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Biomedical subjects

Ming Li

Publications and source records attributed to Ming Li.

At least 199 records · Page 11Linked to original sources
PubMed2005

SIV-specific T lymphocyte responses in PBMC and lymphoid tissues of SIV-infected pigtailed macaques during suppressive combination antiretroviral therapy.

There is currently no SIV macaque model in which the effects of combination antiretroviral therapy on tissue immune responses and latent reservoirs have been measured. This study was performed to define the impact of combination therapy on the specificity and distribution of the T lymphocyte response in multiple tissue compartments. Pigtailed macaques (Macaca nemestrina) were infected with SIV/17E-Fr and treated with combination antiretroviral therapy consisting of 9-R-(2-phosphonomethoxypropyl)adenine (PMPA) and beta-2',3'-dideoxy-3'-thia-5-fluorocytidine (FTC). The SIV-specific T lymphocyte response was measured in peripheral blood, spleen and several lymph nodes at necropsy by IFN-gamma Elispot analysis. Two animals (one treated, one untreated) had high acute peak viremia, which was associated with lower SIV-specific T lymphocyte responses in the peripheral blood and lymphoid tissues. In the treated animal, viremia was controlled to low or undetectable for the study duration, and virus-specific responses remained low. The untreated animal remained viremic throughout the study and developed clinical symptoms of AIDS. In contrast, the two animals that had lower acute peak viremia (one treated, one untreated) had more robust T lymphocyte responses, and controlled viral replication. Virus-specific responses were detected in the treated animal despite 6 months of suppressive therapy. These data suggest that in this model, in the context of acute peak viremia and weak T cell responses, combination therapy may be essential to control virus replication and disease progression. Conversely, in the setting of low initial viremia and robust T lymphocyte responses, treatment does not have a detrimental effect on the immune response.

Adenine↗
PubMed2005

T-type (alpha1G) low voltage-activated calcium channel interactions with nitric oxide-cyclic guanosine monophosphate pathway and regulation of calcium homeostasis in human cavernosal cells.

INTRODUCTION: Nitric oxide-cyclic guanosine monophosphate (NO-cGMP)-mediated relaxation of cavernosal smooth muscle during erection is accompanied by a decrease in intracellular calcium concentrations ([Ca2+](i)). However, it is not known whether and how an increase in [Ca2+](i) is responsible for (i) initiating smooth muscle contraction/detumescence following relaxation; and (ii) maintaining the penis in a flaccid state under nonstimulating conditions. AIM: To elucidate (i) the mechanism(s) of [Ca2+](i) homeostasis regulation in human cavernosal smooth muscle cells (HCSMC); and (ii) how NO-cGMP interacts with such [Ca2+](i) homeostasis. METHODS: We evaluated the expression and function of both T-type and L-type Ca2+ channels in HCSMC by employing selective probes/inhibitors using various cellular and molecular techniques (e.g., reverse transcriptase and real-time polymerase chain reaction, cell proliferation assay, fura-2 Ca2+ fluorescence spectroscopy, enzyme-linked immuno-absorbent assay (ELISA)). MAIN OUTCOME MEASURE: We have demonstrated for the first time significant interactions of NO-cGMP with the T-type (alpha1G) Ca2+ channel in HCSMC. RESULTS: Our results suggest that in addition to NO-induced rapid and transient decrease in [Ca2+](i) that results in smooth muscle relaxation, NO-cGMP also enhanced mRNA expression of the T-type (alpha1G) Ca2+ channel resulting in delayed elevation of [Ca2+](i). This could be abolished by a selective T-channel blocker, NNC 55-0396. Another unique finding of this study is that dose-dependent HCSMC proliferation in vitro by NO is associated with the activation of the T-type (alpha1G) Ca2+ channel that regulates [Ca2+](i) homeostasis in these cells. CONCLUSIONS: Human cavernosal cells express T-type (alpha1G) Ca2+ channels that are involved in maintaining [Ca2+](i) homeostasis and regulation of NO-cGMP-induced smooth muscle relaxation-contraction responsible for penile erection, flaccidity, and tonicity. Targeting these Ca2+ channels may (i) associate various comorbidities with the onset of erectile dysfunction; (ii) provide a biochemical basis for differences between therapeutic profiles of various phosphodiesterase type 5 inhibitors, especially in nonresponders to current therapy; and (iii) provide biochemical basis in understanding mechanism(s) of drug tolerance.

Calcium↗
PubMed2005

Biological activities of purified harpin(Xoo) and harpin(Xoo) detection in transgenic plants using its polyclonal antibody.

Many harpins have been found in plant pathogen bacteria that can elicit disease and insect resistance in plants, and promote plant growth. In this work, we overexpressed and purified Xanthomonas oryzae pv. oryzae harpin, harpin(Xoo), in Escherichia coli BL21/pGEX-hpa1. Harpin(Xoo) was fused to the C-terminus of glutathione S-transferase (GST) and purified using the Bulk GST purification module and thrombin cleavage capture kit. Purified harpin(Xoo) protein was sensitive to protease K and stable to heat treatment, and could not induce a hypersensitive response after treatment with various plant metabolic inhibitors; these characteristics were similar to harpin(Ea) of Erwinia amylovora. The purified harpin(Xoo) showed a similar ability to induce tobacco mosaic virus resistance in tobacco as harpin(Ea). Its antibody worked well in detecting the purified harpin(Xoo), harpin(Xoo) in the total protein of E. coli BL21/pGEX-hpa1 and an hpa1 transgenic rice.

Bacterial Outer Membrane Proteins↗
PubMed2005

Outbreak of infection caused by Enterobacter cloacae producing the novel VEB-3 beta-lactamase in China.

Over a 4-month period from November 2002 to February 2003, 27 ceftazidime-resistant or cefotaxime-resistant nonrepetitive Enterobacter cloacae isolates were collected from 27 patients hospitalized at HuaShan Hospital, Shanghai, People's Republic of China. The Etest did not detect extended-spectrum beta-lactamases (ESBLs) in those 27 isolates; however, screening by the NCCLS ESBL disk test and confirmatory tests detected ESBLs in 4 of 27 isolates and PCR detected ESBLs in 23 of 27 isolates. The majority of ESBL producers exhibited the same repetitive extragenic palindromic PCR pattern but harbored different ESBL genes. CTX-M-3 was the most prevalent ESBL in our study. Interestingly, 12 clonally related E. cloacae isolates possessed a novel bla(VEB)-type beta-lactamase, bla(VEB-3). Bla(VEB-3) was encoded by the chromosome and was located in an integron. Nine of the 12 isolates harbored both the bla(VEB-3) and the bla(CTX-M-3)-like ESBLs. This is the first report of a VEB-1-like ESBL in China and the first report of the simultaneous presence of VEB-1 and CTX-M-3-like ESBLs in an isolate.

Amino Acid Sequence↗
PubMed2005

Human immunodeficiency virus type 1 env clones from acute and early subtype B infections for standardized assessments of vaccine-elicited neutralizing antibodies.

Induction of broadly cross-reactive neutralizing antibodies is a high priority for AIDS vaccine development but one that has proven difficult to be achieved. While most immunogens generate antibodies that neutralize a subset of T-cell-line-adapted strains of human immunodeficiency virus type 1 (HIV-1), none so far have generated a potent, broadly cross-reactive response against primary isolates of the virus. Even small increments in immunogen improvement leading to increases in neutralizing antibody titers and cross-neutralizing activity would accelerate vaccine development; however, a lack of uniformity in target strains used by different investigators to assess cross-neutralization has made the comparison of vaccine-induced antibody responses difficult. Thus, there is an urgent need to establish standard panels of HIV-1 reference strains for wide distribution. To facilitate this, full-length gp160 genes were cloned from acute and early subtype B infections and characterized for use as reference reagents to assess neutralizing antibodies against clade B HIV-1. Individual gp160 clones were screened for infectivity as Env-pseudotyped viruses in a luciferase reporter gene assay in JC53-BL (TZM-bl) cells. Functional env clones were sequenced and their neutralization phenotypes characterized by using soluble CD4, monoclonal antibodies, and serum samples from infected individuals and noninfected recipients of a recombinant gp120 vaccine. Env clones from 12 R5 primary HIV-1 isolates were selected that were not unusually sensitive or resistant to neutralization and comprised a wide spectrum of genetic, antigenic, and geographic diversity. These reference reagents will facilitate proficiency testing and other validation efforts aimed at improving assay performance across laboratories and can be used for standardized assessments of vaccine-elicited neutralizing antibodies.

AIDS Vaccines↗
PubMed2005

Bayesian designs with frequentist and Bayesian error rate considerations.

So far, most Phase II trials have been designed and analysed under a frequentist framework. Under this framework, a trial is designed so that the overall Type I and Type II errors of the trial are controlled at some desired levels. Recently, a number of articles have advocated the use of Bayesian designs in practice. Under a Bayesian framework, a trial is designed so that the trial stops when the posterior probability of treatment is within certain prespecified thresholds. In this article, we argue that trials under a Bayesian framework can also be designed to control frequentist error rates. We introduce a Bayesian version of Simon's well-known two-stage design to achieve this goal. We also consider two other errors, which are called Bayesian errors in this article because of their similarities to posterior probabilities. We show that our method can also control these Bayesian-type errors. We compare our method with other recent Bayesian designs in a numerical study and discuss implications of different designs on error rates. An example of a clinical trial for patients with nasopharyngeal carcinoma is used to illustrate differences of the different designs.

Bayes Theorem↗
PubMed2005

[Preparation of monoclonal antibodies against multiple antigens].

OBJECTIVE: To prepare monoclonal antibodies (mAbs) against multiple antigens by single cell fusion. METHODS: BALB/c mice were immunized with the multiple antigens, namely alpha fetoprotein (AFP), carcinoembryonic antigen (CEA), HBsAgiHBcAg and HBeAg, and hybridomas were employed using PEG as the fusing agent. The hybridoma cells were respectively screened with AFP, CEA, HBsAg, HBcAg and HBeAg by enzyme-linked immunosorbent assay and limited dilution. The mAbs were purified by protein G affinity chromatography, its subtype was identified, the affinity constants (K(a)) were determined and the specificity was analyzed by Western blotting. RESULTS: Twenty hybridoma cell lines were obtained by single cell fusion, including 5 cell lines against AFP, 6 against CEA, 3 against HBsAg, 4 against HBcAg, and 2 against HBeAg. The subtypes of some hybridoma cell lines positive for the mAbs were identified as the immunoglobulin G1 (IgG1), with K(a) ranging from 1x10(9) M(-1) to x10(11) M(-1). Western blot analysis showed that all the mAbs strongly and specifically bound to their respective antigens. CONCLUSION: The mAbs against multiple antigens have been obtained by single cell-fusion, which increases the production of mAbs and reduces the time of preparation.

Animals↗
PubMed2005

[Inhibition of phospholipase C gamma1 signaling pathway promotes apoptosis of human colorectal carcinoma cells].

OBJECTIVE: To investigate the effects of inhibiting phospholipase C gamma1 signaling pathway on the apoptosis of human colorectal carcinoma cells. METHODS: SW620 cells were treated with U73122 in vitro to inhibit the phospholipase C gamma1 signalling pathway and examined under light microscope and transmission electron microscope for analyzing changes in apoptotic behavior of the cells. MTT assay was used to evaluate the cell killing effects, and the percentage of apoptotic cells analyzed using flow cytometry. RESULTS: After inhibition of the phospholipase C gamma1 signaling pathway by U73122, SW620 cells exhibited obvious apoptotic morphology, the viable cells decreased dramatically, and the percentage of apoptotic cells rose to above 50%. CONCLUSION: Inhibition of phospholipase C gamma1 signaling pathway can induce apoptosis of human colorectal carcinoma cells.

Apoptosis↗
PubMed2005

Discovering sequence-structure motifs from protein segments and two applications.

We present a novel method for clustering short protein segments having strong sequence-structure correlations, and demonstrate that these clusters contain useful structural information via two applications. When applied to local tertiary structure prediction, we achieve approximately 60% accuracy with a novel dynamic programming algorithm. When applied to secondary structure prediction based on Support Vector Machines, we obtain a approximately 2% gain in Q3 performance by incorporating cluster-derived data into training and classification. These encouraging results illustrate the great potential of using conserved local motifs to tackle protein structure predictions and possibly other important problems in biology.

Algorithms↗
PubMed2005

[Preparation of time-resolved fluoroimmunoassay kit for human alpha-fetoprotein detection].

OBJECTIVE: To prepare time-resolved fluoroimmunoassay (TRFIA) kit for detecting human alpha-fetoprotein (hAFP). METHODS: Sandwich TRFIA kit for hAFP detection was developed using monoclonal antibody of anti-AFP. RESULTS: AFP-TRFIA kit was capable of detecting AFP within the range of 1-1 000 U/ml with sensitivity of 0.17 U/ml and without cross-reactivity with CEA, CA12-5, CA15-3, or CA19-9. The intra- and inter-assay coefficients of variation were (3.3-5.9)% and (3.7-6.5)%, respectively. The prepared AFP-TRFIA reagent could withstand preservation at 4 degrees celsius; for 1 year and at 37 degrees celsius; for 7 days with the cutoff value of 12 U/ml in healthy subjects (n=426). The correlation coefficient of the detection results between this kit and commercially available AFP kit (Wallac OY, Finland) for 60 blood samples was 0.995. CONCLUSION: The prepared TRFIA kit for hAFP detection meets the demand of clinical application with good sensitivity, precision, specificity, stability and accuracy.

Fluoroimmunoassay↗
PubMed2005

[Replication and expression of recombinant hepatitis C virus in a human liver cancer cell line 7721 in vitro].

OBJECTIVE: To investigate whether recombinant hepatitis C virus (HCV) is capable of long-term replication in vivo in susceptible cells. METHODS: A human liver cell line 7721 was infected with recombinant HCV and after 72-hour incubation, the presence of HCV RNA in the cells and the supernatant were detected by reverse transcriptase (RT)-PCR. The expression of HCV antigen was detected by immunohistochemistry and observed by confocal laser scanning microscope. RESULTS: The plus and minus strands of HCV RNA were detected in the infected cells and the plus strand in the supernatant 72 h after infection. Expressions of HCV core antigens and E1 antigens were found within the cytoplasm. CONCLUSIONS: The recombinant HCV can replicate in 7721 cell line in vitro and this HCV infection model can be useful for studying the mechanism of HCV infection and replication.

Cell Line, Tumor↗
PubMed2005

[Detection of SARS-associated coronavirus N protein by time-resolved fluoroimmunoassay].

OBJECTIVE: To develop a method for quantitative detection of severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV) N protein by timed-resolved fluoroimmunoassay (TRFIA). METHODS: Using a monoclonal antibody (mAb) against SARS-CoV N protein, screened by SARS-CoV N protein and matching experiment, a method for quantitative detection of SARS-CoV N protein by TRFIA was established on the basis of double sandwich enzyme-linked immunosorbent assay (ELISA) and evaluated against the ELISA kit. RESULT: The measurement range of the assay was 0.02-150 ng/ml with a sensitivity of 0.02 ng/ml, the coefficient of variability within runs of 3.3%;-6.2%;, and coefficient of variability between days of 5.3%;-9.6%;. The results of detection were consistent between ELISA and TRFIA. CONCLUSION: TRFIA is a new, sensitive and specific immunoassay for detecting SARS N protein with potential value in clinical applications.

Antibodies, Monoclonal↗
PubMed2005

[Preparation of a monoclonal antibodies against Plasmodium falciparum glutamate dehydrogenase and establishment of colloidal gold-immunochromatographic assay].

OBJECTIVE: To prepare a monoclonal antibodies (mAbs) against glutamate dehydrogenase (GDH) of Plasmodium falciparum (FCC1/HN strain) and establish colloidal gold-immunochromatographic assay (GICA) for diagnosis of Plasmodium falciparum malaria. METHODS: Recombinant GDH was used to immunize Balb/C mice and the mAbs against GDH were prepared using hybridoma technique followed by identification of IgG isotype and its affinity. Protein-G affinity chromatography was employed to purify the antibodies, which were labeled with colloidal gold for establishment of GICA for Plasmodium falciparum detection. RESULTS: Six mAbs were obtained and identified as IgG1(kappa) of IgG isotypes with affinity constants (Kaff) ranging from 1 x 10(-8) to 2.8 x 10(-10). GICA had a sensitivity of 86.66%; and specificity of 96.43%; for Plasmodium falciparum detection compared with routine microscopic examination. CONCLUSION: The established GICA is rapid and accurate for Plasmodium falciparum detection with such potential utility as for instant diagnosis of Plasmodium falciparum malaria.

Animals↗
PubMed2005

[Construction and immunological study of recombinant hBD-2/PSMA chimeric protein eukaryotic expressive plasmid].

The recombinant PSMA DNA vaccine for active immunotherapy of prostate cancer was investigated. Two DNA vaccine recombinant plasmids, pcDNA3.1/PSMA and pcDNA3.1/hBD-2-PSMA, were constructed by inserting the hBD-2 gene and PSMA gene into an eukarytoic expression vector pcDNA3.1. Expression of the two recombinants was detected in transfected COS-7 cells and inoculated mouse muscular cells by RT-PCR and immunohistochemical method. When immunized with pcDNA3.1/PSMA and pcDNA3.1/hBD-2-PSMA, the immunized BALB/c mice acquired specific antibody and T cell response to PSMA. The quantity of the spleen lymphocytes and their CTL activity against PSMA gene transfected-BALB/3T3 cells significantly increased in the immunized mice, and the CTL activity of lymphocytes from pcDNA3.1/hBD-2-PSMA immunized mice was significantly higher than that of pcDNA3.1/PSMA immunized mice. This result suggests that pcDNA3.1/hBD-2-PSMA would probably be developed as a DNA vaccine for the immunotherapy of prostate cancer.

3T3 Cells↗
PubMed2005

[Self-efficacy and health behaviors associated with osteoporosis of middle-aged and elderly subjects: a community-based study].

OBJECTIVE: To understand the self-efficacy and health behaviors with respect to osteoporosis of middle-aged and elderly subjects living in the communities. METHOD: A questionnaire-based survey was conducted using the Osteoporosis Self-Efficacy Scale (designed by Horan M et al ) and osteoporosis health behaviors questionnaire designed by the authors among 290 middle-aged and elderly community dwellers. RESULTS: The mean score of the self-efficacy was 64.75+/-21.93 among the involved subjects without statistically significant difference in terms of gender and intensity of daily exercises. The score differed significantly, however, in terms of age, education background, occupation and calcium intake (P<0.01), and the older group had lower scores than the younger one. Subjects with higher education scored higher, and those with occupations of physical labor scored lower than those with intellectual work. Higher calcium intake also resulted in higher score. It was found in this survey that the preventive behaviors of osteoporosis was insufficient among the subjects, and only 36.1% of the subjects drank milk everyday with 29.2% often having bean products and 49.8% taking calcium supplements. A percentage of 58.1% of these subjects regularly took part in exercises more than 3 times a week for over 30 minutes each time, but they seldom took weight bearing exercise. CONCLUSION: Osteoporosis self-efficacy should enhanced to promote osteoporosis- preventing behaviors in osteoporosis education program for the middle-aged and elderly people in the communities.

Aged↗
PubMed2005

[Clinical and laboratory screening studies on urea cycle defects].

OBJECTIVE: To investigate the incidences of urea cycle defects (UCDs) in the patients with hyperammonemia and study their etiology, clinical and laboratory features. METHODS: In the past 7 years, 26 cases (10.2%) of UCDs were detected from 254 patients with hyperammonemia. The etiological diagnoses were made by blood amino acids analysis, urinary organic acid analysis and blood acylcarnitine profile analysis. Three patients with citrullinemia type II were further confirmed by liver pathological analysis and gene diagnosis. RESULTS: Among 26 cases with UCDs, 15 had ornithine transcarbamylase (OTC) deficiency, 5 had citrullinemia type I, 3 had citrullinemia type II and 3 patients had arginemia. The age of onset of the patients ranged from 3 days to 13 years. Three cases (11.5%) developed hyperammonemic encephalopathy during neonatal period. Thirteen (50.0%), 7 (26.9%) and 3 (11.5%) cases developed clinical symptoms at the age of 1 to 12 months, 1 to 3 years and 6 to 13 years, respectively. Positive family history was found in 11 cases (42.3%). Among 26 patients with UCDs, 9 (34.6%) were hospitalized with the complains of seizures, psychomotor retardation, vomiting and unconsciousness, 8 (30.8%) with recurrent vomiting, headache and coma, 6 due to liver dysfunction. Intrahepatic cholestatic jaundice was found in 3 patients with citrullinemia type II. Blood ammonia ranged from 58 to 259 micromol/L on their first visit to our hospital. Twenty cases (76.9%) had liver dysfunction, 4 patients (15.4%) were diagnosed postmortem. Twenty-one patients got treatment and were followed up. Among them, 7 cases died of hyperammonemic encephalopathy or upper alimentary tract bleeding. Clinical improvement was observed in 14 cases. A boy with OTC deficiency who received a partial liver transplant from his mother showed normal general condition for two years. CONCLUSIONS: UCDs are the most frequent causes of congenital hyperammonemia. In this study, 26 patients (10.2%) with UCDs were identified from 254 patients with hyperammonemia resulting in encephalopathy and liver dysfunction. Early diagnosis and treatment can contribute a lot to improve the prognosis of the patients. Blood ammonia assay and further etiological analysis should be considered in the differential diagnosis of neurological and hepatic abnormality.

Adolescent↗
PubMed2005

[Charactierization of a new antibacterial polypeptide isolated from human cervical mucas].

OBJECTIVE: To identify a new antibactrial polypeptide HCP-1 isolated from human cervical mucus. METHODS: HCP-1 was isolated and purified from human cervical mucus, and N-terminal sequence of HCP-1 was determined. A degenerate primer was designed according to CodeHop methods, and an "anchor-oliga-dT primer" was used for the synthesis of cDNA. Using the degenerate primer and anchor-primer, cDNAs were amplified by PCR. The PCR products were cloned, sequenced, and analyzed by biological software. RESULTS: N-terminat sequence of HCP-1 was PKRKAEGDAK. The full length of HCP-1 cDNA was isolated and of which the sequence was the same as HMG-17. OMIGA software analysis indicated that this molecule contained an alpha-helix region. CONCLUSION: The new antibacterial polypeptide isolated from human cervical mucus is HMG-17. It may play a role in the human cervical mucus and the alpha-helix domain may be its antibacterial activity region.

Adult↗