ; Middle Aged ; Nerve Tissue Proteins ; genetics ; Polymerase Chain Reaction ; Polymorphism, Genetic ; genetics ; Potassium Channels ; genetics ; Sequence Analysis, DNA ; Young Adult

AIMTo study the effect of recombinant hirudin (rH) on tPA-induced fibrinolysis and the possible mechanism of its action.

METHODSThe effect of rH on thrombin-fibrin complex (Th-Fn) was detected by 99mTc labeled rH. In the in vitro clot lysis, tPA as plasminogen activator, and recalcified plasma as plasminogen resource were used to study the influence of rH on fibrinolysis by detecting TAFIa, D-Dimer and FXIII.

RESULTSIn a canine model of femoral artery thrombosis, a clear radioactivity strip was imaged in 30 - 60 min on a part image, and the femoral vein thrombosis developed at 30 min. rH efficiently inhibited clot regeneration. Addition of TM could inhibit clot lysis obviously, and CPI could shorten the delay of clot lysis which due to TAFIa. There was a dose-dependent relationship with TM concentration and TAFI activation. FXIII activation was inhibited by low concentration of rH ( < or = 0.2 u x mL(-1)), and the level of fibrinolysis product, D-Dimer, increased.

CONCLUSIONrH could inhibit the thrombin binding to fibrin. rH inhibited the activation of TAFI and FXIII by combining with thrombin which resulted in enhancement of thrombolysis.

Animals ; Blood Coagulation ; drug effects ; Carboxypeptidase B2 ; metabolism ; Carboxypeptidases ; antagonists & inhibitors ; Dogs ; Factor XIII ; metabolism ; Femoral Artery ; Femoral Vein ; Fibrinolysis ; drug effects ; Fibrinolytic Agents ; pharmacology ; Hirudins ; genetics ; pharmacology ; Male ; Plant Proteins ; pharmacology ; Protease Inhibitors ; Recombinant Proteins ; pharmacology ; Thrombomodulin ; metabolism ; Thrombosis ; metabolism ; Venous Thrombosis ; metabolism

OBJECTIVETo investigate the distribution of &gamma;-glutamyl hydrolase gene (GGH) 452C/T genotype and allele frequency in children with acute leukemia (AL) and healthy children.

METHODSBone marrow samples from 92 children with AL and peripheral blood samples from 124 healthy children were obtained to prepare complementary DNAs (cDNAs). The cDNAs were analyzed for a GGH 452C/T polymorphism by reverse transcriptase-polymerase chain reaction-denaturing gradient gel electrophoresis (RT-PCR-DGGE) and direct sequencing.

RESULTSThe frequencies of the AL patients with TT, CT and CC genotypes were 2.2%, 13.0% and 84.8%, and the frequencies of the control children were 1.6%, 16.9% and 81.5%, respectively. There was no significant difference in GGH genotype or T allele frequency between the two groups (P> 0.05). However, the T allele frequency in Han Chinese children was significantly different from those reported in Japanese, Mexican and African-American populations.

CONCLUSIONThe frequency of 452C/T polymorphism of GGH gene in Han Chinese children has been determined. The results suggested that an ethnic difference may exist.

Acute Disease ; Base Sequence ; Child ; Child, Preschool ; Female ; Gene Frequency ; Genotype ; Humans ; Infant ; Infant, Newborn ; Leukemia ; enzymology ; genetics ; Male ; Polymorphism, Single Nucleotide ; gamma-Glutamyl Hydrolase ; genetics

OBJECTIVE: To explore the interaction between Tubulin beta 4B class IVb (TUBB4B) and Agtpbp1/cytosolic carboxypeptidase- like1 (CCP1) in mouse primary spermatocytes (GC-2 cells) and the role of TUBB4B in regulating the development of GC-2 cells. METHODS: Lentiviral vectors were used to infect GC-2 cells to construct TUBB4B knockdown and negative control (NC-KD) cells. The stable cell lines with TUBB4B overexpression (Tubb4b-OE) and the negative control (NC-OE) cells were screened using purinomycin. RT-qPCR and Western blotting were used to verify successful cell modeling and explore the relationship between TUBB4B and CCP1 expressions in GC-2 cells. The effects of TUBB4B silencing and overexpression on the proliferation and cell cycle of GC-2 cells were evaluated using CCK8 assay and flow cytometry. The signaling pathway proteins showing significant changes in response to TUBB4B silencing or overexpression were identified using Western blotting and immunofluorescence assay and then labeled for verification at the cellular level. RESULTS: Both TUBB4B silencing and overexpression in GC-2 cells caused consistent changes in the mRNA and protein expressions of CCP1 (<i>Pi> < 0.05). Similarly, TUBB4B expression also showed consistent changes at the mRNA and protein after CCP1 knockdown and restoration (<i>Pi> < 0.05). TUBB4B knockdown and overexpression had no significant effect on proliferation rate or cell cycle of GC-2 cells, but caused significant changes in the key proteins of the nuclear factor kappa-B (NF-κB) signaling pathway (p65 and p-p65) and the mitogen-activated protein kinase (MAPK) signaling pathway (ErK1/2 and p-Erk1/2) (<i>Pi> < 0.05); CCP1 knockdown induced significant changes in PolyE expression in GC-2 cells (<i>Pi> < 0.05). CONCLUSIONS TUBB4B and CCP1 interact via a mutual positive regulation mechanism in GC-2 cells. CCP-1 can deglutamize TUBB4B, and the latter is involved in the regulation of NF-κB and MAPK signaling pathways in primary spermatocytes.
Animals ; Male ; Mice ; GTP-Binding Proteins/metabolism* ; Mitogen-Activated Protein Kinases/metabolism* ; NF-kappa B/metabolism* ; RNA, Messenger ; Serine-Type D-Ala-D-Ala Carboxypeptidase/metabolism* ; Signal Transduction ; Spermatocytes ; Tubulin/genetics*

OBJECTIVE: Oxytocin antagonists maybe useful in inhibiting the uterine contractions of preterm labor. One such compound is TT-235. The purpose of this study was to compare the resistance of TT-235 and oxytocin to enzymatic degradation by oxytocinase in pregnant human. METHODS: Blood samples from pregnant women not in labor were incubated in vitro with known amounts of oxytocin and TT-235. Samples were collected at 0, 15, 30, 45 and 60 minute intervals for oxytocin analysis and at 0, 10, 60 and 360 minutes for TT-235 analysis. Oxytocin was analyzed by radioimmunoassay after extraction while TT-235 was analyzed by radioreceptor assay. RESULTS: In human blood, oxytocin was readily metabolized with greater than 83% disappearance over the 60 minute incubation period. In contrast, TT-235 was stable up to 360 minutes of incubation. CONCLUSION: This study suggests that: (1) blood from pregnant human does contain oxytocinase at least in vitro; and (2) TT-235 was resistant to enzymatic degradation by human blood, implying that this oxytocin antagonist may have prolonged activity in vivo in humans.
Cystinyl Aminopeptidase ; Female ; Humans ; Metabolism* ; Obstetric Labor, Premature ; Oxytocin* ; Plasma* ; Pregnancy ; Pregnant Women* ; Radioimmunoassay ; Radioligand Assay ; Uterine Contraction ; Uterus

Gestational diabetes insipidus, which occurs rarely during late pregnancy, may injure the mother and fetus neurologically. It takes place in about 4 of every 100,000 pregnancies. Increased placental-derived vasopressinase in late pregnancy markedly degrades vasopressin. The decreased vasopressin activity causes hypotonic polyuria, polydipsia, and dehydration. We report a woman with gestational diabetes insipidus who had no abnormal laboratory tests before developing symptoms. The diabetes insipidus was controlled well by administering nasal desmopressin (1.desamino.8.D.arginine vasopressin, DDAVP) followed by resolution of the signs and symptoms after delivery. Although gestational diabetes insipidus is rare, a prompt diagnosis and appropriate treatment to reduce the risks of maternal and fetal injury are important.
Cystinyl Aminopeptidase ; Deamino Arginine Vasopressin ; Dehydration ; Diabetes Insipidus ; Diabetes, Gestational ; Female ; Fetus ; Humans ; Hypogonadism ; Mitochondrial Diseases ; Mothers ; Ophthalmoplegia ; Polydipsia ; Polyuria ; Pregnancy ; Pregnancy Trimester, Third ; Vasopressins

OBJECTIVEo clone angiotensin-converting enzyme 2(ACE2) gene, to analyze its amino acids and nucleotides sequence and to investigate tissue distribution of ACE2 in adult mice.

METHODSThe full-length ACE2 encoding sequence was amplified from the RNA of mice kidney tissue by RT-PCR technique, cloned into plasmid pGEM-T easy, then subcloned into plasmid pcDNA3.1+. After identification of DNA sequence, the recombinant plasmid pmACE2 was transfected into Cos7 cells with lipofectin reagent. The transient expression of ACE2 molecule was detected by SDS-PAGE. Sequence analysis was conducted with CLUSTALX program. Tissue distribution of ACE2 in mice was detected by RT-PCR.

RESULTSA fragment about 2.6 kb was amplified and the recombinant plasmid pmACE2 was confirmed by two-enzyme digesting and DNA sequencing. The cloned DNA sequence was consistent with that previously reported, except for 3 variations: A701G, T1102C and T1330C. SDS-PAGE proved that expression of a soluble, truncated products form of ACE2 was a glycoprotein of approximately 80 kD in Cos7 cells. The predicted mice ACE2 sequence contained an N-terminal signal sequence (amino acid residues 1-18), a single HHEMGHIQ zinc-binding domain (amino acid residues 373-380) and C-terminal membrane anchor (amino acid residues 738-765). Mice ACE2 showed 84 % identity with that of human, and 90 % identity with that of rat. Expression of ACE2 was the greatest in lungs, hearts and kidneys, and moderate levels were also detected in testes and livers.

CONCLUSIONMice ACE2 gene has been cloned and successfully expressed in vitro. The tissue-specific expression of ACE2 in different species is not identical.

Amino Acid Sequence ; Animals ; Base Sequence ; Carboxypeptidases ; genetics ; metabolism ; Cloning, Molecular ; DNA, Complementary ; genetics ; Gene Expression ; Kidney ; metabolism ; Lung ; metabolism ; Male ; Mice ; Molecular Sequence Data ; Myocardium ; metabolism ; Peptidyl-Dipeptidase A ; Sequence Analysis ; Tissue Distribution