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Biomedical subjects

Ming Xiao

Publications and source records attributed to Ming Xiao.

At least 19 recordsLinked to original sources
PubMed2019

The 22q11 low copy repeats are characterized by unprecedented size and structural variability.

Low copy repeats (LCRs) are recognized as a significant source of genomic instability, driving genome variability and evolution. The Chromosome 22 LCRs (LCR22s) mediate nonallelic homologous recombination (NAHR) leading to the 22q11 deletion syndrome (22q11DS). However, LCR22s are among the most complex regions in the genome, and their structure remains unresolved. The difficulty in generating accurate maps of LCR22s has also hindered localization of the deletion end points in 22q11DS patients. Using fiber FISH and Bionano optical mapping, we assembled LCR22 alleles in 187 cell lines. Our analysis uncovered an unprecedented level of variation in LCR22s, including LCR22A alleles ranging in size from 250 to 2000 kb. Further, the incidence of various LCR22 alleles varied within different populations. Additionally, the analysis of LCR22s in 22q11DS patients and their parents enabled further refinement of the rearrangement site within LCR22A and -D, which flank the 22q11 deletion. The NAHR site was localized to a 160-kb paralog shared between the LCR22A and -D in seven 22q11DS patients. Thus, we present the most comprehensive map of LCR22 variation to date. This will greatly facilitate the investigation of the role of LCR variation as a driver of 22q11 rearrangements and the phenotypic variability among 22q11DS patients.

22q11 Deletion Syndrome↗
PubMed2006

Rapid DNA mapping by fluorescent single molecule detection.

DNA mapping is an important analytical tool in genomic sequencing, medical diagnostics and pathogen identification. Here we report an optical DNA mapping strategy based on direct imaging of individual DNA molecules and localization of multiple sequence motifs on the molecules. Individual genomic DNA molecules were labeled with fluorescent dyes at specific sequence motifs by the action of nicking endonuclease followed by the incorporation of dye terminators with DNA polymerase. The labeled DNA molecules were then stretched into linear form on a modified glass surface and imaged using total internal reflection fluorescence (TIRF) microscopy. By determining the positions of the fluorescent labels with respect to the DNA backbone, the distribution of the sequence motif recognized by the nicking endonuclease can be established with good accuracy, in a manner similar to reading a barcode. With this approach, we constructed a specific sequence motif map of lambda-DNA. We further demonstrated the capability of this approach to rapidly type a human adenovirus and several strains of human rhinovirus.

Adenoviruses, Human↗
PubMed2006

A simple DNA stretching method for fluorescence imaging of single DNA molecules.

Stretching or aligning DNA molecules onto a surface by means of molecular combing techniques is one of the critical steps in single DNA molecule analysis. However, many of the current studies have focused on lambda-DNA, or other large DNA molecules. There are very few studies on stretching methodologies for DNA molecules generated via PCR (typically smaller than 20 kb). Here we describe a simple method of stretching DNA molecules up to 18 kb in size on a modified glass surface. The very low background fluorescence allows efficient detection of single fluorescent dye labels incorporated into the stretched DNA molecules.

DNA↗
PubMed2006

Mutational analysis of the GDD sequence motif of classical swine fever virus RNA-dependent RNA polymerases.

To define the function of the GDD motif of the RNA-dependent RNA polymerase (RdRp) of classical swine fever virus (CSFV), single amino acid substitutions were introduced into the CSFV NS5B. All substitutions within the GDD motif were detrimental to the polymerase activity, the binding activity and the terminal nucleotidyl transferase activity of the NS5B protein. It was also found that the wild-type NS5B had higher RdRp activity with Mg(+2) than with Mn(+2) whereas some mutants worked better with Mn(+2) than with Mg(+2), suggesting that substitutions within the GDD motif modified the enzyme cation preferences and the GDD sequence of CSFV NS5B might be involved in polymerase-metal interaction. Therefore, the GDD amino acid sequence is important for the function of CSFV RdRp.

Amino Acid Motifs↗
PubMed2006

Expression and colocalization of NADPH-diaphorase and Fos in the subnuclei of the parabrachial nucleus in rats following visceral noxious stimulation.

To investigate whether neural nitric oxide synthase (nNOS) in the parabrachial nucleus (PB) is involved in processing visceral noxious stimulation, we mapped the distribution of histochemical staining for nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d), a marker for nNOS, and immunohistochemical staining for Fos, a neuronal activity marker, in the subnuclei of the PB following 2% formalin injection into the stomach of rats. NADPH-d and noxious-stimuli induced Fos staining were also examined in tissue containing PB cells labeled by the retrograde transport of fluogold (FG) injected into the central nucleus of the amygdala (CeA). We found that the number of Fos immunoreactive (Fos-IR) neurons was significantly increased in the dorsal lateral (dl), external lateral (el) and Kölliker-Fuse (KF) subnuclei of the PB. We observed that intensely labeled (type 1) NADPH-d positive neurons were mainly located in the rostral part of the PB; they extended long processes adjacent Fos-IR neurons, but no Fos/type 1 NADPH-d double-labeled neurons were seen. In contrast, lightly labeled (type 2) NADPH-d positive neurons were principally localized in the dl of the PB, in which a few Fos/type 2 NADPH-d double-labeled neurons were detected. Additionally, a large number of FG/Fos double-labeled neurons were observed to be surrounded closely by the intensive NADPH-d staining in the el of the PB. These results suggest that neurons in the el of the PB that project to the CeA are activated by visceral noxious stimulation and could be indirectly influenced by nitric oxide in the PB.

Analysis of Variance↗
PubMed2006

Characterization of the N-terminal domain of classical swine fever virus RNA-dependent RNA polymerase.

To investigate RNA-dependent RNA polymerase (RdRp) further, mutational analysis of the N-terminal domain of the NS5B protein of Classical swine fever virus was performed. Results show that the N-terminal domain (positions 1-300) of the protein might be divided artificially into four different regions, N1-N4. The N1 region (positions 1-61) contained neither conserved lysine nor conserved arginine residues. NS5B protein with deletion of the N1 region has the capacity for elongative RNA synthesis, but not for de novo RNA synthesis on natural templates. All substitutions of the conserved lysines and arginines in the N2 region (positions 63-216) destroyed RdRp activity completely. Substitutions of the conserved arginines in the N3 region (positions 217-280) seriously reduced RdRp activity. However, all substitutions of the conserved lysines in this region enhanced RNA synthesis and made the mutants synthesize RNA on any template. Substitutions of the conserved arginines in the N4 region (positions 281-300) reduced elongative synthesis and destroyed de novo RNA synthesis. In contrast, substitutions of lysines in this region did not affect RdRp activity significantly. These data indicate that the N3 region might be related to the enzymic specificity for templates, and the conserved lysines and arginines in different regions have different effects on RdRp activity. In combination with the published crystal structure of bovine viral diarrhea virus NS5B, these results define the important role of the N-terminal domain of NS5B for template recognition and de novo RNA synthesis.

Amino Acid Sequence↗
PubMed2006

[Automatic decompression with micro-catheter for open and tension pneumothorax].

OBJECTIVE: To evaluate the feasibility of treating pneumothorax with automatic intermittent decompression with micro-catheter instead of traditional thorax water sealed drainage (TWSD). METHODS: The automatic decompression instrument (ADI), which decompressed intermittently with programmed control, was designed and assembled by the authors (Patent No. ZL 01242081.6). A prospective study of the efficacy of this device was conducted in 87 pneumatothorax cases, and the results were compared with those of TWSD. RESULTS: The average time of closure in ADI group was 4.12+/-0.98 days, which was significantly shorter than that with TWSD (6.83+/-2.06 days, P<0.01). The incidence of complications was also significantly lower in ADI patients (P<0.01), and none of them developed severe complications. Clinical cure was achieved in all the patients in ADI group except for two patients who gave up treatment voluntarily and one transferred for open surgery. CONCLUSIONS: Application of ADI allows faster healing and safer and easier operation, and causes fewer complications and less pain with shortened hospital stay as compared with conventional therapy for open and tension pneumothorax.

Adolescent↗
PubMed2005

In vivo expression of a conditional TGF-beta1 transgene: no evidence for TGF-beta1 transgene expression in SM22alpha-tTA transgenic mice.

Transforming growth beta-1 (TGF-beta1) appears to play a critical role in the regulation of arterial intimal growth and the development of atherosclerosis. TGF-beta1 is expressed at increased levels in diseased arteries; however, its role in disease development remains controversial. Experiments in which TGF-beta1 is overexpressed in the artery wall of transgenic mice could clarify the role of TGF-beta1 in the development or prevention of vascular disease. However, constitutive overexpression of a TGF-beta1 transgene in the mouse artery wall is embryonically lethal. Therefore, to overexpress TGF-beta1 in the artery wall of adult mice, we generated mice that were transgenic for a conditional, tetracycline operator (tetO)-driven TGF-beta1 allele. These mice were viable, and when crossed with mice expressing a tetracycline-regulated transactivator (tTA) in the heart, expressed the TGF-beta1 transgene in a cardiac-restricted and doxycycline-dependent manner. Nevertheless, breeding of the tetO-TGF-beta1 transgene into three lines of mice transgenic for a smooth muscle-targeted tTA (SM22alpha-tTA mice; reported elsewhere to transactivate tetO-driven alleles in smooth muscle cells of large arteries) did not yield expression of the TGF-beta1 transgene. Moreover, tTA expression was not detected in aortae of the SM22alpha-tTA mice. Transgenic mice that express tTA at high levels in vascular smooth muscle and reliably transactivate tetO-driven transgenes would be useful for deciphering the role of TGF-beta1 (or other proteins) in normal arterial physiology and in the development of arterial disease. Currently available SM22alpha-tTA mice were not useful for this purpose. Generation of higher-expressing lines of SM22alpha-tTA mice appears warranted.

Alleles↗
PubMed2005

Polymorphism discovery in 51 chemotherapy pathway genes.

Candidate gene pharmacogenetic studies offer a strategy for the rapid assessment of putative predictive markers. As a first step toward studying the pharmacogenetics of cancer chemotherapy, 51 candidate genes from the pathways of antineoplastic agents were resequenced to identify common genetic polymorphisms that might alter therapeutic response or toxicity. Forty DNA samples were screened from each of three population groups: African-Americans, Asian-Americans and European-Americans. Nearly 378 kb of genomic sequence was obtained from each sample. Nine hundred and four variants were identified, including 139 coding single nucleotide polymorphisms (cSNPs). Three hundred and fifty-six (40%) polymorphisms were common to all three populations and 366 (41%) were population specific. Three hundred and forty-six (38%) variants were novel polymorphisms that were not present in the three public databases that were examined. One hundred and eleven (35%) of the 319 non-synonymous cSNPs that were identified by either resequencing or database mining were predicted by PolyPhen to be either possibly or probably damaging. For the non-synonymous cSNPs identified by resequencing, both the number of cSNPs found and the maximum estimated allele frequency decreased with increasing predicted severity. These results provide experimental validation and estimated allele frequencies for polymorphisms in three common ethnic groups and facilitate applied pharmacogenetic studies of anticancer drugs.

Antineoplastic Agents↗
PubMed2005

The distribution of neural nitric oxide synthase-positive cerebrospinal fluid-contacting neurons in the third ventricular wall of male rats and coexistence with vasopressin or oxytocin.

The detailed distribution of neural nitric oxide synthase (nNOS)-positive cerebrospinal fluid-contacting neurons (CSF-CN) was studied in the wall of the third ventricle of rats by anti-nNOS immunohistochemistry. The coexistence of nNOS and 8-arginine vasopressin (AVP) or oxytocin (OT) was also investigated in the CSF-CN using double labeling immunohistochemistry. The results demonstrated a widespread occurrence of nNOS-CSF-CN throughout the wall of the hypothalamic third ventricle. The vast majority of nNOS-CSF-CN cell bodies were of magnocellular type, commonly classified as oval, fusiform, multipolar, and inverted pear shape. These cell bodies were located in the ependyma, the subependyma, or the parenchyma, and their processes inserted in the ependymal layer or directly contacted with the CSF space. Electron microscopy demonstrated many nNOS-immunoreactive somas, dendrites, and/or axons that were situated at the subependyma, the ependyma, or the supraependyma. Generally, the distribution of OT-CSF-CN in the third ventricular wall was similar to the nNOS-CSF-CN and the ratio of NOS/OT co-expression was approximately 88%. In comparison, the distribution of AVP-CSF-CN was mainly restricted to the rostral part of the third ventricle and the ratio of nNOS/AVP co-expression was only about 6%. The widespread presence of nNOS-CSF-CN-expressing OT in the third ventricular region suggests that NO is an important messenger in the CSF-hypothalamo-hypophyseal neuroendocrine regulation that may in part act in concert with OT.

Animals↗
PubMed2005

Human adult olfactory neural progenitors rescue axotomized rodent rubrospinal neurons and promote functional recovery.

Previously, our lab reported the isolation of patient-specific neurosphere-forming progenitor lines from human adult olfactory epithelium from cadavers as well as patients undergoing nasal sinus surgery. RT-PCR and ELISA demonstrated that the neurosphere-forming cells (NSFCs) produced BDNF. Since rubrospinal tract (RST) neurons have been shown to respond to exogenous BNDF, it was hypothesized that if the NSFCs remained viable following engraftment into traumatized spinal cord, they would rescue axotomized RS neurons from retrograde cell atrophy and promote functional recovery. One week after a partial cervical hemisection, GFP-labeled NSFCs suspended in Matrigel matrix or Matrigel matrix alone was injected into the lesion site. GFP-labeled cells survived up to 12 weeks in the lesion cavity or migrated within the ipsilateral white matter; the apparent number and mean somal area of fluorogold (FG)-labeled axotomized RST neurons were greater in the NSFC-engrafted rats than in lesion controls. Twelve weeks after engraftment, retrograde tracing with FG revealed that some RST neurons regenerated axons 4-5 segments caudal to the engraftment site; anterograde tracing with biotinylated dextran amine confirmed regeneration of RST axons through the transplants within the white matter for 3-6 segments caudal to the grafts. A few RST axons terminated in gray matter close to motoneurons. Matrix alone did not elicit regeneration. Behavioral analysis revealed that NSFC-engrafted rats displayed better performance during spontaneous vertical exploration and horizontal rope walking than lesion Matrigel only controls 11 weeks post transplantation. These results emphasize the unique potential of human olfactory neuroepithelial-derived progenitors as an autologous source of stem cells for spinal cord repair.

Animals↗
PubMed2005

Kinetic fluorescence-quenching detection assay for allele frequency estimation.

The analysis of human genetic variations, such as single-nucleotide polymorphisms (SNPs), has great applications in genome-wide association studies of complex genetic traits. We have developed an SNP genotyping method based on the primer extension assay with fluorescence quenching detection. The template-directed dye-terminator incorporation with fluorescence quenching detection (FQ-TDI) assay is based on the observation that the intensity of fluorescent dye R110- and R6G-labeled acycloterminators is universally quenched once they are incorporated onto a deoxyribonucleic acid (DNA) oligonucleotide primer. By comparing the rate of fluorescence quenching of the two allelic dyes in real time, we have extended this method for allele frequency estimation of SNPs in pooled DNA samples. The kinetic FQ-TDI assay is highly accurate and reproducible both in genotyping and in allele frequency estimation. Allele frequencies estimated by the kinetic FQ-TDI assay correlated well with known allele frequencies, with an r2 value of 0.993. Applying this strategy to large-scale studies will greatly reduce the time and cost for genotyping hundreds and thousands of SNP markers between affected and control populations.

Alleles↗
PubMed2005

Physical and hydraulic properties of baked ceramic aggregates used for plant growth medium.

Baked ceramic aggregates (fritted clay, arcillite) have been used for plant research both on the ground and in microgravity. Optimal control of water and air within the root zone in any gravity environment depends on physical and hydraulic properties of the aggregate, which were evaluated for 0.25-1-mm and 1-2-mm particle size distributions. The maximum bulk densities obtained by any packing technique were 0.68 and 0.64 g cm-3 for 0.25-1-mm and 1-2-mm particles, respectively. Wettable porosity obtained by infiltration with water was approximately 65%, substantially lower than total porosity of approximately 74%. Aggregate of both particle sizes exhibited a bimodal pore size distribution consisting of inter-aggregate macropores and intra-aggregate micropores, with the transition from macro- to microporosity beginning at volumetric water content of approximately 36% to 39%. For inter-aggregate water contents that support optimal plant growth there is 45% change in water content that occurs over a relatively small matric suction range of 0-20 cm H2O for 0.25-1-mm and 0 to -10 cm H2O for 1-2-mm aggregate. Hysteresis is substantial between draining and wetting aggregate, which results in as much as a approximately 10% to 20% difference in volumetric water content for a given matric potential. Hydraulic conductivity was approximately an order of magnitude higher for 1-2-mm than for 0.25-1-mm aggregate until significant drainage of the inter-aggregate pore space occurred. The large change in water content for a relatively small change in matric potential suggests that significant differences in water retention may be observed in microgravity as compared to earth.

Aluminum Silicates↗
PubMed2005

[Clinical nature of tetrodotoxin intoxication and the measures for emergency rescue].

OBJECTIVE: To define the clinical nature of tetrodotoxin intoxication and explore the measures for its emergency rescue. METHODS: The emergency rescue measures and outcomes of 33 cases tetrodotoxin intoxication were reviewed. RESULTS: No obvious effects were observed in these patients after administration of atropine or application of respiratory stimulant, but implementation of mechanical ventilation and comprehensive life support measures resulted in successful rescue of mild to moderate cases within 10 to 14 h, and recovery of spontaneous respiration was achieved in critical cases in 19-24 h. CONCLUSION: Tetrodotoxin selectively blocks the neuromuscular electrical activity, producing the effect of nondepolarizing curine and leading to failure of the respiratory drive without compromising the respiratory center. Atropine, a M-cholinergic receptor blocker, or central stimulants may fail to produce obvious effects, but comprehensive treatments including maintenance of effective ventilation can bring satisfactory effects.

Adolescent↗
PubMed2005

[Inhibition of vascular endothelial growth factor gene expression and proliferation of leukemia cells by RNA interference].

OBJECTIVE: To explore the feasibility of selective inhibiting VEGF expression using VEGF short hairpin RNA (shRNA) interference, and observe the effects of VEGF gene silencing on NB4 cells growth. METHODS: Three 19 bp reverse repeated motifs targeting exons 3, 4, 5 respectively of VEGF gene were synthesized and cloned into eukaryotic expression plasmid pGenesil-1 containing U6 shRNA promoter and termination signal of RNA polymerase. The recombinant plasmids pGenesil-VR1, pGenesil-VR2, pGenesil-VR3 and pGenesil-con (plasmid containing random DNA fragment) were transfected into NB4 cells respectively through lipofectamine reagent. The alteration of VEGF expression was examined by fluorescent real time RT-PCR and Western blot. The proliferation capacity of leukemia cells was measured by trypan blue exclusion, MTT assay, colony formation assay and cell cycles analysis. RESULTS: Recombinant plasmids containing three shRNAs and random fragment were successfully constructed and transfected into NB4 cells respectively by liposome-mediated gene transfer method. shRNA in pGenesil-VR3 cells knocked down the expression of VEGF mRNA and protein dramatically in a sequence-specific manner when compared with that of pGenesil-VR1, Genesil-VR2 and pGenesil-con. The NB4 cells transfected with pGenesil-VR3 (NB4-VR3) had a more significant decrease in proliferation ability than NB4 and that transfected with pGenesil-con (NB4-con). The colony forming efficiencies of NB4-VR3, NB4-con and NB4 cell were (13.3 +/- 3.8)%, (21.3 +/- 6.4)% and (24.5 +/- 5.2)%, respectively (P < 0.05). Higher G(1) and lower S proportion were found in cell cycle distribution in comparison with the control groups by FCM. CONCLUSIONS: The shRNA can efficiently suppress VEGF expression in NB4 cells. Selective VEGF gene silence can inhibit the malignant proliferation of leukemia cells.

Apoptosis↗
PubMed2004

Role of excess inorganic pyrophosphate in primer-extension genotyping assays.

We have developed and genotyped >15,000 SNP assays by using a primer extension genotyping assay with fluorescence polarization (FP) detection. Although the 80% success rate of this assay is similar to those of other SNP genotyping assays, we wanted to determine the reasons for the failures and find ways to improve the assay. We observed that the failed assays fell into three general patterns: PCR failure, excess of heterozygous genotypes, and loss of FP signal for one of the dye labels. After analyzing several hundred failed assays, we concluded that 5% of the assays had PCR failure and had to be redesigned. We also discovered that the other two categories of failures were due to misincorporation of one of the dye-terminators during the primer extension reaction as a result of primer shortening with a reverse reaction involving inorganic pyrophosphate, and to the quenching of R110-terminator after its incorporation onto the SNP primer. The relatively slow incorporation of R110 acycloterminators by AcycloPol compounds the problem with the R110 label. In this report, we describe the source of the problems and simple ways to correct these problems by adding pyrophosphatase, using quenching as part of the analysis, and replacing R110 by Texas red as one of the dye labels. With this new protocol, we have achieved approximately 95% success rate in assay development without the need for optimization.

Alkaline Phosphatase↗
PubMed2004

Relationship between encephalopathy and portal vein-vena cava shunt: value of computed tomography during arterial portography.

AIM: To assess the value of computed tomography during arterial portography (CTAP) in portal vein-vena cava shunt, and analysis of the episode risk in encephalopathy. METHODS: Twenty-nine patients with portal-systemic encephalopathy due to portal hypertension were classified by West Haven method into grade I(29 cases), grade II(16 cases), grade III(10 cases), grade IV( 4 cases). All the patients were scanned by spiral-CT. Plane scans, artery phase and portal vein phase enhancement scans were performed, and the source images were thinly reconstructed to 1.25 mm. We reconstructed the celiac trunk, portal vein, inferior vena cava and their branches and subjected them to three-dimensional vessel analysis by volume rendering (VR) technique and multiplanar volume reconstruction (MPVR) technique. The blood vessel reconstruction technique was used to evaluate the scope and extent of portal vein-vena cava shunt, portal vein emboli and the fistula of hepatic artery-portal vein. The relationship between the episode risk of portal-systemic encephalopathy and the scope and extent of portal vein-vena cava shunt, portal vein emboli and fistula of hepatic artery- portal vein was studied. RESULTS: The three-dimensional vessel reconstruction technique of spiral-CT could display celiac trunk, portal vein, inferior vena cava and their branches at any planes and angles and the scope and extent of portal vein-vena cava shunt, portal vein emboli and the fistula of hepatic artery- portal vein. In twenty-nine patients with portal-systemic encephalopathy, grade I accounted for 89.7% esophageal varices, 86.2% paragastric varices; grade II accounted for 68.75% cirsomphalos, 56.25% paraesophageal varices, 62.5% retroperitoneal varices and 81.25% dilated azygos vein; grade III accounted for 80% cirsomphalos, 60% paraesophageal varices, 70% retroperitoneal varices, 90% dilated azygos vein, and part of the patients in grades II and III had portal vein emboli and fistula of hepatic artery-portal vein; grade IV accounted for 75% dilated left renal vein, 50% paragallbladder varices, all the patients had fistula of hepatic artery- portal vein. CONCLUSION: The three-dimensional vessel reconstruction technique of spiral-CT can clearly display celiac trunk, portal vein, inferior vena cava and their branches at any planes and angles and the scope and extent of portal vein-vena cava shunt. The technique is valuable for evaluating the episode risk in portal-systemic encephalopathy.

Adult↗