A phage-displayed single chain variable fragment that interacts with hepatitis B core antigen: library construction, selection and diagnosis.

BACKGROUND: Phage display is an alternative method for constructing and selecting antibodies with desired specificity towards an antigen. OBJECTIVES: To construct a library of single chain variable fragment (ScFv) towards hepatitis B core antigen (HBcAg). To isolate a ScFv phage clone that interacts with HBcAg and to develop a phage-ELISA for detecting the antigen. STUDY DESIGN: Mice were inoculated with HBcAg and RNA was extracted from their spleen cells. The genes encoding heavy (V(H)) and light (V(L)) chains were amplified, linked via PCR and cloned into a phagemid vector. Phage particles displaying ScFv were panned against HBcAg and a selected clone was characterized and employed as a diagnostic reagent for detecting HBcAg in serum samples. RESULTS: A phage clone that interacts with HBcAg was selected from the antibody library. The binding of the phage to HBcAg was inhibited by a cyclic peptide bearing the WSFFSNI sequence. A phage-ELISA was established using the recombinant phage and as low as 10ng of HBcAg can be detected by the assay. CONCLUSION: The ScFv displayed on the surface of filamentous phage is an alternative choice for diagnosis of HBcAg in serum samples.     

人心连续组织切片的计算机三维重建

用表面重建法对人心连续组织切片进行了三维重建，重建中应用了火棉胶断面切片技术及图像拟合与插值处理，与以往的解剖断面重建比较，图像更加清晰、逼真，可从不同观察心脏的外部轮廓与内部结构。作为三维重建的基础性研究，可为心血管影像诊断提供解剖形态学依据和可靠的三维软件。     

Nonspecific induction of immunoglobulin M antibodies to periodontal disease-associated microorganisms after polyclonal human B-lymphocyte activation by Fusobacterium nucleatum.

The production of antibodies to oral bacteria was determined in lymphocyte cultures stimulated with sonicated Fusobacterium nucleatum, a potent inducer of polyclonal B-cell activation. After 9 days the cultures were examined by a microenzyme-linked immunosorbent assay for immunoglobulin M (IgM) antibodies to F. nucleatum, Bacteroides gingivalis, Actinomyces viscosus, and Streptococcus sanguis. Antibodies to these four bacteria were detected in cultures stimulated with polyclonal B-cell activation-inducing concentrations of F. nucleatum. However, significant concentrations of antibodies to F. nucleatum, but not to the other three microorganisms, were produced in cultures that received suboptimal polyclonal B-cell activation-inducing doses of F. nucleatum. Absorption studies indicated the specificity of the antibodies to each of the bacteria tested. IgM antibody production induced by F. nucleatum was enhanced by the addition of T cells. The production of IgM antibodies to the bacteria was reproducible in cultures from a single person tested on 3 consecutive days. The concentration of antibodies in replicate cultures, however, fluctuated greatly. To obtain consistent responses on successive days, multiple replicate cultures were required. These results suggest that F. nucleatum, which is frequently present in subgingival plaque, could induce the production of antibodies not only to F. nucleatum, but also to other microorganisms associated with periodontal diseases.     

N(omega)-nitro-L-arginine methyl ester attenuates lithium-induced c-Fos, but not conditioned taste aversion, in rats

Lithium chloride (LiCl) at doses sufficient to induce conditioned taste aversion (CTA) causes c-Fos expression in the relevant brain regions and activates the hypothalamic-pituitary-adrenal (HPA) axis. It has been suggested that nitric oxide (NO) in the central nervous system may play a role not only in the activation of HPA axis but also in CTA learning, and that LiCl may activate the brain NO system. To determine the role of NO in lithium-induced CTA, we examined the lithium-induced CTA, brain c-Fos expression, and plasma corticosterone level with Nomega-nitro-L-arginine methyl ester (L-NAME) pretreatment. Intraperitoneal L-NAME (30 mg/kg) given 30 min prior to LiCl significantly decreased lithium-induced c-Fos expression in the brain regions implicated in CTA learning, such as the hypothalamic paraventricular nucleus (PVN), central nucleus of amygdala (CeA), and nucleus tractus of solitarius. However, either the lithium-induced CTA acquisition or the increase in plasma corticosterone was not attenuated by l-NAME pretreatment. These results suggest that NO may be involved in lithium-induced neuronal activation of the brain regions, but not in the CTA acquisition or the HPA axis activation.     

Posttransplant primary cutaneous Ki-1 (CD30)+/CD56+ anaplastic large cell lymphoma

An anaplastic large cell lymphoma that was negative for Epstein-Barr virus and positive for Ki-1 (CD30) presented as a polypoid scalp mass in a 56-year-old man 16 years after renal transplantation. The lymphoma was of the CD4+ cytotoxic T-cell lineage, and the tumor cells also expressed CD56. Despite reduction in the dose of immunosuppression and localized radiotherapy, the tumor had rapidly progressed to involve the soft tissue of the right hand. Systemic chemotherapy induced complete regression of the soft tissue lesion. This case illustrates that posttransplant primary cutaneous CD30+ anaplastic large cell lymphomas may assume an aggressive clinical course but can still be controlled by systemic chemotherapy.     

Epidemiologic characteristics of death by poisoning in 1991-2001 in Korea

The purpose of this study was to investigate the epidemiologic characteristics of the death by poisoning in Korea. We recoded the Death Certificates Database by injury based on the short version of the International Classification of External Causes of Injuries (ICECI). We evaluated the mortality rate by total injury and poisoning, and analyzed the mortality rate by age, gender, year and month, toxic agent, and intent. Adjusted odds ratios were calculated to evaluate the effects of socioeconomic factors on suicidal poisoning death. The total number of death cases by injury was 346,656. The proportion of death cases by injury decreased from 13.53% of all death cases in 1991 to 11.89% in 2001. However, the mortality rate by poisoning increased rapidly from 1998, and then remained stable. The number of suicidal poisoning deaths has gradually increased, and its mortality rate was 6.41 (per 100,000) in 2001. Major toxic agents were pesticides and herbicides (50.90%) in 2001. Adjusted odds ratios of suicidal poisoning versus other poisonings showed significant differences in education attainment, region, and marital status. In conclusion, the mortality rate by poisoning has increased, and the proportion of suicidal poisoning also has increased compared to that of accidental poisoning.     

Loss of heterozygosity on chromosomes 3p,8p,9p and 17p in the progression of squamous cell carcinoma of the larynx

Previous molecular genetic studies of laryngeal squamous cell carcinoma (SCC)have shown certain chromosomal regions with recurring alterations. But studies of sequential molecular alterations and genetic progression model of laryngeal SCC have not been clearly defined. To identify the chromosomal alterations associated with the carcinogenesis of laryngeal SCC, we analyzed genomic DNA from microdissected squamous metaplasia, squamous dysplasia, invasive SCC, and metastatic carcinoma samples from 22 laryngeal SCC patients for loss of heterozygosity (LOH) at microsatellite loci. Ten microsatellite markers on chromosome 3p, 8p, 9p, and 17p were used. LOH at 9p21 was observed in the all stages including squamous metaplasia, squamous dysplasia, invasive SCC and metastatic carcinoma. LOH at 17p13.1, 3p25 and 3p14.2 was observed from the squamous dysplasia, invasive SCC and metastatic carcinoma. LOH at 8p21.3-p22 was observed mainly from the invasive SCC and metastatic carcinoma. The results suggest that 9p21 in the early event, 17p13.1, 3p25 and 3p14.2 in the intermediate event and 8p21.3- p22 in the late event may be involved in the laryngeal carcinogenesis.     

Analytical model to describe fluorescence spectra of normal and preneoplastic epithelial tissue: comparison with Monte Carlo simulations and clinical measurements

Fluorescence spectroscopy has shown promise for the detection of precancerous changes in vivo. The epithelial and stromal layers of tissue have very different optical properties; the albedo is relatively low in the epithelium and approaches one in the stroma. As precancer develops, the optical properties of the epithelium and stroma are altered in markedly different ways: epithelial scattering and fluorescence increase, and stromal scattering and fluorescence decrease. We present an analytical model of the fluorescence spectrum of a two-layer medium such as epithelial tissue. Our hypothesis is that accounting for the two different tissue layers will provide increased diagnostic information when used to analyze tissue fluorescence spectra measured in vivo. The Beer-Lambert law is used to describe light propagation in the epithelial layer, while light propagation in the highly scattering stromal layer is described with diffusion theory. Predictions of the analytical model are compared to results from Monte Carlo simulations of light propagation under a range of optical properties reported for normal and precancerous epithelial tissue. In all cases, the mean square error between the Monte Carlo simulations and the analytical model are within 15%. Finally, model predictions are compared to fluorescence spectra of normal and precancerous cervical tissue measured in vivo; the lineshape of fluorescence agrees well in both cases, and the decrease in fluorescence intensity from normal to precancerous tissue is correctly predicted to within 5%. Future work will explore the use of this model to extract information about changes in epithelial and stromal optical properties from clinical measurements and the diagnostic value of these parameters.     

Methods for derivation of human embryonic stem cells

The expanded blastocysts, developed from 2PN-stage embryos, are generally divided into three categories: a good blastocyst containing a large and distinguishable inner cell mass (ICM), a blastocyst with a small and distinct ICM, and a blastocyst with a poorly defined ICM. In this study, we introduce methods for the derivation of human embryonic stem cells (hESCs) depending on the quality of the blastocysts. An immunosurgical method was used for the good expanded blastocysts. This method, however, raises the probability of ICM loss in cases of hESC derivation from blastocysts with smaller or indistinct ICMs. Furthermore, this method is also associated with a risk of the contamination of the hESCs with animal pathogens. To overcome these shortcomings, the partial- or whole-embryo culture method was used. For blastocysts with no visible ICM, the whole-embryo culture method was used to establish hESCs via the seeding of the entire blastocyst without its zona pellucida directly on a STO feeder layer. However, trophectodermal overgrowth tends to hinder the expansion of the ICM during the initial steps of hESC derivation. Therefore, the partial-embryo culture method was developed to establish hESCs from blastocysts with smaller ICMs. The surgical isolation of the region containing the ICM with an ultra-fine glass pipette alleviates trophectoderm overgrowth. This method is also applicable to blastocysts with large and distinct ICMs, and the efficiency of this method is comparable to that of the immunosurgical method.