BACKGROUND: Dipeptidyl peptidase 4/CD26 (DPP-4) is a widely expressed cell surface serine protease. DPP-4 inhibitors, one of common anti-diabetic agents play a protective role in bone metabolism in recent studies. A soluble form of DPP-4 is found in serum, and exhibits DPP-4 enzymatic activity. However, the physiological role of serum or soluble DPP-4 and its relationship with DPP-4 enzymatic function remain poorly understood. The aims of current study were to determine the association between serum DPP-4 activity and bone mineral density (BMD) in postmenopausal women. METHODS: We recruited data and serum samples from 124 consecutive healthy postmenopausal women aged >50 years. We divided study subjects into obese (body mass index [BMI] ≥25 kg/m2) and non-obese (BMI <25 kg/m2) postmenopausal women and examined the correlation between serum DPP-4 activity and clinical variables in each groups. RESULTS: A total of 124 postmenopausal women was enrolled, with a mean age of 59.9±7.1 years. The mean BMI of the study patients was 24.4±2.8 kg/m2. Regarding bone turnover markers, serum DPP-4 activity was positively correlated with serum calcium concentrations, intact parathyroid hormone, and serum C-telopeptide levels in all of the study subjects. However, there was no association between serum DPP-4 activity and BMD in the spine or femoral neck in all of the study subjects. Serum DPP-4 activity was negatively correlated (R=−0.288, P=0.038) with BMD of the spine in obese postmenopausal women. CONCLUSION: This study demonstrated for the first time that serum soluble DPP-4 activity was negatively correlated with BMD in obese postmenopausal women.
Biological Markers ; Bone Density* ; Calcium ; Dipeptidyl Peptidase 4 ; Female ; Femur Neck ; Humans ; Metabolism ; Parathyroid Hormone ; Postmenopause ; Serine Proteases ; Spine

OBJECTIVE: To observe the expression of proteinase transmembrane protease, serine 3 (TMPRSS3) in mouse cochlea, and to investigate the significance of TMPRSS3 in the inner ear. METHODS: The protein expression of TMPRSS3 in C57/BL mouse cochlea was identified and detected by immunohistochemistry and immunofluorescence. Different cochlear tissues, such as spiral ganglion neurons, corti organ, stria vascularis and so on, were separated to detect the gene expression of TMPRSS3 by real-time fluorescence quantitative polymerase chain reaction (qPCR). The cochlear tissues with different ages were collected and the expression of TMPRSS3 mRNA was detected by qPCR. RESULTS: TMPRSS3 was mainly expressed in the spiral ganglion neurons, and there was TMPRSS3 mRNA in the cochlea in groups with different age. The expression level of TMPRSS3 mRNA was much weaker. CONCLUSION The distribution of TMPRSS3 was observed in many regions of the mouse cochlea, but mainly in the spiral ganglion neurons. This indicates that TMPRSS3 may be involved in the physiological functional regulation of the spiral ganglion neurons.
Animals ; Cochlea ; metabolism ; Female ; Male ; Membrane Proteins ; genetics ; metabolism ; Mice ; Mice, Inbred C57BL ; RNA, Messenger ; genetics ; metabolism ; Serine Proteases ; genetics ; metabolism

OBJECTIVETo explore the significance of plasma levels of mannan-binding lectin (MBL)-associated serine protease 2 (MASP2) in children with upper respiratory tract infection (URTI).

METHODSA total of 103 children with URTI and 35 healthy children were examined for plasma levels of MASP2 and C-reactive protein (CRP). According to CRP levels, white blood cell count (WBC), stage of infection, and administration of treatments, the children with URTI were divided into the elevated CRP group (n=48) and the normal CRP group (n=54), elevated WBC group (n=61) and normal WBC group (n=40), the early stage of infection without treatment group (n=68) and mid-late stage of infection with treatment group (n=35).

RESULTSPlasma MASP2 levels was significantly higher in URTI group than in the healthy control group (P<0.001) and showed a close correlation with age (r=0.302, P<0.01). Plasma MASP2 level was significantly correlated with CRP level in elevated CRP group (r=0.310, P<0.05) but not in normal CRP group (P>0.05), correlated with WBC in elevated WBC group (r=0.392, P<0.01) but not in normal WBC group (P>0.05), and was significantly higher in early stage infection without treatment group than in mid-late stage of infection with treatment group (P<0.01). MASP2, MBL2 and CRP genes had a common binding site for the transcription factor HNF-4α.

CONCLUSIONSMASP2 may be an acute-phase protein, and its plasma level might serve as a new reference index in the diagnosis of URTI in children.

C-Reactive Protein ; metabolism ; Case-Control Studies ; Child ; Humans ; Leukocyte Count ; Mannose-Binding Protein-Associated Serine Proteases ; metabolism ; Respiratory Tract Infections ; blood

Aprotinin is a serine protease inhibitor and a powerful antifibrinolytic agent, derived from the inhibition of plasmin and kallikrein. Therefore, it is widely used in cardiopulmonary bypass surgery or major surgery for reducing bleeding and blood transfusion requirements. Aprotinin is rapidly eliminated from the circulation by glomerular filtration and is actively reabsorbed in the renal tubular system, where it is stored, metabolized, and eliminated over the following 5-6 days. Because of this metabolism, concerns have been raised regarding the possibility that aprotinin may impair renal function due to a toxic effects on proximal tubular cells. We report three cases of postoperative renal failure after aprotinin had been used during surgery. Two patients, Jehovah's Witnesses who refused blood transfusion, required hemodialysis. One patient, who underwent spinal orthopedic surgery, was administered aprotinin to reduce intraoperative blood loss, and developed acute renal dysfunction. The patient recovered after supportive therapy.
Acute Kidney Injury* ; Aprotinin* ; Blood Transfusion ; Cardiopulmonary Bypass ; Fibrinolysin ; Filtration ; Hemorrhage ; Humans ; Jehovah's Witnesses ; Kallikreins ; Metabolism ; Orthopedics ; Renal Dialysis ; Renal Insufficiency ; Serine Proteases

Proteases in the skin are essential to epidermal permeability barrier homeostasis. In addition to their direct proteolytic effects, certain proteases signal to cells by activating protease-activated receptors (PARs), the G-protein-coupled receptors. The expression of functional PAR-2 on human skin and its role in inflammation, pruritus, and skin barrier homeostasis have been demonstrated. Atopic dermatitis (AD) is a multifactorial inflammatory skin disease characterized by genetic barrier defects and allergic inflammation, which is sustained by gene-environmental interactions. Recent studies have revealed aberrant expression and activation of serine proteases and PAR-2 in the lesional skin of AD patients. The imbalance between proteases and protease inhibitors associated with genetic defects in the protease/protease inhibitor encoding genes, increase in skin surface pH, and exposure to proteolytically active allergens contribute to this aberrant protease/PAR-2 signaling in AD. The increased protease activity in AD leads to abnormal desquamation, degradation of lipid-processing enzymes and antimicrobial peptides, and activation of primary cytokines, thereby leading to permeability barrier dysfunction, inflammation, and defects in the antimicrobial barrier. Moreover, up-regulated proteases stimulate PAR-2 in lesional skin of AD and lead to the production of cytokines and chemokines involved in inflammation and immune responses, itching sensation, and sustained epidermal barrier perturbation with easier allergen penetration. In addition, PAR-2 is an important sensor for exogenous danger molecules, such as exogenous proteases from various allergens, and plays an important role in AD pathogenesis. Together, these findings suggest that protease activity or PAR-2 may be a future target for therapeutic intervention for the treatment of AD.
Anti-Infective Agents/pharmacology ; Dermatitis, Atopic/*enzymology ; Endopeptidases/metabolism ; Homeostasis ; Humans ; Hydrogen-Ion Concentration ; Inflammation ; Models, Biological ; Models, Genetic ; Peptide Hydrolases/*metabolism ; Receptor, PAR-2/*metabolism ; Serine Proteases/metabolism ; Signal Transduction ; Skin/enzymology/pathology ; Treatment Outcome

Mannan-binding lectin (MBL) is a soluble innate immune protein that binds to glycosylated targets. MBL acts as an opsonin and activates complement, contributing to the destruction and clearance of infecting microorganisms. Hepatitis C virus (HCV) encodes two envelope glycoproteins E1 and E2, expressed as non-covalent E1/E2 heterodimers in the viral envelope. E1 and E2 are potential ligands for MBL. Here we describe an analysis of the interaction between HCV and MBL using recombinant soluble E2 ectodomain fragment, the full-length E1/E2 heterodimer, expressed in vitro, and assess the effect of this interaction on virus entry. A binding assay using antibody capture of full length E1/E2 heterodimers was used to demonstrate calcium dependent, saturating binding of MBL to HCV glycoproteins. Competition with various saccharides further confirmed that the interaction was via the lectin domain of MBL. MBL binds to E1/E2 representing a broad range of virus genotypes. MBL was shown to neutralize the entry into Huh-7 cells of HCV pseudoparticles (HCVpp) bearing E1/E2 from a wide range of genotypes. HCVpp were neutralized to varying degrees. MBL was also shown to neutralize an authentic cell culture infectious virus, strain JFH-1 (HCVcc). Furthermore, binding of MBL to E1/E2 was able to activate the complement system via MBL-associated serine protease 2. In conclusion, MBL interacts directly with HCV glycoproteins, which are present on the surface of the virion, resulting in neutralization of HCV particles.
Binding, Competitive ; Glycosylation ; Hepacivirus ; genetics ; pathogenicity ; physiology ; Humans ; Mannose-Binding Lectin ; metabolism ; Mannose-Binding Protein-Associated Serine Proteases ; metabolism ; Monosaccharides ; metabolism ; Protein Binding ; Protein Multimerization ; Tumor Cells, Cultured ; Viral Envelope Proteins ; metabolism ; Virion ; pathogenicity ; physiology ; Virus Internalization

Earthworm fibrinolytic enzyme (EFE) is a group of protease having fibrinolytic and plasminogen-activator activities isolated from earthworm. Molecular biology research showed that there were 21 EFE coding sequences, in which only one sequence, AY438624, whose translated protein had similar N-terminal amino-acid sequence to EfP-I purified from Eisenia fetida. To obtain coding sequence of EfP-I , we designed specific primers according to 5' and 3' sequences of AY438624. A new DNA sequence was obtained by RT-PCR, sequence analysis showed that the protein translated from the coding sequence had identical N-terminal amino-acid sequence with EfP-I purified from Eisenia fetida and Lumbricus rubellus. Analysis by using ScanProsite prediction programs proved that the sequence had high similarity to AY438624 and belonged to trypsin family of serine protease. But there was difference between two sequences, that was there was a domain of characteristic amino acids of N-glycosylation site Asn-Xaa-Ser/Thr(N-x-S/T)in the new sequence (DQ418454). Then the expressed vector pMAL-c2X-Efp-I was constructed by cloning the gene into the plasmid pMAL-c2X, and was transformed to E. coli TB1. After induction and expression of the recombinant, the product MBP-EfP-I was purified by MBP affinity chromatography. Western blotting analysis showed that the product reacted with both anti-MBP and anti-EfP-I -1 serum. Casein plate test and fibrin plate test showed that the protein expressed had fibrinolytic activity.
Animals ; Biocatalysis ; Blotting, Western ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Fibrin ; metabolism ; Fibrinolysis ; Gene Expression Regulation, Enzymologic ; Oligochaeta ; enzymology ; genetics ; Recombinant Proteins ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Serine Proteases ; genetics ; metabolism

Serine proteases form one of the most important families of enzymes and perform significant functions in a broad range of biological processes, such as intra- and extracellular protein metabolism, digestion, blood coagulation, regulation of development, and fertilization. A number of serine proteases have been identified in parasitic helminths that have putative roles in parasite development and nutrition, host tissues and cell invasion, anticoagulation, and immune evasion. In this review, we described the serine proteases that have been identified in parasitic helminths, including nematodes (Trichinella spiralis, T. pseudospiralis, Trichuris muris, Anisakis simplex, Ascaris suum, Onchocerca volvulus, O. lienalis, Brugia malayi, Ancylostoma caninum, and Steinernema carpocapsae), cestodes (Spirometra mansoni, Echinococcus granulosus, and Schistocephalus solidus), and trematodes (Fasciola hepatica, F. gigantica, and Schistosoma mansoni). Moreover, the possible biological functions of these serine proteases in the endogenous biological phenomena of these parasites and in the host-parasite interaction were also discussed.
Animals ; Cestoda/classification/*enzymology/growth & development/physiology ; Host-Parasite Interactions ; Life Cycle Stages ; Nematoda/classification/*enzymology/growth & development/physiology ; Serine Proteases/genetics/*metabolism ; Trematoda/classification/*enzymology/growth & development/physiology

OBJECTIVETo understand the role that esophageal cancer related gene-1 (ECRG-1) plays and to search for ECRG-1-interacting proteins.

METHODSA DNA fragment encoding the carboxy-terminus of ECRG-1 (amino acids 40 - 418) was inserted into pGBKT7-DNA-BD vector and fused in-frame to the DNA-binding domain of GAL4. Then, it was used as a bait to screen the human fetal liver cDNA library by yeast two-hybrid, with the cDNA fragment inserted into pACT2 vector and fused in-frame to the Gal4 activation domain. If ECRG-1 interacted with a protein encoded by a cDNA fragmant in the yeast, the transcription of reporter Gene could be activated. With the false positive clonies eliminated, the inserts in the positive plasmids were sequenced and compared to those in the GenBank.

RESULTSIn approximately 3 x 10(6) independent tansformants screened, 23 clonies exhibited the expression of reporter gene. After eliminating the false positive clonies, two cDNA fragments were obtained. DNA sequencing revealed that one encoded Miz-1 (Myc-interacting Zn finger protein-1), and another encoded FLNA (actin-binding protein-280), Miz-1, being a Zn finger protein, could be bound to p15 promotor and activated the transcription. FLNA, being an actin-binding protein took part in the TGF-beta pathway via interaction with Smad.

CONCLUSIONECRG-1 is able to be specifically bound to Miz-1 and FLNA in the yeast. It may play a role in the regulation of cell cycle via interaction with Miz-1 and FLNA.

Contractile Proteins ; metabolism ; DNA-Binding Proteins ; metabolism ; Escherichia coli Proteins ; metabolism ; physiology ; Filamins ; Gene Library ; Humans ; Kruppel-Like Transcription Factors ; Liver ; embryology ; physiology ; Membrane Proteins ; Microfilament Proteins ; metabolism ; Recombinant Fusion Proteins ; metabolism ; physiology ; Serine Proteases ; Transcription Factors ; Two-Hybrid System Techniques ; Yeasts ; genetics

The deubiquitinating enzyme ubiquitin specific peptidase 15 (USP15) is regarded as a regulator of TGFβ signaling pathway. This process depends on Smad7, the inhibitory factor of the TGFβ signal, and type I TGFβ receptor (TβR-I), one of the receptors of TGFβ. The expression level of USP15 seems to play vital roles in the pathogenesis of many neoplasms, but so far there has been no report about USP15 in psoriasis. In this study, immunohistochemical staining of USP15, TβR-I and Smad7 was performed in 30 paraffin-embedded psoriasis specimens and 10 normal specimens to investigate the expression of USP15, TβR-I and Smad7 in psoriasis and to explore the relevance among them. And USP15 small interfering RNA (USP15 siRNA) was used to transfect Hacat cells to detect the mRNA expression of TβR-I and Smad7. Of 30 cases of psoriasis in active stage, 28, 24 and 26 cases were positive for USP15, TβR-I and Smad7 staining, respectively. The positive rates of USP15 and Smad7 were significantly higher in psoriasis specimens than in normal skin specimens (44.1%±26.0% vs. 6.1%±6.6%, 47.2%±27.1% vs. 6.6%±7.1%), and positive rate of TβR-I (20.3%±22.2%) in psoriasis was lower than that in normal skin specimens (46.7%±18.2%). There was a significant positive correlation between USP15 and Smad7 expression, and significant negative correlations between USP15 and TβR-expression, an I d between TβR- and Smad7 expression I in psoriasis. After transfection of USP15 siRNA in Hacat cells, the expression of TβR-mRNA was up I -regulated and that of Smad7 was down-regulated. It is concluded that USP15 may play a role in the pathogenesis of psoriasis through regulating the TβR-I/Smad7 pathway and there may be other cell signaling pathways interacting with USP15 to take part in the development of psoriasis.
Adult ; Cell Line ; Female ; Gene Expression ; Humans ; Immunohistochemistry ; Keratinocytes ; cytology ; metabolism ; Male ; Middle Aged ; Protein-Serine-Threonine Kinases ; biosynthesis ; genetics ; Psoriasis ; genetics ; metabolism ; RNA Interference ; Receptors, Transforming Growth Factor beta ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; genetics ; Skin ; metabolism ; Smad7 Protein ; biosynthesis ; genetics ; Ubiquitin-Specific Proteases ; biosynthesis ; genetics ; Young Adult