Cardiomyopathy, Dilated ; Cardiomyopathy, Restrictive/genetics* ; Humans ; Mutation ; Pedigree ; Tropomyosin/genetics*

Objective: To investigate the expression pattern of tropomyosin 2(TPM2) in aorta of patients with aortic dissection and explore its clinical implication. Methods: Thirteen cases with acute type A aortic dissection(TAAD) diagnosed by transabdominal aortic angiography from 2015 in Tongji Hospital were included. During the operation, the aortic wall tissues of these patients were collected. Ten patients with heart transplantation were selected as control group, and normal aortic wall tissues were taken. The hematoxylin-eosin (HE) and Verhoeff's Van Gieson (EVG) staining were performed to observe the morphological changes of aorta. The mRNA expression level of TPM2 was measured by real-time fluorescent quantitative-PCR, and the protein levels of TPM2 were detected by Western blot and immunohistochemical staining. Image The J software was used to collect the optical density values of each point on the image, obtain the integrated optical density(IOD) value, and calculate the average density(%, IOD/area of the target distribution area). Results: HE and EVG staining revealed medial degeneration and broken elastic fiber in aorta of TAAD patients. The mRNA expression levels of TPM2 were significantly upregulated in aorta of TAAD patients as compared to the control group (P<0.05), so as the TPM2 protein expression levels ((9.73±1.20)% vs. (0.11±0.04)%, P<0.05). And TPM2 was mainly expressed in cytoplasm. Conclusion: The increased expression of TPM2 in TAAD patients hints that TPM2 might be involved in the pathogenesis of aortic dissection.
Aneurysm, Dissecting/genetics* ; Aorta ; Aortic Aneurysm, Thoracic/genetics* ; Gene Expression ; Humans ; RNA, Messenger ; Tropomyosin/metabolism*

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

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

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 construct an eukaryotic expression vector of tumor associated gene tropomyosin 2 (TM2) tagged with green fluorescence protein and study its biological role in tumorigenesis. METHODS; Whole length of TM2 cDNA sequence was amplified from human gastric cancer cell line AGS by reverse transcript PCR. The recombinant TM2 was inserted into eukaryotic expression vector pIRES2 tagged with enhanced green fluorescence protein. pIRES2-EGFP was transfected into human hepatoma cell line QGY-7703 cells by liposome technique and detected by fluorescence microscopy and flow cytometry. TM2 protein level was determined by Western blot. The in vitro invasion and motility were further studied. Its tumorigenesis was assessed on nude mice.

RESULTSThe recombinant TM2 was shown to be expressed in QGY-7703 cells by fluorescence microscopy and flow cytometry. The protein level of TM2 was increased. The invasion ability of TM2- transfected cells was increased(45.6 +/- 9.9) than that in controls(21.6 +/- 3.3, P < 0.05), as well as mobility(41.4 +/- 11.8 vs. 16.7 +/- 3.7). The tumor volume was (241.5 +/- 95.1) mm3, significantly higher than that in blank-vector controls (100.6 +/- 85.4) mm3 and negative-controls (123.7 +/- 92.3) mm3.

CONCLUSIONTM2 plays a role in growth and metastasis of hepatic tumor.

Animals ; Cell Line, Tumor ; Cell Movement ; DNA, Complementary ; genetics ; Genetic Vectors ; Green Fluorescent Proteins ; metabolism ; Humans ; Liver Neoplasms ; metabolism ; pathology ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Invasiveness ; Neoplasm Transplantation ; Random Allocation ; Recombinant Fusion Proteins ; genetics ; metabolism ; Stomach Neoplasms ; genetics ; pathology ; Transfection ; Tropomyosin ; genetics ; metabolism ; physiology ; Tumor Burden

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