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Wei Zeng

Publications and source records attributed to Wei Zeng.

At least 19 recordsLinked to original sources
PubMed2026

Clinical and endocrine correlates of genetic etiologies in severe hypospadias: Study from 34 patients.

OBJECTIVE: Hypospadias is a prevalent congenital anomaly (0.3%-1.0%); however, severe hypospadias (defined as proximal cases with the meatus at the penoscrotal junction, scrotum, or perineum) is a rare and clinically challenging entity with a multifactorial etiology. This study aimed to characterize the interrelationships among the clinical, endocrine, and genetic profiles in children with severe hypospadias. MATERIALS AND METHODS: We conducted a comprehensive analysis of 34 male patients with severe hypospadias. Preoperative hormone levels were measured using two methods: chemiluminescent immunoassay for luteinizing hormone and follicle-stimulating hormone, and liquid chromatography-tandem mass spectrometry for testosterone (T), dihydrotestosterone (DHT), dehydroepiandrosterone (DHEA), 17α-hydroxyprogesterone (17α-OHP), and other steroids. Genetic analysis was conducted via whole exome sequencing. RESULTS: The diagnostic yield of clinically relevant genetic variants (including pathogenic and likely pathogenic, and variants of uncertain significance) in our cohort was 41.2% (14/34) of patients. Patients carrying these variants exhibited a more complex phenotypic profile compared to non-carriers, including a significantly higher rate of patients with ≥3 associated malformations and a greater prevalence of cryptorchidism. Furthermore, the group with clinically relevant variants showed selective elevations in adrenal-derived precursors, specifically 17α-OHP and DHEA. Correlation analysis revealed significant positive associations of both 17α-OHP levels and the T/DHT ratio with the number of associated malformations. CONCLUSION: This study reveals significant genetic heterogeneity in patients with severe hypospadias. Those carrying genetic variants was associated with more severe clinical phenotypes, while certain endocrine variations, including the elevation of adrenal-derived hormones, were also observed in this cohort.

Humans↗
PubMed2007

Metabolism and excretion of the dipeptidyl peptidase 4 inhibitor [14C]sitagliptin in humans.

The metabolism and excretion of [(14)C]sitagliptin, an orally active, potent and selective dipeptidyl peptidase 4 inhibitor, were investigated in humans after a single oral dose of 83 mg/193 muCi. Urine, feces, and plasma were collected at regular intervals for up to 7 days. The primary route of excretion of radioactivity was via the kidneys, with a mean value of 87% of the administered dose recovered in urine. Mean fecal excretion was 13% of the administered dose. Parent drug was the major radioactive component in plasma, urine, and feces, with only 16% of the dose excreted as metabolites (13% in urine and 3% in feces), indicating that sitagliptin was eliminated primarily by renal excretion. Approximately 74% of plasma AUC of total radioactivity was accounted for by parent drug. Six metabolites were detected at trace levels, each representing <1 to 7% of the radioactivity in plasma. These metabolites were the N-sulfate and N-carbamoyl glucuronic acid conjugates of parent drug, a mixture of hydroxylated derivatives, an ether glucuronide of a hydroxylated metabolite, and two metabolites formed by oxidative desaturation of the piperazine ring followed by cyclization. These metabolites were detected also in urine, at low levels. Metabolite profiles in feces were similar to those in urine and plasma, except that the glucuronides were not detected in feces. CYP3A4 was the major cytochrome P450 isozyme responsible for the limited oxidative metabolism of sitagliptin, with some minor contribution from CYP2C8.

Adenosine Deaminase↗
PubMed2007

Reversible movement of switch 1 loop of myosin determines actin interaction.

The conserved switch 1 loop of P-loop NTPases is implicated as a central element that transmits information between the nucleotide-binding pocket and the binding site of the partner proteins. Recent structural studies have identified two states of switch 1 in G-proteins and myosin, but their role in the transduction mechanism has yet to be clarified. Single tryptophan residues were introduced into the switch 1 region of myosin II motor domain and studied by rapid reaction methods. We found that in the presence of MgADP, two states of switch 1 exist in dynamic equilibrium. Actin binding shifts the equilibrium towards one of the MgADP states, whereas ATP strongly favors the other. In the light of electron cryo-microscopic and X-ray crystallographic results, these findings lead to a specific structural model in which the equilibrium constant between the two states of switch 1 is coupled to the strength of the actin-myosin interaction. This has implications for the enzymatic mechanism of G-proteins and possibly P-loop NTPases in general.

Actins↗
PubMed2006

Activation of EDTA-resistant gelatinases in malignant human tumors.

Among the many proteases associated with human cancer, seprase or fibroblast activation protein alpha, a type II transmembrane glycoprotein, has two types of EDTA-resistant protease activities: dipeptidyl peptidase and a 170-kDa gelatinase activity. To test if activation of gelatinases associated with seprase could be involved in malignant tumors, we used a mammalian expression system to generate a soluble recombinant seprase (r-seprase). In the presence of putative EDTA-sensitive activators, r-seprase was converted into 70- to 50-kDa shortened forms of seprase (s-seprase), which exhibited a 7-fold increase in gelatinase activity, whereas levels of dipeptidyl peptidase activity remained unchanged. In malignant human tumors, seprase is expressed predominantly in tumor cells as shown by in situ hybridization and immunohistochemistry. Proteins purified from experimental xenografts and malignant tumors using antibody- or lectin-affinity columns in the presence of 5 mmol/L EDTA were assayed for seprase activation in vivo. Seprase expression and activation occur most prevalently in ovarian carcinoma but were also detected in four other malignant tumor types, including adenocarcinoma of the colon and stomach, invasive ductal carcinoma of the breast, and malignant melanoma. Together, these data show that, in malignant tumors, seprase is proteolytically activated to confer its substrate specificity in collagen proteolysis and tumor invasion.

Animals↗
PubMed2006

Recombinant M. smegmatis vaccine targeted delivering IL-12/GLS into macrophages can induce specific cellular immunity against M. tuberculosis in BALB/c mice.

In the present study, we constructed a viable therapeutic vaccine of recombinant M. smegmatis mediated IL-12/GLS (granulysin) gene transfer into murine macrophages to exert the immunotherapy effects on the Mycobacterium tuberculosis infection. We tested this recombinant therapeutic vaccine in an in vivo study to determine its capability of stimulating host specific immune responses against M. tuberculosis. BALB/c mice intranasally immunized with the therapeutic vaccine developed an efficient Th1 protective immune response against M. tuberculosis which was equal to that of the BCG strain. Inoculation intranasally with this viable vaccine induced high level of serum IFN-gamma, IL-12 and IgG2a. The viable vaccine was capable of inducing purified protein derivative (PPD) antigen-specific splenocytes proliferation and IFN-gamma production from T cells in spleens of the immunized mice. In addition, intranasally inoculation with the viable vaccine can induce PPD antigen-specific sIgA production in the broncho-alveolar lavage fluid (BALF) of the immunized mice. No change of IL-4 level was found in all groups. The therapeutic mechanism of this viable vaccine against M. tuberculosis infection observed here appeared to be a result of the specific Th1 immune response activated by mycobacterium antigen from M. smegmatis and the expression of sIL-12/GLS in alveolar macrophages via the M. smegmatis-mediated gene transfer method. This research demonstrates that the therapeutic gene can be introduced into a host by viable mycobacteria works to induce the host specific immune response against M. tuberculosis infection in vivo. Since this therapeutic vaccine can strongly induce specific Th1 responses against M. tuberculosis in BALB/c mice and has no obviously harmfulness to the host simultaneously, the recombinant vaccine might be a potential candidate therapeutic vaccine against tuberculosis.

Animals↗
PubMed2006

Resonance energy transfer between green fluorescent protein variants: complexities revealed with myosin fusion proteins.

Green fluorescent protein and its variants are frequently used as Förster (fluorescence) resonance energy transfer (FRET) pairs to determine the proximity of protein domains. We prepared fusion proteins comprising yellow fluorescent protein-Dictyostelium myosin II motor domain-cyan fluorescent protein (YFP-myosin-CFP) and compared their FRET properties with an existing construct (GFP-myosin-BFP), containing a green fluorescent protein acceptor and blue fluorescent protein donor [Suzuki, Y., Yasunaga, T., Ohkura, R., Wakabayashi, T. and Sutoh, K. (1998) Nature 396, 380-383]. The latter construct showed an apparent 40% reduction in acceptor fluorescence on ATP addition, when excited via the donor, compared with the YFP-myosin-CFP constructs which showed a small increase (<or=5%). We propose that this disparity primarily arises from the differential response of GFP and YFP on intramolecular association with the donor probe. Studies with isolated GFP and YFP at high concentrations show that they dimerize with similar K(d) values but the spectrum shifts toward the protonated state only with GFP. On excitation at 380 nm, the protonated GFPH emits at 510 nm via excited-state proton transfer, giving the appearance of extensive FRET. These findings have important implications for FRET measurements using GFP-type probes because they give rise to changes in donor and acceptor emission ratios through processes other than FRET and complicate the extraction of the true degree of energy transfer from experimental data. Furthermore, the unknown orientation factor prevents the distance of the lever arm swing from being derived from these FRET changes.

Animals↗
PubMed2006

Effect of single oral doses of sitagliptin, a dipeptidyl peptidase-4 inhibitor, on incretin and plasma glucose levels after an oral glucose tolerance test in patients with type 2 diabetes.

CONTEXT: In response to a meal, glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP) are released and modulate glycemic control. Normally these incretins are rapidly degraded by dipeptidyl peptidase-4 (DPP-4). DPP-4 inhibitors are a novel class of oral antihyperglycemic agents in development for the treatment of type 2 diabetes. The degree of DPP-4 inhibition and the level of active incretin augmentation required for glucose lowering efficacy after an oral glucose tolerance test (OGTT) were evaluated. OBJECTIVE: The objective of the study was to examine the pharmacodynamics, pharmacokinetics, and tolerability of sitagliptin. DESIGN: This was a randomized, double-blind, placebo-controlled, three-period, single-dose crossover study. SETTING: The study was conducted at six investigational sites. PATIENTS: The study population consisted of 58 patients with type 2 diabetes who were not on antihyperglycemic agents. INTERVENTIONS: Interventions included sitagliptin 25 mg, sitagliptin 200 mg, or placebo. MAIN OUTCOME MEASURES: Measurements included plasma DPP-4 activity; post-OGTT glucose excursion; active and total incretin GIP levels; insulin, C-peptide, and glucagon concentrations; and sitagliptin pharmacokinetics. RESULTS: Sitagliptin dose-dependently inhibited plasma DPP-4 activity over 24 h, enhanced active GLP-1 and GIP levels, increased insulin/C-peptide, decreased glucagon, and reduced glycemic excursion after OGTTs administered at 2 and 24 h after single oral 25- or 200-mg doses of sitagliptin. Sitagliptin was generally well tolerated, with no hypoglycemic events. CONCLUSIONS: In this study in patients with type 2 diabetes, near maximal glucose-lowering efficacy of sitagliptin after single oral doses was associated with inhibition of plasma DPP-4 activity of 80% or greater, corresponding to a plasma sitagliptin concentration of 100 nm or greater, and an augmentation of active GLP-1 and GIP levels of 2-fold or higher after an OGTT.

Administration, Oral↗
PubMed2006

Cell movement during chick primitive streak formation.

Gastrulation in amniotes begins with extensive re-arrangements of cells in the epiblast resulting in the formation of the primitive streak. We have developed a transfection method that enables us to transfect randomly distributed epiblast cells in the Stage XI-XIII chick blastoderms with GFP fusion proteins. This allows us to use time-lapse microscopy for detailed analysis of the movements and proliferation of epiblast cells during streak formation. Cells in the posterior two thirds of the embryo move in two striking counter-rotating flows that meet at the site of streak formation at the posterior end of the embryo. Cells divide during this rotational movement with a cell cycle time of 6-7 h. Daughter cells remain together, forming small clusters and as result of the flow patterns line up in the streak. Expression of the cyclin-dependent kinase inhibitor, P21/Waf inhibits cell division and severely limits embryo growth, but does not inhibit streak formation or associated flows. To investigate the role off cell-cell intercalation in streak formation we have inhibited the Wnt planar-polarity signalling pathway by expression of a dominant negative Wnt11 and a Dishevelled mutant Xdd1. Both treatments do not result in an inhibition of streak formation, but both severely affect extension of the embryo in later development. Likewise inhibition of myosin II which as been shown to drive cell-cell intercalation during Drosophila germ band extension, has no effect on streak formation, but also effectively blocks elongation after regression has started. These experiments make it unlikely that streak formation involves known cell-cell intercalation mechanisms. Expression of a dominant negative FGFR1c receptor construct as well as the soluble extracellular domain of the FGFR1c receptor both effectively block the cell movements associated with streak formation and mesoderm differentiation, showing the importance of FGF signalling in these processes.

Animals↗
PubMed2006

Distinct roles for cysteine cathepsin genes in multistage tumorigenesis.

Multiple types of degradative enzymes, including cathepsins of the cysteine protease family, have been implicated in the regulation of angiogenesis and invasion during cancer progression. Several cysteine cathepsins are up-regulated in a mouse model of pancreatic islet cell carcinogenesis (RIP1-Tag2), and tumor progression is impaired following their collective pharmacologic inhibition. Using null mutations of four of the implicated cysteine cathepsins, we have now dissected their individual roles in cancer development. Mutants of cathepsins B or S impaired tumor formation and angiogenesis, while cathepsin B or L knockouts retarded cell proliferation and tumor growth. Absence of any one of these three genes impaired tumor invasion. In contrast, removal of cathepsin C had no effect on either tumor formation or progression. We have identified E-cadherin as a target substrate of cathepsins B, L, and S, but not cathepsin C, potentially explaining their differential effects on tumor invasion. Furthermore, we detected analogous increases in cathepsin expression in human pancreatic endocrine neoplasms, and a significant association between increased levels of cathepsins B and L and tumor malignancy. Thus individual cysteine cathepsin genes make distinctive contributions to tumorigenesis.

Animals↗
PubMed2006

A new approach for evaluating carryover and its influence on quantitation in high-performance liquid chromatography and tandem mass spectrometry assay.

In a high-performance liquid chromatography (HPLC)-based analytical method, carryover denotes one type of systematic error that is derived from a preceding sample and introduced into the next sample. For typical bioanalytical method development, a significant amount of time and resources are spent on reducing carryover for some analytes. In this paper, the statistical characteristics of carryover were analyzed based on the experimental results. The relative carryover (RC), defined as the peak area ratio of a blank sample to the preceding sample, was constant for the analyte and independent of the concentration of the preceding sample. The influence of carryover on the quantitation of the next injected sample or the 'following' sample was proportional to the concentration ratio of two consecutive samples and the relative carryover. Based on these experiments and analyses, the influence of carryover on the quantitation of unknown samples in an HPLC assay can be evaluated by the estimated carryover influence (ECI), which is the product of the relative carryover and the concentration ratio. This new approach provides a quantitative estimation for the influence of carryover on the quantitation of the unknown sample, and removes the limit put on the dynamic range of the assay by the current criterion of carryover. In general, if the relative standard deviation (RSD) of a validated bioanalytical method is less than 10%, the carryover will not have a significant effect on the accuracy of the assay when the estimated carryover influence is less than 5%.

Blood Chemical Analysis↗
PubMed2006

Determination of MK-0431 in human plasma using high turbulence liquid chromatography online extraction and tandem mass spectrometry.

A robust and sensitive method using high turbulence liquid chromatography (HTLC) online extraction with tandem mass spectrometry (MS/MS) for the determination of MK-0431 in human plasma was developed and validated to support the clinical studies. This HTLC online extraction method eliminated the time-consuming offline sample extraction procedures and significantly increased productivity. A narrow bore large particle size reversed-phase column (Cyclone, 50 x 1.0 mm, 60 microm) and a BDS Hypersil C18 column (30 x 2.1 mm, 3 microm) were used as extraction and analytical columns, respectively. The linear dynamic range of the calibration curve was 0.5 to 1000 ng/mL. Intraday validation was conducted using five calibration curves prepared in five lots of human control plasma, and the intraday precision (RSD%) was from 2.4 to 9.0% and the accuracy was from 98.0 to 103% of the nominal value. The intraday precision (RSD%, n = 5) for plasma quality control (QC) samples varied from 2.0 to 5.3% and accuracy from 103 to 105% of the nominal value. The interday precision (RSD%) for 100 sets of plasma QC samples in 29 analytical runs varied from 6.3 to 9.0% and the accuracy from 98.8 to 104% of the nominal value. No significant difference was observed between the interday and intraday precision and accuracy of the QC samples.

Chromatography, High Pressure Liquid↗
PubMed2006

Pharmacokinetic and pharmacodynamic properties of multiple oral doses of sitagliptin, a dipeptidyl peptidase-IV inhibitor: a double-blind, randomized, placebo-controlled study in healthy male volunteers.

BACKGROUND: Dipeptidyl peptidase-IV (DPP-IV) inhibitors represent a new class of oral antihyperglycemic agents. Sitagliptin is an orally active and selective DPP-IV inhibitor currently in Phase III development for the treatment of type 2 diabetes mellitus. OBJECTIVE: The aim of this study was to assess the pharmacokinetic and pharmacodynamic (PK/PD) properties and tolerability of multiple oral once-daily or twice-daily doses of sitagliptin. METHODS: This double-blind, randomized, placebo-controlled,incremental oral-dose study was conducted at SGS Biopharma, Antwerp, Belgium. Healthy, nonsmoking male volunteers aged 18 to 45 years with a creatinine clearance rate of >80 mL/min and normoglycemia and weighing within 15% of their ideal height/weight range were randomly assigned to 1 of 8 treatment groups: sitagliptin 25, 50, 100, 200, or 400 mg or placebo, QD for 10 days; a single dose of sitagliptin 800 mg administered on day 1 followed by 600 mg QD on days 3 to 10; or sitagliptin 300 mg BID for 10 days. For analysis of PK properties, plasma and urine samples were obtained before study drug administration on day 1 and at 0.5, 1, 2, 4, 6, 8, 10, 12, and 16 hours after study drug administration on day 1; before study drug administration on days 2 to 9; and every 24 hours for 96 hours after the last dose on day 10, and analyzed for sitagliptin concentrations. Assays were used to measure inhibition of plasma DPP-IV activity and plasma concentrations of active and total glucagon-like peptide-1 (GLP-1), glucose, and glucagon, and serum concentrations of insulin, C-peptide, insulin-like growth factor-1, and insulin like growth factor binding protein-3. Tolerability was assessed throughout the study using physical examination, including vital sign measurements; 12-lead electrocardiography; and laboratory analysis, including hematology, biochemistry (hepatic aminotransferase and creatine phosphokinase), and urinalysis. RESULTS: Seventy subjects were enrolled (mean age, 32.9 years [range, 18-45 years]; mean weight, 79.7 kg [range, 63.4-97.7 kg]; 8 patients per sitagliptin study group and 14 patients in the control group). In the sitagliptin groups, the plasma concentration-time profiles and principal PK parameters (T(max), C(max), and t((1/2))) were statistically similar at days 1 (single dose) and 10 (steady state). In the groups receiving sitagliptin QD doses, accumulation of sitagliptin was modest (AUC accumulation ratio [day 10/day 1] range, 1.05-1.29), and the apparent terminal elimination t((1/2)) was 11.8 to 14.4 hours. At steady state in the sitagliptin QD groups, the mean proportion of drug excreted unchanged in the urine was approximately 70.6%. Dose-dependent inhibition of plasma DPP-IV activity was apparent, and the pattern of inhibition at steady state (day 10) was statistically similar to that observed on day 1. Day-10 weighted mean inhibition of plasma DPP-IV activity over 24 hours was > or = 80% for doses of > or = 50 mg QD. After a standard meal, active GLP-1 concentrations were significantly increased in the sitagliptin groups by approximately 2-fold compared with that in the control group, a finding consistent with near-maximal acute glucose lowering in preclinical studies. Across doses, no apparent adverse effects, including hypoglycemia, were found or reported. CONCLUSIONS: The results from this study in a select population of healthy male volunteers suggest that multiple oral doses of sitagliptin inhibited plasma DPP-IV activity and affected active GLP-1 concentrations in a dose-dependent manner, without producing hypoglycemia. Multiple dosing of sitagliptin exhibited a PK/PD profile consistent with that of a QD regimen and was well tolerated.

Administration, Oral↗
PubMed2006

Pharmacokinetics and pharmacodynamic effects of the oral DPP-4 inhibitor sitagliptin in middle-aged obese subjects.

Sitagliptin (MK-0431) is an oral, potent, and selective dipeptidyl peptidase-IV (DPP-4) inhibitor developed for the treatment of type 2 diabetes. This multicenter, randomized, double-blind, placebo-controlled study examined the pharmacokinetic and pharmacodynamic effects of sitagliptin in obese subjects. Middle-aged (45-63 years), nondiabetic, obese (body mass index: 30-40 kg/m2) men and women were randomized to sitagliptin 200 mg bid (n = 24) or placebo (n = 8) for 28 days. Steady-state plasma concentrations of sitagliptin were achieved within 2 days of starting treatment, and >90% of the dose was excreted unchanged in urine. Sitagliptin treatment led to approximately 90% inhibition of plasma DPP-4 activity, increased active glucagon-like peptide-1 (GLP-1) levels by 2.7-fold (P < .001), and decreased post-oral glucose tolerance test glucose excursion by 35% (P < .050) compared to placebo. In nondiabetic obese subjects, treatment with sitagliptin 200 mg bid was generally well tolerated without associated hypoglycemia and led to maximal inhibition of plasma DPP-4 activity, increased active GLP-1, and reduced glycemic excursion.

Adenosine Deaminase↗
PubMed2006

[Expression of human granulysin in murine macrophages and its effects on cell damage and apoptosis].

AIM: To construct an eukaryotic expression vector pBudCE4.1/GLS encoding human granulysin(GLS) derived from activated CTLs and observe its effects on cell damage and apoptosis when expressed in macrophages of various murine germ lines. METHODS: The gene sequence coding GLS carried on pEGFP-C1/GLS was subcloned into pBudCE4.1 plasmid and then transfected into different murine macrophages. The expression of GLS was detected by RT-PCR and immunocytochemical staining. The impairment and apoptosis of the host cells was assesed by measuring LDH in culture media and fluorescent staining of nuclear DNA, respectively. RESULTS: The pBudCE4.1/GLS recombinant containing accurate ORF of GLS was obtained and successfully expressed in the target cells at 48 h after transfection. No obvious effects on cell damage and apoptosis was detected during this course. CONCLUSION: The gene coding human GLS can be expressed in murine macrophages. The expressed product has no obvious effects on cell damage and apoptosis.

Animals↗
PubMed2006

[Study on determining the composition of ternary complex (AI-CAS-CTMAB) with the ratio of double peak values at double wavelengths by UV spectrophotometry].

In the present paper, the composition of complex Al-CAS-CTMAB was studied with the ratio of double peak values at dual-wavelengths by UV spectrophotometry. First, the composition of complex of Al-CAS was determined in this method, Al:CAS = 1:2. The absorption of ternary complex(Al-CAS-CTMAB) was determined with reference reagent of the complex Al-CAS. The epsilon values of Al-CAS and Al-CAS-CTMAB were determined with water reference. The composition of complex Al (CAS)n1 (CTMAB)n2 was calculated with the formula: [formula: see text] CAS:CTMAB = 1:1; Al:CAS:CTMAB = 1:2:2. The calculated value and that obtained by traditional method are agreeable. The method is reliable for determining composition of ternary complexes.

English Abstract↗
PubMed2006

[Investigation of biodegradable materials as polymeric gene carriers].

This is a study on the biodegradable polymers as gene controlled-released coatings for gene transfer. The PELA (poly (Dl-lactic acid)-co-poly (ethylene glycol), and PLGAE (poly (lactic acid)-co-poly (ethylene glycol)-co-poly (glycolic acid) random copolymer) were synthesized and prepared as the coatings of plasmid pCH110 in the transfection. All kinds of factors affecting the loading efficiency, cytotoxicity, transfection efficiency and the course of the degradation and release in vitro were discussed. The average diameters of microspheres of PELA and PLGAE were 1-3 microm and 0.72 microm respectively. The loading efficiency levels of them were 62% and 70% respectively. The transfection efficiency levels of two kinds of pCH110 delivery system for COS-1 cells were higher and two of them had few cytotoxicity. After transfection, the X-gal assay was performed and reported positive for 96 h. The biodegradable polymeric materials as gene carriers possess their potential superiority.

Biocompatible Materials↗
PubMed2006

[The cloning and expression of rhlR, a quorum sensing gene in Pseudomonas aeruginosa PAO1, and a study of its effect on bacterial clearance rate in mouse lung].

OBJECTIVE: To inquire about the molecular characteristics of rhlR, a Quorum Sensing gene in Pseudomonas aeruginosa (P. aeruginosa) PAO1, and to explore the immunogenicity of RhlR protein in mouse. METHODS: The rhlR gene of PAO1 was amplified by PCR and cloned into pGEX4T-1 plasmid. The recombination was expressed in E. coli BL21 (DE3) and analyzed by SDS-PAGE and Western blotting. The fusion protein (GST-RhlR) was purified by GST purification Kit and the purified protein was used to immunize mice. P. aeruginosa PA0315 was injected into mouse lung to explore the immuno-protection of the protein. RESULTS: The 726 bp DNA fragment of rhlR was amplified from PAO1 general DNA. The restriction enzyme map showed that the inserted part of rhlR-pGEX4T-1 was successfully constructed and the gene was 100% homologous to rhlR in GenBank. The recombinant plasmid expressed a 54 kDa fusion protein (rhlR-GST) in E. coli BL21 (DE3) after induction by IPTG. The fusion protein could be recognized by mouse polyvalent antiserum against P. aeruginosa. The results showed that the bacterial clearance rate in mouse lung was 86. 92% in rhlR groups and 49.44% in the control group. CONCLUSION: A 54 kDa protein (RhlR-GST) has been successfully expressed in E. coli BL21 (DE3). The RhlR could increase the bacterial clearance rate in mouse lung and may serve as immunoprotective antigen to develop the genetic engineering vaccine.

Animals↗