BACKGROUND: Neurotrophin-3 (NT-3), a member of the NT family, has only been considered an ancillary compound that provides anti-apoptotic benefits by inactivating tropomyosin receptor kinase C (TrkC)-induced apoptotic signals. However, little is known about the clinical relevance of NT-3 expression itself in neuroblastoma. The purpose of this study was to assess NT-3 expression in patients with neuroblastoma and its relevance to clinicopathologic findings and treatment outcomes. METHODS: In this study, expression of NT-3 and TrkC was analyzed using immunohistochemistry in 240 patients with newly diagnosed neuroblastoma. RESULTS: The results of the study revealed that NT-3 expression was associated with older age at diagnosis, localized tumors, and more differentiated tumors but was not associated with early treatment response (degree of residual tumor volume after three cycles of chemotherapy) and progression-free survival (PFS). However, when analysis was confined to patients with MYCN amplified tumors, NT-3 expression was associated with better early treatment response with borderline significance (P = 0.092) and higher PFS (86.9% vs. 58.2%; P = 0.044). In multivariate analysis in patients with MYCN amplified tumors, NT-3 was independent prognostic factor (hazard ratio, 0.246; 95% confidence interval, 0.061–0.997; P = 0.050). In another subgroup analysis, the early treatment response was better if NT-3 was expressed in patients without TrkC expression (P = 0.053) while it was poorer in patients with TrkC expression (P = 0.023). CONCLUSION: This study suggests that NT-3 expression in neuroblastoma has its own clinical significance independent of TrkC expression, and its prognostic significance differs depending on the status of MYCN amplification and/or TrkC expression.
Diagnosis ; Disease-Free Survival ; Humans ; Immunohistochemistry ; Multivariate Analysis ; Neoplasm, Residual ; Neuroblastoma ; Phosphotransferases ; Tropomyosin

Pneumocystis carinii causes serious pulmonary infection in immunosuppressed patients. This study was undertaken to observe the cytoskeletal proteins of P. carinii by immuno-electron microscopy. P. carinii infection was experimentally induced by immunosuppression of Sprague-Dawley rats for seven weeks, and their lungs were used for the observations of this study. The gold particles localized actin, tropomyosin, and tubulin. The actin was irregularly scattered in the cytoplasm of the trophic forms but was much more concentrated in the inner space of the cell wall of the cystic forms called the inner electron-lucent layer. No significant amount of tropomyosin was observed in either trophic forms or cystic forms. The tubulin was distributed along the peripheral cytoplasm and filopodia of both the trophic and cystic forms rather than in the inner side of the cytoplasm. Particularly, in the cystic forms, the amount of tubulin was increased and located mainly in the inner electron-lucent layer of the cell wall where the actin was concentrated as well. The results of this study showed that the cell wall of P. carinii cystic forms is a structure whose inner side is rich in actin and tubulin. The location of the actin and tubulin in P. carinii suggests that the main role of these proteins is an involvement in the protection of cystic forms from the outside environment by maintaining rigidity of the cystic forms.
Actins/analysis ; Animals ; Cytoskeletal Proteins/*analysis ; Fungal Proteins/*analysis ; Histocytochemistry ; Microscopy, Immunoelectron ; Pneumocystis/*chemistry/cytology ; Rats ; Rats, Wistar ; Support, Non-U.S. Gov't ; Tropomyosin/analysis ; Tubulin/analysis

BACKGROUND AND OBJECTIVES: Left ventricle burdened by longstanding volume-overload, undergoes various structural and functional alterations. Accordingly, the expressions of multiple classes of genes are likely to be altered. However, the profile of gene expressions, specifically in a volume-overloaded left ventricle in humans, has not been explored. SUBJECTS AND METHODS: The pattern of gene expression was studied, using a cDNA microarray, in myocardium from 4 normal subjects and 5 patients with chronic regurgitant valvular heart disease whose end-diastolic left ventricular dimension measures 65 mm or more, but whose systolic function remained preserved. RESULTS: We identified 58 differentially expressed genes that were functionally classifiable in the volume-overloaded myocardium. Those genes involved in cell cycle/growth (up/down-regulation: 9/1), signal transduction (4/1) were mostly overexpressed in the volume-overloaded myocardium. The distributions of the gene expressions were variable for those involved in transcription/translation (up/down-regulation: 6/7) and apoptosis (2/2). The genes related to the myocyte structure (troponin T3, tropomyosin, etc)(up/down-regulation: 1/10), as well as those related to metabolism (2/5), were underexpressed. The gene expression patterns from RT-PCR and Western blot, with randomly selected genes, were similar to those from the cDNA microarray. CONCLUSION: Altered expression was identified in multiple genes in the volume-overloaded human left ventricle prior to the development of heart failure. The genes related to cell growth and signal transduction were mostly overexpressed, while those related to cellular structure and metabolism appeared to be underexpressed. These results might help in the elucidation of cellular mechanisms for the remodeling process associated with chronic volume-overloading.
Apoptosis ; Blotting, Western ; Cellular Structures ; Gene Expression* ; Heart Failure* ; Heart Valve Diseases ; Heart Ventricles ; Heart* ; Humans* ; Metabolism ; Muscle Cells ; Myocardium* ; Oligonucleotide Array Sequence Analysis ; Signal Transduction ; Transcriptome* ; Tropomyosin

OBJECTIVETo perform cloning of the gene encoding Chinese Schistosoma japonicum tropomyosin (SjcTM) and its expression in Escherichia coli.

METHODSSjcTM cDNA fragment, except for 14 amino acids at the amino terminus, was obtained by reverse transcriptase-polymerase chain reaction (RT-PCR) with total RNA extracted from adult worms of S. japonicum. The RT-PCR product was cloned into T vector and sequenced. The SjcTM cDNA, derived from the constructed TA clone pGEM-SjcTM, was then subcloned into the expressing vector pBV220. After characterization by agarose gel electrophoresis, endonucleases digestion and PCR, the resultant recombinant plasmid was used for expression under the temperature-dependent condition.

RESULTSThe RT-PCR product, cloned into a T vector, was sequenced and shown to be 96.5% identical at the nuclei acid level and 98.1% identical in deduced amino acid sequence to that of S. mansoni tropomyosin. The target DNA fragment was then subcloned into a prokaryotic vector pBV220. Induced expression in E. coli DH5alpha cells resulted in a constant level of recombinant protein production. The results of SDS-PAGE and Western blot revealed that the molecular weight of non-fusion recombinant protein (rSjcTM) was approximately 32 kDa and could be recognized specifically by a polyclonal antiserum specific for native S. japonicum tropomyosin (SjcTM).

CONCLUSIONThe engineering of the cDNA encoding S. japonicum tropomyosin and its bacterial expression was successfully made.

Amino Acid Sequence ; Animals ; Base Sequence ; Cloning, Molecular ; DNA, Complementary ; analysis ; chemistry ; Escherichia coli ; genetics ; Molecular Sequence Data ; Recombinant Proteins ; biosynthesis ; Schistosoma japonicum ; genetics ; Tropomyosin ; biosynthesis ; chemistry ; genetics

Renal cell carcinoma (RCC) is one of the most malignant tumors in urology, and due to its insidious onset patients frequently have advanced disease at the time of clinical presentation. Thus, early detection is crucial in management of RCC. To identify tumor specific proteins of RCC, we employed proteomic analysis. We prepared proteins from conventional RCC and the corresponding normal kidney tissues from seven patients with conventional RCC. The expression of proteins was determined by silver stain after two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). The overall protein expression patterns in the RCC and the normal kidney tissues were quite similar except some areas. Of 66 differentially expressed protein spots (p<0.05 by Student t-test), 8 different proteins from 11 spots were identified by MALDI-TOF-MS. The expression of the following proteins was repressed (p<0.05); aminoacylase-1, enoyl-CoA hydratase, aldehyde reductase, tropomyosin alpha-4 chain, agmatinase and ketohexokinase. Two proteins, vimentin and alpha-1 antitrypsin precursor, were dominantly expressed in RCC (p<0.05).
Aged ; Aldehyde Reductase/analysis ; Amidohydrolases/analysis ; Carcinoma, Renal Cell/*metabolism/pathology ; Comparative Study ; Electrophoresis, Gel, Two-Dimensional ; Enoyl-CoA Hydratase/analysis ; Female ; Fructokinases/analysis ; Humans ; Kidney Neoplasms/*metabolism/pathology ; Male ; Middle Aged ; Proteome/*analysis ; Proteomics/*methods ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Tropomyosin/analysis ; Ureohydrolases/analysis ; Vimentin/analysis ; alpha 1-Antitrypsin/analysis

PURPOSE: Gastric cancer is one of the most prevalent cancers worldwide. 5-fluorouracil (5-FU) and cisplatin are the most commonly used drugs for the treatment of gastric cancer. However, a significant number of tumors often fail to respond to chemotherapy. MATERIALS AND METHODS: To better understand the molecular mechanisms underlying drug resistance in gastric cancer the gene expression in gastric cancer cells, which were either sensitive or resistant to 5-FU and cisplatin, were examined using cDNA microarray analysis. To confirm the differential gene expression, as determined using the microarray, semiquantitative RT-PCR was performed on a subset of differentially expressed cDNAs. RESULTS: 69 and 45 genes, which were either up-regulated (9 and 22 genes) or down-regulated (60 and 25 genes), were identified in 5-FU- and cisplatin-resistant cells, respectively. Several genes, such as adaptor-related protein complex 1 and baculoviral IAP repeat-containing 3, were up-regulated in both drug-resistant cell types. Several genes, such as the ras homolog gene family, tropomyosin, tumor rejection antigen, protein disulfide isomerase-related protein, melanocortin 1 receptor, defensin, cyclophilin B, dual specificity phosphatase 8 and hepatocyte nuclear factor 3, were down-regulated in both drug-resistant cell types. CONCLUSION: These findings show that cDNA microarray analysis can be used to obtain gene expression profiles that reflect the effect of anticancer drugs on gastric cancer cells. Such data may lead to the assigning of signature expression profiles of drug-resistant tumors, which may help predict responses to drugs and assist in the design of tailored therapeutic regimens to overcome drug resistance.
Adaptor Protein Complex 1 ; Cisplatin* ; Cyclophilins ; DNA, Complementary* ; Drug Resistance ; Drug Therapy ; Dual-Specificity Phosphatases ; Fluorouracil* ; Gene Expression* ; Hepatocytes ; Humans ; Oligonucleotide Array Sequence Analysis* ; Receptor, Melanocortin, Type 1 ; Stomach Neoplasms* ; Transcriptome ; Tropomyosin

Sheldon-Hall syndrome (SHS) is a rare autosomal dominant, inherited arthrogryposis syndrome characterized by multiple congenital contractures of the distal limbs. To date, four genes that encode the skeletal muscle fiber complex have been confirmed as the causative genes. Mutations in MYH3 have been identified most frequently and few cases of SHS caused by TPM2 mutations have been reported worldwide. This report describes, for the first time, a Korean family with two generations of SHS resulting from a rare TPM2 mutation, p.R133W. The affected mother and daughter manifested typical facial features of SHS including a triangular face with downslanting palpebral fissures, small mouth, high arched palate, and prominent nasolabial folds, and showed camptodactyly of fingers and deformities of feet with congenital vertical tali. Generalized myopathy with relative sparing of the slow-twitch muscle fibers was also revealed by electromyography in the affected mother.
Alleles ; Arthrogryposis/*genetics ; Asian Continental Ancestry Group/*genetics ; Exons ; Female ; Finger Phalanges/radiography ; Foot Bones/radiography ; Humans ; Infant, Newborn ; Mutation ; Pedigree ; Phenotype ; Republic of Korea ; Sequence Analysis, DNA ; Tropomyosin/*genetics

OBJECTIVETo screen the hypertrophic scar related cytoskeletal genes during early postburn stage, so as to explore their roles in postburn scar contraction.

METHODScDNA microarray chips containing 4096 human cDNAs were employed to investigate the cytoskeletal gene expression of the scar samples from human postburn hypertrophic scar. Furthermore, the expression of one of the cytoskeletal genes in hypertrophic scar tissue was studied by in situ hybridization.

RESULTSThirteen up - regulated cytoskeletal genes in 3 early postburn hypertrophic scar samples were identified. Moreover, the cells expressing human tropomyosin TM30 mRNA, one of the up - regulated cytoskeletal genes, were found increased in the early postburn hypertrophic scar samples.

CONCLUSIONIn this study up - regulated expression of many hypertrophic scar related cytoskeletal genes was found in the scar samples during early postburn stage, and they might be important factors leading to postburn hypertrophic scar formation and contraction.

Adolescent ; Adult ; Burns ; genetics ; Child ; Child, Preschool ; Cicatrix, Hypertrophic ; genetics ; Cytoskeleton ; genetics ; Gene Expression Profiling ; Humans ; Male ; Oligonucleotide Array Sequence Analysis ; RNA, Messenger ; genetics ; metabolism ; Tropomyosin ; genetics ; Up-Regulation

Paramyosin is a myofibrillar protein present in helminth parasites and plays multifunctional roles in host-parasite interactions. In this study, we identified the gene encoding paramyosin of Clonorchis sinensis (CsPmy) and characterized biochemical and immunological properties of its recombinant protein. CsPmy showed a high level of sequence identity with paramyosin from other helminth parasites. Recombinant CsPmy (rCsPmy) expressed in bacteria had an approximate molecular weight of 100 kDa and bound both human collagen and complement 9. The protein was constitutively expressed in various developmental stages of the parasite. Imunofluorescence analysis revealed that CsPmy was mainly localized in the tegument, subtegumental muscles, and the muscle layer surrounding the intestine of the parasite. The rCsPmy showed high levels of positive reactions (74.6%, 56/75) against sera from patients with clonorchiasis. Immunization of experimental rats with rCsPmy evoked high levels of IgG production. These results collectively suggest that CsPmy is a multifunctional protein that not only contributes to the muscle layer structure but also to non-muscular functions in host-parasite interactions. Successful induction of host IgG production also suggests that CsPmy can be applied as a diagnostic antigen and/or vaccine candidate for clonorchiasis.
Amino Acid Sequence ; Animal Structures/chemistry ; Animals ; Antibodies, Helminth/blood ; Cloning, Molecular ; Clonorchis sinensis/chemistry/*genetics ; Collagen/metabolism ; Complement C9/metabolism ; Helminth Proteins/chemistry/*genetics/immunology/metabolism ; Immunoglobulin G/blood ; Molecular Sequence Data ; Molecular Weight ; Protein Binding ; Rats ; Rats, Sprague-Dawley ; Sequence Alignment ; Sequence Analysis, DNA ; Sequence Homology, Amino Acid ; Tropomyosin/chemistry/*genetics/immunology/metabolism