Biomedical subjects
Publications and source records attributed to C T Lin.
Pseudoaneurysm of the splenic artery has been rarely reported and the Doppler echocardiographic finding seldom described. Herein we report a rare case of huge extrasplenic pseudoaneurysm, which was detected by color flow Doppler ultrasonography and successfully treated by ligation of the splenic artery and resection of the pseudoaneurysm.
Venous thrombosis induced by a transvenous permanent pacemaker is a common complication. However, superior vena cava (SVC) syndrome caused by pacemaker leads is only occasionally seen and its prevalence has been estimated to be less than 1 in 1000 pacemaker patients. Herein, we report a Taiwanese patient of high grade AV block, who presented with SVC syndrome 2 years after transvenous permanent pacemaker implantation. This case features fibrotic stenosis of the junction of right brachiocephalic trunk and SVC, and an extensive thrombus formation resulting in complete obliteration of the left brachiocephalic vein. The collateral circulation was so delicate that he still could lead a rather normal life, even if anticoagulant therapy proved to be ineffective from an angiographic point of view.
The cadherin/catenin complex plays a key role in the initiation of cell-cell recognition, and adhesion, and the elaboration of structural and functional organization in multicellular tissues and organs. It is associated with tumor metastasis and also acts as an "invasion suppressor" of cancer cells. Nasopharyngeal carcinoma (NPC) is notorious for its highly metastatic nature. The expression of the E-cadherin/catenin complex is down-regulated in NPC tumor specimens. To obtain better insight into the intercellular adhesive property of NPC cells, we used immunofluorescence microscopy, immunoprecipitation, and immunoblot analysis to examine the expression of the classical cadherins and beta-catenin in a NPC cell line, TW-039. The results demonstrate a change in the distribution of E-cadherin from cytosolic flakes to cell-cell contacts with increasing time in culture. Between days 1 and 5 after plating, the detergent-insoluble fraction of E-cadherin increased from 20% to 37% of total E-cadherin, and that for P-cadherin increased from 33% to 40%. By contrast, the values for beta-catenin remained unchanged (26% and 25%). Both immunofluorescence and immunoblot studies suggested that P-cadherin may be involved in pioneer contact adhesion of TW-039 cells. Interestingly, E-, P-, and N-cadherin are co-expressed in this cell line. Immunoprecipitation studies also showed that other members of the cadherin family may be involved in the contact adhesion of TW-039 cells.
BACKGROUND: This study was designed to determine whether screening for high-risk human papillomaviruses testing could improve the detection of cervical dysplasia and cancer in assistance with conventional Papanicoloau (Pap) smears. METHODS: The study was based on 114 patients with abnormal Pap smears referred for colposcopy from Feb. 1997 to Dec. 1997. The presence of high-risk human papillomavirus (HPV) DNA was determined with the Hybrid Capture method (including HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, and 68). Cytologic examination by Papanicolaou smear was based on the Bethesda system and cervical biopsy was done via colposcopy. RESULTS: Cytologic examination demonstrated high-grade squamous intraepithelial lesions (HSIL) in 24 patients with HPV positive (75%), low-grade squamous intraepithelial lesions (LSIL) in 38 with 61% HPV positive, and atypical squamous cells of undetermined significance (ASCUS) in 52 with 37% HPV positive. Among patients with a cytologic diagnosis of borderline abnormalities (ASCUS or LSIL), those with who were HPV positive were significantly more likely to have cervical dysplasia (both p < 0.05). The sensitivity of combined HPV assay and/or cytology for detection of noninvasive precursor (91%) was significantly greater than those of cytology (68%) or HPV assay (81%) alone. CONCLUSION: The addition of the hybrid capture high-risk HPV DNA assay to cytologic examination of cervical smears appears to increase the sensitivity of cervical screening. Our findings suggest that HPV DNA may be a useful adjunct marker for early detection of cervical dysplasia in women with minimally abnormal Pap smears (ASCUS/low-grade SIL).
The prognosis of recurrent endometrial carcinomas is generally poor, except for isolated vaginal relapse. We report a case of recurrent endometrial cancer in a 58-year-old woman who initially received a type I extended hysterectomy with bilateral salpin-go-oophorectomy and bilateral para-aortic and pelvic lymph node dissection. The first recurrence occurred in the left parametrium 7 months after the primary surgery. The salvage therapy consisted of radiotherapy combined with hormonal therapy (tamoxifen and Megace). Complete remission was achieved initially. Subsequently, the patient accepted six courses of chemotherapy (cisplatin and Adriamycin) for progressive elevation of cancer antigen 125 (CA-125). The CA-125 levels remained elevated with titers fluctuating around 100 U/ml until a second recurrence at the left iliac 75 months following salvage therapy. The second salvage treatment consisted of maximal debulking of the pelvic mass and intraoperative radiotherapy (IORT) followed by four courses of chemotherapy with paclitaxel and carboplatin. Complete remission was again accomplished, with clinical investigations and molecular markers returning to normal. The patient has been clinically free of disease for more than 2 years since the second relapse of cancer. In this particular case, we found that repeated recurrence could occur after a long complete remission following salvage therapy; however, the disease could be recontrolled with further aggressive salvage efforts. A multimodality approach with combinations of radical resection, IORT, and paclitaxel-based chemotherapy can be offered to patients with localized recurrent or repeatedly recurrent endometrial carcinoma after previous cisplatin-based chemotherapy and pelvic radiation.
Previously, we have identified the association of G protein beta subunit (Gbeta) with mitotic spindles in various mammalian cells. Since microtubules are the main component of mitotic spindles, here we have isolated bovine brain microtubules and purified Gbeta subunit to identify the close association of Gbeta subunit with purified brain microtubules and have shown the direct incorporation of Gbeta subunit into the microtubules both in vitro and in vivo. It was found that: (1) microtubular fraction isolated from bovine brain contained Gbeta subunit, (2) coimmunoprecipitation demonstrated that Gbeta subunit could be coprecipitated with tubulin, (3) addition of purified Gbeta subunit into cytosolic extract for microtubule assembly caused direct incorporation of Gbeta subunit into assembled microtubules and increased the association of microtubule-associated proteins with microtubules, and (4) incubation of exogenous Gbeta subunit with detergent-permeabilized cells resulted in direct incorporation of Gbeta subunit into microtubule fibers and depolymerized tubulin molecules. We conclude that G protein beta subunit is closely associated with microtubules and may play an important role in the regulation of microtubule formation in addition to its regulatory role in cellular signal transduction.
A subtractive cDNA cloning strategy was used to isolate canine retina-specific genes. Canine phosducin cDNA was cloned from a canine subtracted retinal cDNA library and was analysed as a candidate for canine generalized progressive retinal atrophies (gPRA). Canine phosducin cDNA is 1230 bp in length encoding 245 amino acids. The nucleotide and amino acid sequences of canine phosducin are highly conserved when compared with those of five other mammalian species, namely human, cat, cow, rat, and mouse. Northern blot analysis demonstrated that the mRNA transcript for phosducin was approximately 1.3 kb in size and was present in canine retina, but showed no visible signals in 13 other canine tissues. The phosducin gene was examined for polymorphisms in a total of 101 pedigree dogs of eight breeds, including normal, obligate gPRA carriers, and gPRA-affected dogs, by single-stranded conformation polymorphisms (SSCP) analysis. Polymorphisms in the phosducin gene were detected only in the 3' untranslated region of the gene in two breeds of dogs: allelic heterozygous polymorphisms in miniature poodles suffering from one form of gPRA (progressive rod-cone degeneration, prcd), and a different polymorphism in a single normal Irish wolfhound. The polymorphisms of phosducin in prcd-affected miniature poodles did not segregate with the autosomal recessive form of gPRA. Heterozygous inheritance of the polymorphisms suggests that phosducin is very unlikely to carry the mutation causing prcd, so phosducin was probably excluded as a candidate for prcd-affected miniature poodles in this study.
Microbial risk assessment is emerging as a new discipline in risk assessment. A systematic approach to microbial risk assessment is presented that employs data analysis for developing parsimonious models and accounts formally for the variability and uncertainty of model inputs using analysis of variance and Monte Carlo simulation. The purpose of the paper is to raise and examine issues in conducting microbial risk assessments. The enteric pathogen Escherichia coli O157:H7 was selected as an example for this study due to its significance to public health. The framework for our work is consistent with the risk assessment components described by the National Research Council in 1983 (hazard identification; exposure assessment; dose-response assessment; and risk characterization). Exposure assessment focuses on hamburgers, cooked a range of temperatures from rare to well done, the latter typical for fast food restaurants. Features of the model include predictive microbiology components that account for random stochastic growth and death of organisms in hamburger. For dose-response modeling, Shigella data from human feeding studies were used as a surrogate for E. coli O157:H7. Risks were calculated using a threshold model and an alternative nonthreshold model. The 95% probability intervals for risk of illness for product cooked to a given internal temperature spanned five orders of magnitude for these models. The existence of even a small threshold has a dramatic impact on the estimated risk.
Using confocal laser scanning microscopy with a dual-wavelength laser system, the behaviour of BCECF [(2',7'-bis-2-carboxyethyl)-5-(and-6)carboxyfluorescein] was investigated in a variety of cell lines. Selection of a small area for monitoring allowed discrimination between various intracellular organelles, whose identity was established by vital staining. It was found that, after loading the cells with BCECF, both the nucleus and the mitochondria showed a higher level of fluorescence than the cytoplasm. Calibration of the pH-sensitivity of these fluorescence signals using the nigericin method yielded identical curves, as did exposure of the cells to NH4Cl. These studies suggest that BCECF, despite its inhomogeneous intracellular distribution, reports the pH of only one cellular compartment, the cytosol.
The differential diagnosis of familial macrothrombocytopenia and idiopathic thrombocytopenic purpura (ITP) may be difficult owing to the similarities in their clinical and laboratory presentations, but it is important because of dissimilarities in their management and prognosis. We investigated two families with familial macrothrombocytopenia and granulocyte inclusion. The probands of both families presented with mild bleeding tendency, macrothrombocytopenia, normal bone marrow, and increased percentages of platelet-associated immunoglobulin G (IgG) and reticulated platelets. ITP had been misdiagnosed in both patients initially. Both probands failed to respond to steroid therapy. Family study revealed an autosomal dominant pattern of heredity in both families, with absence of Alport's syndrome-like features (hearing impairment, congenital cataract, and interstitial nephritis). All thrombocytopenic family members showed blue cytoplasmic inclusions in neutrophils on peripheral blood smears. Ultrastructurally, distinct granulocyte inclusions comprising clusters of rough endoplasmic reticulum, smooth endoplasmic reticulum, and polysomes were detected, without the presence of parallel filaments. The clinical, laboratory, and hereditary findings were consistent with a diagnosis of Sebastian platelet syndrome in both families. In conclusion, caution should be exercised when interpreting the percentages of platelet-associated IgG in thrombocytopenic patients, as overinterpretation may lead to misdiagnosis of macrothrombocytopenia as ITP. Family history is important, as familial ITP is rare, and careful examination of blood smears is essential.
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Here we describe a two-stepped photobiotin-based procedure to enrich a target (canine retinal) cDNA library for tissue specific clones without removing those containing repetitive ( SINE ) elements, despite the presence of these elements in the driver population. In a first hybridization excess SINE elements were hybridized to a driver (canine cerebellar) cDNA. In a second hybridization target cDNA was added to this reaction. The resulting cDNA library was enriched for retinal specific clones, but contained the same ratio of clones with SINE elements found in the unsubtracted library.
Small inverted repeats (small palindromes) on plasmids have been shown to mediate a recombinational rearrangement event in Escherichia coli leading to the formation of inverted dimers (giant palindromes). This recombinational rearrangement event is efficient and independent of RecA and RecBCD. In this report, we propose a cruciform-dumbbell model to explain the inverted dimer formation mediated by inverted repeats. In this model, the inverted repeats promote the formation of a DNA cruciform which is processed by an endonuclease into a linear DNA with two hairpin loops at its ends. Upon DNA replication, this linear dumbbell-like DNA is then converted to the inverted dimer. In support of this model, linear dumbbell DNA molecules with unidirectional origin of DNA replication (ColE1 ori ) have been constructed and shown to transform E.coli efficiently resulting in the formation of the inverted dimer. The ability of linear dumbbell DNA to transform E.coli suggests that the terminal loops may be important in bypassing the requirement of DNA supercoiling for initiation of replication of the ColE1 ori.
The long terminal repeats of maedi visna virus strain 1514 contain a consensus AP-1 binding site which has been shown to be important in controlling virus transcription. However, this consensus site is absent in strain EV-1. Here, we have compared the ability of oligonucleotides corresponding to LTR sequences from EV-1 with those from 1514 to bind transcription factors in competitive gel retardation assays and activate reporter gene expression. The experiments demonstrated no observable binding of AP-1 to the EV-1-derived sequences and significant differences in the abilities of the 1514 and EV-1 sequences to activate transcription. However, both viral sequences interacted with a second, previously undetected, transcription factor. This factor gave specific gel shifts which were competed by an oligonucleotide containing the consensus sequence for the AML/PEBP2/CBF family of transcriptional factors, but not by control AP-1 or OCT-1 oligonucleotides. The factor was therefore denoted AML (vis). A second AML (vis) site, noted upstream of the TATA box proximal AP-1 site, gave single shifts which were competed by the downstream AML (vis) oligonucleotide. Both sites were functional in transfection assays. In gel shift retardation assays, polyclonal antisera directed against known runt domain proteins were able to supershift part of the AML (vis) binding activity in nuclear extracts from physiologically relevant cell types. The results thus suggest that the AML (vis) binding factor belongs to the AML/PEBP2/CBF family of transcription factors and may be important in controlling virus replication in these and other strains of ruminant lentiviruses.
This study evaluates the effects of a food simulating solution (75 vol% ethanol/water) and an artificial saliva (Moi-Stir) on dentine structure and chemistry, using scanning electron microscopic examination and Fourier transform infra-red (FTIR) microscopic characterization. The effect on the bonding of composites to the conditioned dentine were evaluated by shear bond strength (SBS) tests. Three adhesive/composite systems were examined: Tenure/Marathon One, Scotchbond Multi-Purpose/Z100, and Optibond/Herculite XRV. Control specimens were stored in either distilled water or tested without storage. Dentine surface exposure to ethanol resulted in partial loss of the smear layer and of plugs, as well as possible perturbation of collagen. Dentine surfaces exposed to artificial saliva or to distilled water had no evidence of any change from normal appearance of the smear layer. The measured FTIR spectra for most specimens conditioned in these two liquids appeared to be similar to those obtained from fresh dentine. SBS data were analysed using ANOVA and the Tukey LSD test. The SBS value for the non-preconditioned control (23.0 +/- 3.7 MPa) or for the dentine preconditioned in distilled water (22.9 +/- 4.2 MPa) was significantly higher (P < 0.05) than that for dentine pre-conditioned in ethanol (20.0 +/- 3.5 MPa). The SBS (13.3 +/- 3.4 MPa) of all bonding systems was reduced by 40-50% (P < 0.001) when artificial saliva pre-conditioned dentine was used. The failure mode at the dentine-bonding agent interface for the artificial saliva group was adhesive in nature. This is in contrast to the complex cohesive fracture mode found in the control groups and in most ethanol conditioned groups. Dentine structure and chemistry, shear bond strength, and the subsequent debonded mode can be significantly affected by exposure to oral environment prior to conditioning.
Generalized progressive retinal atrophy (gPRA) represents a genetically heterogenous group of retinal degenerations affecting pedigree dogs. Currently, we are using a candidate gene approach in an attempt to identify mutations causing gPRA in dogs. Here we report the cloning, sequencing and analysis of canine rom-1, a structural gene of the rod photoreceptor. Single-stranded conformation polymorphism (SSCP) analysis was used to look for polymorphisms segregating with gPRA in the English cocker spaniel, Labrador retriever, miniature poodle, miniature long-haired dachshund, Tibetan terrier, miniature schnauzer, Cardigan Welsh corgi and Irish wolfhound. Further investigation involved DNA sequencing and restriction fragment length polymorphism (RFLP) analysis. Our studies revealed the presence of three polymorphisms, none of which segregated with disease phenotype. Haplotype analysis identified four rom-1 alleles. Our results indicate that rom-1 is unlikely to be a cause of gPRA in the breeds of dog examined.
Recently, we have established nine nasopharyngeal carcinoma (NPC) cell lines in which only one cell line showed the p53 mutation. For investigation of the p53 mutation in this line, immunostaining using anti-p53 antibody was applied and showed the presence of p53 protein in the cytoplasm but not in the nucleus. Single strand conformation polymorphism analysis of the p53 gene showed one normal and one additional DNA band. Cloning and sequencing of PCR-amplified DNA showed an AGA (arginine) to ACA (threonine) heterozygous point mutation at codon 280. Transfection of the p53 DNA binding sequence and chloramphenicol acetyltransferase assay revealed loss of transcriptional activation function of endogenous p53 protein. Co-localization of the endogenous and the transfected exogenous p53 protein by polyclonal antibodies to anti-p53 protein revealed strong exogenous p53 staining in the transfected nuclei and weak staining of endogenous p53 protein in the cytoplasm. We concluded that (a) a heterozygous point mutation at codon 280 was identified in the NPC-TW 06 cell line; (b) the point mutation may cause the stagnation of mutant p53 protein in the cytoplasm, and loss of its transcriptional activation function; (c) endogenous and exogenous p53 protein can be co-localized at the same time in the transfected cells; and (d) 280 mutant p53 protein in NPC cells does not cause a decrease or increase in sensitivity to chemotherapy.
At National Taiwan University Hospital, from 1986 to 1996, autopsies were performed on 16 patients with acquired immunodeficiency syndrome. There were 15 men and 1 woman. Fourteen of these male patients had contracted the disease as a result of sexual practice, among which 9 were homosexual, 1 was bisexual and 4 were heterosexual. One of the patients had become infected by sharing a syringe during intravenous drug use. The female was a sex worker. Among these patients, only 2 had been tested for HIV before developing AIDS. On autopsy, lymphoid depletion and thymus atrophy were found in all patients. Testicular atrophy was noted in all the male patients. Three patients died of malignant lymphoma. Twelve patients died of opportunistic infections and 1 committed suicide. The initial opportunistic infection was usually oral candidiasis. Pneumocystis carinii pneumonia (PCP) was the most common opportunistic infection developed in the early stage while cytomegalovirus (CMV) infection was the most common one found in the late stage. Mycobacterium infection had developed in 8 patients. Six patients had disseminated Kaposi's sarcoma (KS) and 4 of them were homosexual. In 4 patients, biopsy specimens were proved to have KS associated viral (HHV-8) genome. Malignant lymphoma was found in 4 cases, all were of high grade B cell type. Epstein-Barr virus (EBV) encoding small RNA (EBER1) was demonstrated in all the lymphomas. In conclusion, (1) the prevalence of tuberculosis (38%) in patients with AIDS in Taiwan is high; (2) the most common opportunistic infections in this series are candidiasis, PCP and CMV infections; (3) the incidence of AIDS related non-Hodgkin's lymphoma in Taiwan has increased since 1995.