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Biomedical subjects

Y H Hsu

Publications and source records attributed to Y H Hsu.

At least 37 records · Page 2Linked to original sources

A defective RNA associated with bamboo mosaic virus and the possible common mechanisms for RNA recombination in potexviruses.

A naturally occurring 1.1 kb RNA was isolated from purified virions of bamboo mosaic potexvirus isolate S (BaMV-S). This RNA is a defective RNA (D RNA) derived from a single internal deletion of the BaMV genome. A cDNA clone representing the complete nucleotide sequence of the BaMV-S D RNA was generated and its nucleotide sequence was determined. The BaMV D cDNA is 1015 nts in length [excluding the poly(A) tail] and consists of two regions corresponding to 867 nts of the 5' terminus and 148 nts of the 3' terminus of the BaMV genomic RNA. BaMV D cDNA contains a single open reading frame (ORF) encoding a putative 29.7 kDa protein comprised of a fusion of the first 258 amino acids of BaMV ORF 1 and the last 2 amino acids of coat protein. The coding capacity of D RNA was verified by in vitro translation of native BaMV-S D RNA and of 1.1 kb RNA transcribed in vitro from the full-length D cDNA. BaMV D RNA can be reproducibly generated by serial passages of BaMV-S in Nicotiana benthamiana and is the first D RNA in the potexvirus group shown to be generated de novo. Alignments of sequences surrounding the 5' and 3' junction borders of reported potexvirus D RNAs reveal a 65.2-84.6% sequence identity, suggesting that common mechanisms for viral RNA recombination are involved in the generation of potexvirus D RNAs.

Amino Acid Sequence↗

Identification of the RNA-binding sites of the triple gene block protein 1 of bamboo mosaic potexvirus.

The triple gene block protein 1 (TGBp1) encoded by open reading frame 2 of bamboo mosaic potexvirus (BaMV) was overexpressed in Escherichia coli and purified in order to test its RNA-binding activity. UV crosslinking assays revealed that the RNA-binding activity was present mainly in the soluble fraction of the refolded TGBp1. The binding activity was nonspecific and salt concentration-dependent: activity was present at 0-50 mM NaCl but was almost abolished at 200 mM. The RNA-binding domain was located by deletion mutagenesis to the N-terminal 3-24 amino acids of TGBp1. Sequence alignment analysis of the N-terminal 25 amino acids of the TGBp1 homologues of potexviruses identified three arginine residues. Arg-to-Ala substitution at any one of the three arginines eliminated most of the RNA-binding activity, indicating that they were all critical to the RNA-binding activity of the TGBp1 of BaMV.

Amino Acid Sequence↗

Sufficient length of a poly(A) tail for the formation of a potential pseudoknot is required for efficient replication of bamboo mosaic potexvirus RNA.

RNAs transcribed from a full-length infectious cDNA clone of the bamboo mosaic potexvirus (strain O) genome, pBaMV-O, were infectious to Nicotiana benthamiana plants. Mutant genomes in which the poly(A) tail is absent or replaced by a 3' tRNA-like structure from turnip yellow mosaic virus RNA failed to amplify detectably in N. benthamiana protoplasts. No amplification was detected in protoplasts inoculated with transcripts containing 4, 7, or 10 adenylate residues at the 3' end, whereas transcript inocula with 15 adenylate residues resulted in coat protein accumulation to a level 26% of that resulting from inoculation with transcripts with 25 adenylate residues (designated as wild type). Coat protein accumulation levels of 69 and 98% relative to wild type were observed after inoculation of protoplasts with transcripts bearing poly(A) tails 18 and 22 nucleotides long, respectively. The presence of a putative 3' pseudoknot structure including at least 13 adenylate residues of the 3'-terminal poly(A) tail was supported by enzymatic and chemical structural analysis. The functional relevance of this putative pseudoknot was tested by mutations that affected basepairing within the pseudoknot. These results support the existence of functional 3' pseudoknot that includes part of the 3' poly(A) tail.

Base Sequence↗

Bamboo mosaic potexvirus satellite RNA (satBaMV RNA)-encoded P20 protein preferentially binds to satBaMV RNA.

A satellite RNA of 836 nucleotides [excluding the poly(A) tail] depends on the bamboo mosaic potexvirus (BaMV) for its replication and encapsidation. The BaMV satellite RNA (satBaMV) contains a single open reading frame encoding a 20-kDa nonstructural protein (P20). The P20 protein with eight histidine residues at the C terminus was overexpressed in Escherichia coli. Experiments of gel retardation, UV cross-linking, and Northwestern hybridization demonstrated that purified P20 was a nucleic-acid-binding protein. The binding of P20 to nucleic acids was strong and highly cooperative. P20 preferred binding to satBaMV- or BaMV-related sequences rather than to nonrelated sequences. By deletion analysis, the P20 binding sites were mainly located at the 5' and 3' untranslated regions of satBaMV RNA, and the RNA-protein interactions could compete with the poly(G) and, less efficiently, with the poly(U) homopolymers. The N-terminal arginine-rich motif of P20 was the RNA binding domain, as shown by in-frame deletion analysis. This is the first report that a plant virus satellite RNA-encoded nonstructural protein preferentially binds with nucleic acids.

Amino Acid Sequence↗

Clinical experience with endoscopic stents for treatment of common bile duct stones.

Endoscopic removal of common bile duct (CBD) stones after endoscopic sphincterotomy (EST) is now a widely accepted procedure. Surgery is usually recommended when extraction of stones after EST fails. For patients with major medical problems or who are at high surgical risk, however, endoscopic stent placement may help to prevent stone impaction and cholangitis. In this report, we describe the long-term effects and complications of biliary stent use in elderly patients with CBD stones. From August 1995 to June 1998, 19 patients with CBD stones underwent stent placement by duodenoscopy. Three of these patients underwent this procedure for temporary treatment while awaiting surgery or EST. In the remaining 16 patients (6 men and 10 women, mean age 76 +/- 10 years), invasive management carried a high risk of complications. We used a 7F straight stent for the first patient, while the remaining 15 received 7F pigtail stents. During a mean follow-up period of 34 months, two patients were lost to follow-up and two patients had migration of the stents. Three patients had acute cholangitis with stents in situ. Of these, one underwent stent exchange 8 months later, while the CBD stones were cleared either by endoscopy or surgery in the other two patients. Five patients died of nonbiliary diseases during the follow-up period. Our results show that long-term biliary stent placement is an advisable alternative therapeutic modality for high-risk and debilitated patients with CBD stones.

Aged↗

Biological responses of dog prostate and adjacent structures after meso-tetra-(m-hydroxyphenyl) chlorin and aluminum disulfonated phthalocyanine based photodynamic therapy.

Further to our work on the feasibility of application of photodynamic therapy (PDT) to the canine prostate, this study evaluates the biological responses of the prostate and adjacent vital structures with meso-tetra-(m-hydroxylphenyl) chlorin (mTHPC) or aluminum disulfonated phthalocyanine (AlS2Pc) based PDT as a preparatory step for clinical trials. Skin photosensitivity was not particularly problematic if light protection could be implemented properly for 2 weeks following sensitization. Prostate PDT was well tolerated by the experimental animals with only minor physical distress. mTHPC was more powerful than AlS2Pc in terms of prostate lesions induced. A large portion of prostate tissue could be destroyed by PDT with 4 punctures. Physical distress was probably caused by severe urethral irritation and aching from acute swelling of the prostate. Although the voiding condition normalized within 10 days, regeneration of urethral epithelium was not complete until 3-4 weeks after PDT. Improper placement of laser fiber caused extensive ecchymosis of the retroperitoneal organs. The biological significance of PDT induced hyperemia in the periprostatic structures remains poorly defined. Neither periprostatic nerve damage nor rectal lesions were seen in dogs receiving either mTHPC or AlS2Pc. Glandular atrophy with papillary cystic regeneration of the prostate was the most prominent finding 90 days after PDT. The glandular architecture was well preserved because the interlobular collagens were less affected than the cellular components of the glands. Our study suggests that PDT with mTHPC and AlS2Pc is safe and promising for necrosing a substantial amount of prostate tissue. The completeness of treatment and long-term therapeutic effectiveness for prostate cancer, however, remains to be determined through further investigation.

Animals↗

The expression of cytokeratin 18 in transitional cell carcinoma comparing with hepatoma.

The epithelium in kidneys and urinary bladders contain CK18 as in liver cells. The modulation of cytokeratin 18 during tumor transformation in hepatoma had been previously recognized through a series of biochemical and immunological approaches. A 14 KD hepatoma related molecules was found in the previous studies. We would like to utilize the hepatoma transformation model to study the changes in CK18 in transitional cell carcinoma, using immunoblotting and western blotting techniques. The result is that transitional cell carcinoma retain their CK18 molecule. Furthermore, CK18 related molecules similar to those seen in hepatoma also present in transitional cell carcinoma. The conclusions are transitional cell carcinoma contains CK18 related proteins similar to those seen in hepatoma tissues. We suggest that this element would be responsible for the change during the malignant transformation processes.

Blotting, Western↗

Stabilizing of cytokeratin in PLC/PRF/5 cells.

There are two sorted groups of cytokeratin 18 (CK18) in forms of assembly and disassembly in PLC/PRF/5 cells. A subcellular shifting is found in association with conditions of microtubule networks. The finding shows that CK18 mostly in forms of assembly, when microtubule networks are in status. The result also reveals that CK18 is relatively in forms of disassembly, while microtubule networks are disrupted in steps. It indicates that intact microtubule networks are probably a stabilizing factor of assembled CK18. It implies that CK18 is not a stable molecule when the cell is under environmental stress.

Blotting, Western↗

Subgenomic RNAs of bamboo mosaic potexvirus-V isolate are packaged into virions.

Purified virions of bamboo mosaic potexvirus-V isolate (BaMV-V) were found to contain three major RNA species, the 6.4 kb genomic RNA and two RNAs of 2.0 and 1.0 kb, in addition to associated satellite RNA (0.85 kb). Results of Northern blot hybridization, primer extension analysis and cDNA sequencing showed that the packaged 2.0 and 1.0 kb RNAs of BaMV-V were subgenomic RNAs. In contrast, in the BaMV-O isolate, only genomic RNA was packaged and encapsidated subgenomic RNAs were not detectable. The transcription initiation sites for the 2.0 and 1.0 kb subgenomic RNAs of BaMV-V were located 1 1 and 16 nt upstream of the initiation codon of open reading frames (ORFs) 2 and 5, respectively. The 2.0 and 1.0 kb subgenomic RNAs functioned as messengers for the ORF2 protein and capsid protein, respectively. Packaging of the 1.0 kb subgenomic RNAs resulted in the formation of rod-shaped particles about 70 nm in length. Our results indicate that BaMV isolates have evolved distinctly for packaging of subgenomic RNAs.

Capsid↗

Identification and characterization of the Escherichia coli-expressed RNA-dependent RNA polymerase of bamboo mosaic virus.

Bamboo mosaic virus (BaMV), a member of the potexvirus group, infects primarily members of the Bambusoideae. The open reading frame 1 (ORF1) of BaMV encodes a 155-kDa polypeptide that was postulated to be involved in the replication and the formation of cap structure at the 5' end of the viral genome. To characterize the activities associated with the 155-kDa viral protein, it was expressed in Escherichia coli BL21(DE3) cells with thioredoxin, hexahistidine, and S. Tag fused consecutively at its amino terminus, and the fusion protein was purified by metal affinity chromatography. Several RNA fragments, prepared by in vitro transcription, were tested as substrates for the RNA-dependent RNA polymerase (RdRp) activity. Among them, the expressed fusion enzyme was able to generate a 32P-labeled RNA product when 3'-end RNA fragments of the positive strand or negative strand of BaMV were included in the assay mixture. Dot hybridization assay revealed that the reaction products are complementary to their RNA substrates. Taken together, the evidence suggests that the 155-kDa protein encoded by ORF1 of BaMV has an RdRp activity and should be involved in the replication of BaMV. Mutational analyses demonstrate the importance of the GDD motif in the polymerase activity, and deletion studies suggest that the polymerase activity resides in the carboxyl terminus of the 155-kDa viral protein.

Base Sequence↗

Focal segmental glomerulosclerosis after poststreptococcal glomerulonephritis in the elderly: a case report.

Acute poststreptococcal glomerulonephritis (PSGN) is quite rare in the elderly population. It is very difficult to recognize because the initial symptoms are similar to those of more common diseases of this age group. Compared with younger patients, elderly patients with PSGN tend to have a high incidence of renal function impairment and anuria or prolonged oliguria. Recovery from acute PSGN in the elderly is less predictable than in children and younger adults, especially when associated with severe impairment of renal function. We present the case of an elderly patient with PSGN, who had a skin infection of the face and anuria on admission. Percutaneous left renal biopsy was performed in the initial hospitalization period and in the recovery phase of the disease. Renal function recovered within two months. However, increased urinary white blood cell sediments with normal serum creatinine concentrations persisted in the successive three years of follow-up. Renal biopsy was repeated and showed evidence of focal segmental glomerulosclerosis.

Aged↗

Molecular evolution and phylogeny of satellite RNA associated with bamboo mosaic potexvirus.

Satellite RNA of bamboo mosaic potexvirus (satBaMV) is a linear RNA molecule which encodes a 20-kDa nonstructural protein. Sequences of seven different satBaMV isolates from bamboo hosts in three genera showed 0.7% to 7.5% base variation which spanned the whole RNA molecule. However, the putative 20-kDa open reading frame was all preserved in these isolates. The phylogenetic relationship based on the nucleotide sequence did not show particular grouping of satBaMV from the host in one genus; neither was the grouping of satBaMV evident by location of sampling. Putative secondary structures of the 3' untranslated regions showed a basic pattern with conserved hexanucleotides (ACCUAA) and polyadenylation signal (AAUAAA) located in the loop regions. Although the satBaMV-encoded 20-kDa protein is a nonstructural protein, its predicted secondary structure contains eight-stranded beta-sheets which may form "jelly-roll" structure similar to that found in capsid protein encoded by satellite virus of panicum mosaic virus.

Amino Acid Sequence↗

Intracranial meningeal melanocytoma: CT and MRI.

We report the MRI and CT findings of an intracranial meningeal melanocytoma (IMM) arising from Meckel's cave and review the imaging characteristics of IMM. On CT, IMM constantly appear as well-circumscribed, isodense to slightly dense, extra-axial tumours with homogeneous contrast enhancement. This appearance is non-specific and similar to that of meningiomas or small neuromas. On MRI, the signal of IMM is strongly related to the amount of melanin pigment: the more melanin, the more shortening of T1 and T2 relaxation times. Only when it shows as a homogeneous mass, bright on T1 and dark on T2 weighting, can a specific diagnosis of a melanin-containing tumour be made. However, this still cannot provide a distinction between IMM and malignant meningeal melanoma.

Adult↗

Reoperation for recurrent trigeminal neuralgia after microvascular decompression.

BACKGROUND: Microvascular decompression (MVD) is an effective technique for those who have trigeminal neuralgia (TN) but cannot tolerate, or show no response to medicine. Though the initial success rate is high, some patients may develop severe recurrent neuralgia, especially after a longer period of follow-up. The efficacy of reoperation needs to be evaluated. To know the possible risk factors of recurrence after initial MVD is mandatory to the management of recurrent TN. METHODS: Among the 80 cases of TN treated with MVD, five cases showed severe recurrent symptoms within a follow-up period from 9 months-4 years. The symptoms recurred on the same side of the face, and were unresponsive to medical treatment. Brain computed tomography (CT) and magnetic resonance imaging (MRI) may reveal the etiology of recurrence. Repeat decompression of the trigeminal nerve was the main goal of reoperation, which was done via a suboccipital approach. RESULTS: Over the past 17 years, 80 MVDs for TN have been performed at Chang Gung Memorial Hospital. There were five cases of serious postoperative recurrence, which could not be relieved by medicine. Recurrence occurred 1 day-12 months after the initial surgery. Three cases were due to vascular compression, while two were caused by the local effect of Teflon felt. Reoperation produced complete remission in four patients, and partial remission in one. CONCLUSIONS: An increasing number of patients may experience severe recurrent TN after initial MVD during a long period of follow-up. Reoperation is safe and beneficial for these patients, but the results are dependent on the etiology of the recurrence. Further vascular compression of the trigeminal nerve can be relieved by MVD. Otherwise, in cases of severe adhesion caused by Teflon, complete microneural lysis can achieve satisfactory results.

Adult↗

Arsenic methylation capacity, body retention, and null genotypes of glutathione S-transferase M1 and T1 among current arsenic-exposed residents in Taiwan.

In order to elucidate the relationships among arsenic methylation capacity, body retention, and genetic polymorphisms of glutathione S-transferase (GST) M1 and T1, a total of 115 study subjects were recruited from Lanyang Basin located on the northeast coast of Taiwan. Specimens of drinking water, blood, urine, hair and toenail were collected from each study subject. Urinary inorganic and methylated arsenic were speciated by high performance liquid chromatography combined with hydride-generation atomic absorption spectrometry. Arsenic concentration in hair and toenail were quantitated by atomic absorption spectrophotometry. The polymerase chain reaction was used to determine genetic polymorphisms of GST M1 and T1. Arsenic concentrations in urine, hair, and toenail of study subjects were positively correlated with arsenic levels in their drinking water. Percentages of various arsenic species in urine (mean +/- standard error (SE) were 11.8 +/- 1.0, 26.9 +/- 1.2 and 61.3 +/- 1.4, respectively, for inorganic arsenic, monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA). Men and women had similar arsenic methylation capability. No associations were observed between arsenic methylation capability and arsenic content in either drinking water or urine. Ratios of arsenic contents in hair and toenail to urinary arsenic content (mean +/- standard error) were 6.2 +/- 0.7 and 16.5 +/- 1.7, respectively. Genetic polymorphisms of GST M1 and T1 were significantly associated with arsenic methylation. Subjects having the null genotype of GST M1 had an increased percentage of inorganic arsenic in urine, while those with null genotype of GST T1 had an elevated percentage of DMA in urine. Arsenic contents in hair and toenail were significantly correlated with the increase in arsenic concentrations of drinking water and urine, while no significant associations were observed between arsenic contents in hair and toenail and polymorphisms of GST M1 and T1. The relationship between arsenic methylation capability and body retention was modified by genetic polymorphisms of GST M1 and T1. Arsenic contents in hair and toenail were negatively associated with MMA percentage and positively associated with DMA percentage among subjects having null genotypes of GST M1 and T1, but not among those with non-null genotypes.

Arsenic↗