Biomedical subjects

Xiao Yang

Publications and source records attributed to Xiao Yang.

At least 55 records · Page 3Linked to original sources

PubMed2004

Regulation of large-scale chromatin unfolding by Smad4.

The tumor suppressor Smad4 plays a critical role in the transforming growth factor-beta (TGF-beta signaling pathway. Smad4 is an essential component of transcriptional complexes mediating the activation of Smad-dependent target genes. However, how Smad4 regulates various chromosomal events remains unknown. Here, we report that Smad4 induces large-scale chromatin unfolding by targeting Smad4 to an amplified, lac operator-containing chromosome region in mammalian cells. This unfolding activity maps to the subdomains within the transactivation domain of Smad4. Both the N-terminal missense and the C-terminal truncated tumor-derived mutations increased the chromatin unfolding activity of Smad4. These data identify a novel function of Smad4 and suggest that regulation of large-scale chromatin unfolding by Smad4 may be responsible for cancer development.

Alleles↗

PubMed2004

HBsAg and HBx knocked into the p21 locus causes hepatocellular carcinoma in mice.

Hepatocellular carcinoma (HCC) affects males in a significantly higher proportion than females and is one of the human cancers etiologically related to viral factors. Many studies provide strong evidence of the direct role that hepatitis B virus (HBV) plays in hepatic carcinogenesis, but the functions of HBV surface antigen (HBsAg) and X protein (HBx) in hepatocarcinogenesis through direct or indirect mechanisms are still being debated. We generated two HBV gene knock-in transgenic mouse lines by homologous recombination. HBsAg and HBx genes were integrated into the mouse p21 locus. Both male and female p21-HBx transgenic mice developed HCC after the age of 18 months; however, male p21-HBsAg transgenic mice began to develop HCC 3 months earlier. The expression of a number of genes related to metabolism and genomic instability largely resembled the molecular changes during the development of HCC in humans. ER-beta (estrogen receptor-beta) was extremely up-regulated only in tumor tissues of male p21-HBsAg mice, providing genetic evidence that HBsAg might be the major risk factor affecting the gender difference in the causes of HCC. In conclusion, these mice might serve as good models for studying the different roles of HBsAg and HBx in early events of HBV-related hepatocarcinogenesis.

Animals↗

PubMed2004

[A transgenic mouse that targets the expression of Cre recombinase in hepatocyte].

OBJECTIVE: To construct a mouse that specifically expresses Cre recombinase in hepatocyte. METHODS: A hepatocyte specific transgenic construct containing mouse albumin promoter, the Cre recombinase gene and the poly (A) of human growth factor gene was generated. The linearized constructs were introduced into the fertilized eggs by microinjection to obtain the transgenic mice. The transcriptional specificity of Cre recombinase was detected by reverse transcription polymerase chain reaction (RT-PCR). The expression and function of Cre recombinase were detected by PCR and Southern Blot after crossing the Alb-Cre transgenic mice with the Smad4 conditional knockout mice. RESULTS: The linearized constructs were microinjected into 837 fertilized eggs, and then the 797 effective eggs of microinjected eggs were implanted into the oviducts of 27 pseudo pregnant mice. In the 53 offspring, there were 6 mice carrying the transgene identified by polymerase chain reaction (PCR) and Southern Blot. Cre recombinase transcripts were detected in the livers and testis of the Alb-Cre transgenic mice using RT-PCR. The Cre recombinase was expressed in the livers of the double heterozygous for Alb-Cre and Smad4 floxed allele, and the exon 8 floxed by loxP site was deleted. CONCLUSION: A hepatocyte-specific Cre transgenic mouse was generated successfully. The Cre recombinase expressed specifically in liver and could mediate the recombination between loxP sites in vivo.

Animals↗

PubMed2004

Role of connective growth factor in plasminogen activator inhibitor-1 and fibronectin expression induced by transforming growth factor beta1 in renal tubular cells.

BACKGROUND: Connective tissue growth factor (CTGF) contributes greatly to renal tubulointerstitial fibrosis, which is the final event leading to end-stage renal failure. This study was designed to investigate the effects of CTGF antisense oligodeoxynucleotides (ODNs) on the expressions of plasminogen activator inhibitor-1 (PAI-1) and fibronectin in renal tubular cells induced by transforming growth factor beta1 (TGF-beta1) in addition to the role of CTGF in the accumulation and degradation of renal extracellular matrix (ECM). METHODS: A human proximal tubular epithelial cell line (HKC) was cultured in vitro. Cationic lipid-mediated CTGF antisense ODNs were transfected into HKC cells. After HKC cells were stimulated with TGF-beta1 (5 microg/L), the mRNA levels of PAI-1 and fibronectin were measured by RT-PCR. Intracellular PAI-1 protein synthesis was assessed by flow cytometry. The secreted PAI-1 and fibronectin in the medium were determined by Western blot and ELISA, respectively. RESULTS: TGF-beta1 was found to induce tubular CTGF, PAI-1, and fibronectin mRNA expression. PAI-1 and fibronectin mRNA expression induced by TGF-beta1 was significantly inhibited by CTGF antisense ODNs. CTGF antisense ODNs also inhibited intracellular PAI-1 protein synthesis and lowered the levels of PAI-1 and fibronectin protein secreted into the medium. CONCLUSIONS: CTGF may play a crucial role in the accumulation and degradation of excessive ECM during tubulointerstitial fibrosis, and transfecting CTGF antisense ODNs may be an effective way to prevent renal fibrosis.

Cells, Cultured↗

PubMed2004

[The expression of migration inhibitory factor in peripheral blood mononuclear cell and its correlation with disease activity in systemic lupus erythematosus].

OBJECTIVE: To investigate the change of macrophage migration inhibitory factor (MIF) mRNA expression in peripheral blood mononuclear cell (PBMC) and serum MIF and its correlation with disease active index in patients with systemic lupus erythematosus (SLE). METHODS: MIF mRNA expression of PBMC and serum MIF in patients with active and inactive SLE was measured by reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. The correlations of serum MIF with SLE disease active index, serum ds-DNA antibody, ANA, C3, C4 and IgG were examined. RESULTS: There was marked increase in MIF mRNA expression of PBMC and serum MIF in patients with SLE than that in normal volunteers (P < 0.01). Level of MIF in patients with active SLE was higher than that with inactive SLE (P < 0.01). There were significant correlation between the blood MIF concentration, MIF mRNA expression and SLE disease active index, ds-DNA antibody, C3, C4 and IgG, but no significant different was observed between the blood MIF and antinuclear antibody. CONCLUSIONS: MIF mRNA expression in PBMC and serum MIF concentration is significantly increased in patients with SLE and correlates with SLE disease activity. Serum MIF levels may be a useful parameter for monitoring disease active in patients with SLE.

Adolescent↗

PubMed2004

[Study on a human tissue-type plasminogen activator mutant cDNA knocked-in the beta-casein gene site of murine ES cells].

A murine beta-casein gene targeting vector was constructed using the cloned genomic sequence. The short arm was 2.7 kb including mouse beta-casein gene 5' flanking sequence, exon1, intron1 and partial exon2. The long arm is a 3.4 kb fragment including partial intron2, exon3 approximately 7, intron3 approximately 6 and partial intron7. The human t-PA mutant cDNA was subcloned in the exon2 and fused with the mice beta-casein signal peptide sequence. The positive selective marker neo was placed in the middle of intron2. A tk negative selective marker was just outside the short arm. TC-1 ES cells were cultured and amplified on G418 resistant feeder layer. The linearized targeting construct DNAs of 45 microg were introduced into 2 x 10(7) ES cells by electroporation. Totally 192 ES clones were picked up after cultured in G418 and Gancyclovir for 7 days. The colonies were amplified and subjected to genomic DNA preparation. The genomic DNAs were digested with EcoR I and used for Southern blot analysis. A probe inside the 5' homologous arm was used for hybridization. A 9.8 kb band was found in wild type, but the band was shift down from 9.8 kb to 6.6 kb in the beta-casein gene targeted allele because a new EcoR I site was introduced into the exon2 along with the human t-PA mutant gene. There were 9.8 kb and 6.6 kb bands in targeted ES cells. One clone of targeted ES cells with correct homologous recombination events was obtained among 78 analyzed clones. It lays foundation for gene targeted mice making.

Animals↗

PubMed2004

Clinical features and surgical treatment of A-pattern exotropia.

PURPOSE: To investigate the clinical characteristics and determine the effective surgical managements of A-pattern exotropia. METHODS: Thirty-two patients with A-pattern exotropia underwent superior oblique muscle weakening procedures, medial rectus resection or (and) lateral rectus recession. Pre- and post-operative eye position, deviation angle, superior oblique function and binocular vision were examined and analyzed in the cases. RESULTS: Overaction of the superior oblique muscles (31/32) and underaction of the medial rectus muscle (20/32) were presented in the cases. Postoperatively, a satisfactory ocular alignment was obtained in 28 cases (87.5%), and the A-pattern was corrected in 31 cases (96.9%). Four cases got binocular vision after surgery. CONCLUSIONS: As one of the most common forms of A and V patterns, A-pattern exotropia showed clinical characteristics of superior oblique muscle overaction and medial rectus muscle underacion, which should be the primary factors in the etiology of A-pattern exotropia, and superior oblique weakening procedures combined with horizontal surgery should be an effective approach to A-pattern exotropia treatment.

Adolescent↗

PubMed2004

Establishment of liver specific glucokinase gene knockout mice: a new animal model for screening anti-diabetic drugs.

AIM: To characterize the liver-specific role of glucokinase in maintaining glucose homeostasis and to create an animal model for diabetes. METHODS: We performed hepatocyte-specific gene knockout of glucokinase in mice using Cre-loxP gene targeting strategy. First, two directly repeated loxP sequences were inserted to flank the exon 9 and exon 10 of glucokinase in genomic DNA. To achieve this, linearized targeting vector was electroporated into ES cells. Then G418- and Gancyclovir-double-resistant clones were picked and screened by PCR analysis and the positives identified by PCR were confirmed by Southern blot. A targeted clone was selected for microinjection into C57BL/6J blastocysts and implanted into pseudopregnant FVB recipient. Chimeric mice and their offspring were analyzed by Southern blot. Then by intercrossing the Alb-Cre transgenic mice with mice containing a conditional gk allele, we obtained mice with liver-specific glucokinase gene knockout. RESULTS: Among 161 double resistant clones 4 were positive to PCR and Southern blot and only one was used for further experiments. Eventually we generated the liver specific glucokinase knockout mice. These mice showed increased glucose level with age and at the age of 6 weeks fasting blood glucose level was significantly higher than control and they also displayed impaired glucose tolerance. CONCLUSION: Our studies indicate that hepatic glucokinase plays an important role in glucose homeostasis and its deficiencies contribute to the development of diabetes. The liver glucokinase knockout mouse is an ideal animal model for MODY2, and it also can be applied for screening anti-diabetic drugs.

Animals↗

PubMed2004

[The transgenic mice specifically express Cre recombinase in central nerve system].

For specific expressing Cre recombinase in central nerve system (CNS), a transgenic construct (pGFAP-Cre-hGH), containing the beta-globin insulators, 1.8 kb of glial fibrillary acidic protein gene (GFAP) 5' end regulation region, Cre gene and polyA of human growth hormone gene (hGH) was generated, in which the 5' end regulation region of GFAP was isolated from a 129sv mouse genomic DNA library with PCR-screening. 7.6 kb of pGFAP-Cre-hGH DNA fragment was introduced into 191 fertilized eggs by microinjection. 176 injected eggs were implanted into the oviducts of eight female mice respectively, from which 25 offspring were obtained. Seven mice carry the Cre genes by the identification of PCR and Southern blotting, and the integration efficiency is 28%. The GFAP-Cre transgenic mice were crossed with ROSA26 mice whose genomic DNA is integrated by LoxP sites and LacZ expression frame to check the activity and the tissue-specific expression of Cre recombinase and recombination with its mediation between two LoxP sites. The results of LacZ dying showed that the Cre recombinase was expressed only in CNS and successfully mediated the recombination between the LoxP sites in vivo.

Animals↗

PubMed2004

[Efficacy of rigid gas permeable contact lens for the correction of mixed astigmatism].

PURPOSE: To evaluate the efficacy of rigid gas permeable contact lens (RGPCL) for the correction of mixed astigmatism. METHODS: Patients with mixed astigmatism by mydriasis refraction were selected as subjects. Routine eye check was performed to exclude contraindication of contact lens. Investigation included visual acuity, keratometry and corneal topography. Average keratometry value was chose as base curve of the first trial lens. Final base curve, diameter, material and edge of lens were prescribed according to the centralized position, movement and fluorescence pattern of trial lens. After over refraction, final refractive power of lens was prescribed. Corrective visual acuity with spectacle and rigid gas permeable contact lens were compared in 24 patients (31 eyes). RESULTS: Corrective visual acuity of patients with RGPCL was much better than those with spectacle. CONCLUSIONS: RGPCL is safe and effective to correct mixed astigmatism.

Adolescent↗

PubMed2004

Graphical approach to weak motif recognition.

We address the weak motif recognition problem in DNA sequences, which extends the general motif recognition to more difficult cases, allowing more degenerations in motif instances. Several algorithms have earlier attempted to find weak motifs in DNA sequences but with limitations. In this paper, we propose a graph-based algorithm for weak motif detection, which uses dynamic programming approach to find cliques indicating motif instances. The experiments on synthetic datasets show that the algorithm finds weak motif instances more accurately and efficiently compared to earlier approaches. Its performances on real datasets in finding transcription factor binding sites are comparable with the existing techniques.

Algorithms↗

PubMed2003

Kinetic substrate quantification by fitting the enzyme reaction curve to the integrated Michaelis-Menten equation.

The reliability of kinetic substrate quantification by nonlinear fitting of the enzyme reaction curve to the integrated Michaelis-Menten equation was investigated by both simulation and preliminary experimentation. For simulation, product absorptivity epsilon was 3.00 mmol(-1) L cm(-1) and K(m) was 0.10 mmol L(-1), and uniform absorbance error sigma was randomly inserted into the error-free reaction curve of product absorbance A(i) versus reaction time t(i) calculated according to the integrated Michaelis-Menten equation. The experimental reaction curve of arylesterase acting on phenyl acetate was monitored by phenol absorbance at 270 nm. Maximal product absorbance A(m) was predicted by nonlinear fitting of the reaction curve to Eq. (1) with K(m) as constant. There were unique A(m) for best fitting of both the simulated and experimental reaction curves. Neither the error in reaction origin nor the variation of enzyme activity changed the background-corrected value of A(m). But the range of data under analysis, the background absorbance, and absorbance error sigma had an effect. By simulation, A(m) from 0.150 to 3.600 was predicted with reliability and linear response to substrate concentration when there was 80% consumption of substrate at sigma of 0.001. Restriction of absorbance to 0.700 enabled A(m) up to 1.800 to be predicted at sigma of 0.001. Detection limit reached A(m) of 0.090 at sigma of 0.001. By experimentation, the reproducibility was 4.6% at substrate concentration twice the K(m), and A(m) linearly responded to phenyl acetate with consistent absorptivity for phenol, and upper limit about twice the maximum of experimental absorbance. These results supported the reliability of this new kinetic method for enzymatic analysis with enhanced upper limit and precision.

Algorithms↗

PubMed2003

Mutation analysis of the Smad3 gene in human osteoarthritis.

Osteoarthritis (OA) is the most common joint disease worldwide. Recent studies have shown that targeted disruption of Smad3 in mouse results in OA. To reveal the possible association between the Smad3 gene mutation and human OA, we employed polymerase chain reaction-single strand conformation polymorphism and sequencing to screen mutations in all nine exons of the Smad3 gene in 32 patients with knee OA and 50 patients with only bone fracture. A missense mutation of the Smad3 gene was found in one patient. The single base mutation located in the linker region of the SMAD3 protein was A --> T change in the position 2 of codon 197 and resulted in an asparagine to isoleucine amino-acid substitution. The expressions of matrix metalloproteinase 2 (MMP-2) and MMP-9 in sera of the patient carrying the mutation were higher than other OA patients and controls. This is the first report showing that the Smad3 gene mutations could be associated with the pathogenesis of human OA.

DNA Mutational Analysis↗

PubMed2003

Squamous cell carcinoma and mammary abscess formation through squamous metaplasia in Smad4/Dpc4 conditional knockout mice.

Smad4 is a central mediator for TGFbeta signals, which play important functions in many biological processes. To study the role of Smad4 in mammary gland development and neoplasia, we disrupted this gene in mammary epithelium using a Cre-loxP approach. Smad4 is expressed in the mammary gland throughout development; however, its inactivation did not cause abnormal development of the gland during the first three pregnancies. Instead, lack of Smad4 gradually induced cell proliferation, alveolar hyperplasia and transdifferentiation of mammary epithelial cells into squamous epithelial cells. Consequently, all mutant mice developed squamous cell carcinoma and/or mammary abscesses between 5 and 16 months of age. We demonstrated that absence of Smad4 resulted in beta-catenin accumulation at onset and throughout the process of transdifferentiation, implicating beta-catenin, a key component of the Wnt signaling pathway, in the development of squamous metaplasia in Smad4-null mammary glands. We further demonstrated that TGFbeta1 treatment degraded beta-catenin and induced epithelial-mesenchymal transformation in cultured mammary epithelial cells. However, such actions were blocked in the absence of Smad4. These findings indicate that TGFbeta/Smad4 signals play a role in cell fate maintenance during mammary gland development and neoplasia.

Abscess↗

PubMed2003

IL-17 induces autoantibody overproduction and peripheral blood mononuclear cell overexpression of IL-6 in lupus nephritis patients.

OBJECTIVE: To investigate the role of IL-17 in the overproduction of autoantibodies and IL-6 overexpression by peripheral blood mononuclear cells (PBMC) of lupus nephritis (LN) patients. METHODS: Fifteen consecutively hospitalized LN patients were selected as subjects and 15 healthy adults as normal controls. PBMC were obtained by Ficoll density gradient centrifugation. IgG, anti-dsDNA antibody and IL-6 protein levels were assessed using enzyme-linked immunosorbent assays (ELISA) on the supernatant of cultured PBMC of LN patients or normal controls. IL-6 mRNA levels in PBMC were measured using reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: In medium culture, IgG, anti-dsDNA and IL-6 protein levels of the supernatant of PBMC from LN patients were significantly higher than those from normal controls (1492.1 +/- 73.2 ng/ml vs 636.7 +/- 51.9 ng/ml for IgG, 306.6 +/- 53.7 IU/ml vs 95.8 +/- 11.6 IU/ml for anti-dsDNA and 50.92 +/- 15.92 ng/ml vs 1.77 +/- 0.73 ng/ml for IL-6, all P < 0.001). In LN patients, IgG, anti-dsDNA and IL-6 protein levels were higher in the supernatants of PBMC in the IL-17-stimulated culture than the medium culture, but in normal controls, only the IL-6 protein levels were significantly higher. The increase in IgG, anti-dsDNA and IL-6 protein levels induced by IL-17 was dose-dependent and could be completely blocked by IL-17 monoclonal antibody mIgG(28) and partially blocked by dexamethasone. Similarly, IL-6 mRNA overexpression of PBMC in LN patients or normal controls induced by IL-17 was both dose- and time-dependent. During medium culture, IL-6 mRNA levels in LN patients were significantly higher than those in normal controls (1.80 +/- 0.11 vs 0.36 +/- 0.07). During stimulation with IL-17, IL-6 mRNA levels in LN patients were higher than those in normal controls (3.21 +/- 0.24 vs 1.30 +/- 0.14, P < 0.05) and also significantly higher when comparing the stimulated culture with the medium culture either in LN patients or normal control. CONCLUSIONS: IL-17 may play an important role in the pathogenesis of LN through the induction of IgG, anti-dsDNA overproduction and IL-6 overexpression of PBMC in LN patients.

Adolescent↗

PubMed2003

[Clinical features and surgical treatment of A-pattern exotropia].

OBJECTIVE: To investigate the clinical features of A-pattern exotropia and to evaluate the efficacy of surgical treatment for the correction of A-pattern exotropia (A-XT). METHODS: Pre- and post-operative eye position, superior oblique muscle function, clinical features and binocular function were examined and analyzed in 32 patients with A-pattern exotropia. RESULTS: Overaction of the superior oblique (SO) (31/32) and insufficiency of medial rectus muscle (20/32) were commonly seen in the group of patients. The tenectomy of the SO and the resection of the medial rectus muscle were effective in most of A-pattern exotropia. 87.5% (28/35) of A-pattern exotropia in our study were successfully corrected. Four cases got binocular function after surgery. CONCLUSIONS: Based on the clinical features and the effects of surgical treatment, we conclude that the superior oblique muscle overaction and medial rectus muscle insufficiency are the primary factors in the etiology of A-pattern exotropia that is the most common form of A-V syndrome. Weakening the overaction of superior oblique muscle combined with the correction of exotropia is an effective way to correct A-pattern exotropia.

Adolescent↗

PubMed2003

[Topographical evaluation on decentration of orthokeratology lenses].

OBJECTIVE: To evaluate the degree and correlative factors of decentration of orthokeratology lenses and its effect on the visual function. METHODS: Two different kinds of orthokeratology lenses were fitted to 270 eyes of 135 patients [initial mean refractive error: (-3.98 +/- 1.51) D]. Humphery Instruments ATLAS 8.0 was used for the computer-assisted analysis of corneal differential topographical maps. The examination of corneal topography was proceeded on the patients before the fitting of orthokeratology lenses and 6-month later. The distance from center of optic zone to apex of the cornea was measured as the value of decentration of orthokeratology lenses. The factors influenced the value of decentration were analyzed, including the initial refraction error, astigmatism, keratometry values, corneal eccentricity, and the diameter of the lens. The complaints of patients were recorded. Questionnaires, involving the symptoms of monocular diplopia and glare, were used to evaluate the effects of decentration of orthokeratology lenses on the visual function. RESULTS: The mean distance of decentration was (0.49 +/- 0.34) mm after one night fitting, the mean distance of decentration after follow-up for 1 month, 3 months and 6 months was (0.57 +/- 0.41) mm, (0.55 +/- 0.48) mm and (0.59 +/- 0.39) mm, respectively. After one month, the distance of decentration was less than 0.5 mm in 51.1% eyes, 0.5 - 1.0 mm in 35.6% eyes and more than 1.0 mm in 13.3% eyes. The direction of decentration in eyes with more than 0.50 mm decentration was mainly in the temporal side (48.5%). Patients with greater initial astigmatism and smaller diameter of lens showed greater distance of decentration (P < 0.05). There was no statistically significant difference in the distance of decentration between two groups with different corneal eccentricities and keratometry values (P > 0.05). The distance of decentration was greater in patients with monocular diplopia and glare. CONCLUSIONS: The degree of decentration of orthokeratology depends on the degree of initial refractive error, astigmatism and the design of orthokeratology lenses. The degree of decentration can influence the visual function.

Adolescent↗

PubMed2003

[Establishment of keratinocyte-specific Cre recombinase transgenic mice].

A keratinocyte-specific Cre transgenic construct (pK5-Cre) containing the keratin 5 promoter, Cre recombinase gene and polyA of human growth hormone gene was generated. The 4.2 kb DNA fragment of K5-Cre-hGH was introduced into 720 fertilized zygotes by microinjection. 695 injected eggs were implanted into the oviduct of 29 female mice respectively, from which 48 off-spring were obtained. Twelve mice carrying the transgene were identified by genotyping, and the integration efficiency is 25%. The K5-Cre transgenic mice were crossed with Smad4 conditional gene targeting mice to check the tissue-specific expression of the Cre recombinase and the Cre mediated recombination in multiple tissues. The results showed that the Cre recombinase was expressed in skin tissue only and successfully mediated the recombination between the loxP sites in vivo.

Animals↗