Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Reproductive toxicity is one of the potential side effects of anticancer alkylating agents, with potential effects on vimentin intermediate filaments, one of the main components of the Sertoli cytoskeleton. Research suggests (Aumuller et al, 1988; Aumuller et al, 1992) that the highly organized and active Sertoli cytoskeleton is important in spermatogenesis. The aim of the current study was to investigate the effects of alkylating agents on vimentin filament expression in vitro. Sertoli cells, isolated from 20-day-old mice testes, were cultured for 5 days and then incubated with 0, 50, 100, and 200 micromol/L nitrogen mustard (HN2). Morphologic changes in Sertoli cells were observed per 30-minute interval at 12-hour exposure time points to 100 micromol/L HN2. Vimentin expression was investigated by immunocytochemistry at 6 hours and 24 hours posttreatment and reverse transcriptase polymerase chain reaction and Western blot at 12 hours posttreatment with 50, 100, and 200 micromol/L HN2. Exposure to HN2 resulted in a comparatively small Sertoli cell body with diminished cytoplasm. Sertoli cells were shrunk or detached. Cytoskeletal disruption increased with increasing HN2 concentration. The optical density values of vimentin antibody and expression of vimentin mRNA and protein were significantly decreased with increasing concentration of HN2. Significant treatment dose-dependent and time-dependent differences of vimentin mRNA and protein expression levels were also noted. Our data suggest that the change in the biochemical properties of vimentin may indicate that one of the mechanisms of reproductive toxicity resulting from HN2 is disruption of Sertoli cell vimentin filament structure, accompanied by a down-regulation of vimentin expression.
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PMID:Cytoskeleton vimentin disruption of mouse sertoli cells injured by nitrogen mustard in vitro. 1719

Voltage-gated chloride channels (ClCs) are important mediators of cellular ion homeostasis and volume regulation. In an earlier study, we used immunohistochemical, Western blot, and reverse transcriptase PCR (RT-PCR) approaches to identify ClC-K variants in types II, IV, and V fibrocytes of the rodent spiral ligament. We have now confirmed the expression of ClC-K2 in these cells by in situ hybridization. All three of these fibrocyte subtypes are thought to be involved in cochlear K(+) recycling; thus, it is important to understand the precise mechanisms regulating their membrane conductance and the role played by ClCs in this process. In this study, we report the characterization of a secondary cell line derived from explants from the region of the rat spiral ligament underlying and inferior to the spiral prominence. The cultured cells were immunopositive for vimentin, Na,K/ATPase, Na,K,Cl-cotransporter, carbonic anhydrase isozyme II, and creatine kinase isozyme BB, but not for cytokeratins or Ca/ATPase, an immunostaining profile indicative of the type IV subtype. Evaluation of the cultures by RT-PCR and Western blot analysis confirmed the presence of both ClC-2 and -K2. Whole-cell patch clamp recordings identified two biophysically distinct Cl(-) currents in the cultured cells. One, an inwardly rectifying Cl(-) current activated by hyperpolarization or decreasing extracellular pH corresponded with the properties of ClC-2. The other, a weak outwardly rectifying Cl(-) current regulated by extracellular pH, Cl(-), and Ca(2+) resembled the channel characteristics of ClC-K2 when expressed in Xenopus oocytes. These findings suggest that at least two functionally different chloride channels are involved in regulating membrane anion conductance in cultured type IV spiral ligament fibrocytes.
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PMID:Identification of ClC-2 and CIC-K2 chloride channels in cultured rat type IV spiral ligament fibrocytes. 1733 50

Rat osteoblasts were cultured for 4 or 5 days aboard the Space Shuttle and solubilized during spaceflight. Post-flight analyses by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) determined the relative mRNA levels of matrix proteins, adhesion molecules, and cytoskeletal proteins including osteopontin (OP), osteonectin (ON), CD44, alpha-tubulin, actin, vimentin, fibronectin (FN), and beta1-integrin. The mRNA levels of OP and alpha-tubulin in the flight cultures were decreased by 30% and 50% on day 4 and day 5 of flight, as compared to the ground controls. In contrast, the CD44 mRNA levels in the flight cultures increased by 280% and 570% of the ground controls on day 4 and day 5. The mRNA levels of ON and FN in the flight cultures were slightly increased as compared to ground controls. The mRNA levels of actin, vimentin, or beta1-integrin did not change in spaceflight conditions. The matrix proteins, adhesion molecules, and cytoskeletal proteins may form dynamic network complexity with signaling molecules as an adaptive response to perturbation of mechanical stress under microgravity.
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PMID:Microgravity signal ensnarls cell adhesion, cytoskeleton, and matrix proteins of rat osteoblasts: osteopontin, CD44, osteonectin, and alpha-tubulin. 1738 75

The aim of this study is to investigate the ability of adult human bone marrow mesenchymal stem cells to differentiate towards a cardiomyogenic phenotype in vitro. Bone marrow samples have been aspirated from 30 patients undergoing open heart surgery. Mesenchymal stem cells were isolated and cultured in enriched medium. Second passaged cells were treated with 10 microM 5-azacytidine for 24 h. Selected surface antigens were analyzed by flow cytometry. Morphologic characteristics were analyzed by confocal and electron microscopy. Expression of cytoskeletal protein vimentin and muscle specific myosin heavy chain were analyzed by immunohistochemistry. Expression of alpha-cardiac actin, beta-myosin heavy chain and cardiac troponin-T was detected by reverse transcriptase polymerase chain reaction. Mesenchymal stem cells were spindle-shaped with irregular processes. Cells treated with 5-azacytidine have assumed a stick-like morphology. They were connecting with adjoining cells forming myotube-like structures. Numerous myofilaments were detected in induced cells running in a parallel fashion without forming sarcomeres that were immunohistochemically positive for myosin heavy chain and vimentin. The mRNAs of alpha-cardiac actin, beta-myosin heavy chain and troponin-T were expressed in both induced and uninduced cells. These results indicate that adult human bone marrow mesenchymal stem cells can differentiate towards a cardiomyogenic lineage in vitro.
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PMID:In vitro cardiomyogenic differentiation of adult human bone marrow mesenchymal stem cells. The role of 5-azacytidine. 1767 Jul 26

We report 20 cases of a distinct, previously unrecognized renal neoplasm, anaplastic sarcoma of the kidney with polyphenotypic features. The tumors were identified by re-reviewing tumors with unusual anaplastic features from the National Wilms Tumor Study Pathology Center, the International Society of Pediatric Oncology and the United Kingdom Children's Cancer Study Group trials. Patients ranged in age from 10 months to 41 years (median age 5 y, mean age 12 y) and females predominated (1.5:1). Twelve tumors presented in the right kidney, and 5 in the left (laterality was unknown in 3 cases). The most common presentation was a renal mass. Grossly, most tumors were large, measured 4 to 21 cm (mean 12.7 cm) and weighed 115 to 1820 g (mean 835 g). Seven out of 12 tumors suitable for assessment had a distinct cystic component. The tumors involved the pelvi-calyceal system in 5 of the cases. Histologically, all tumors showed a spindle cell component which contained either multiple foci or diffuse, widespread anaplastic changes with bizarre pleomorphic cells and very atypical mitotic figures. Chondroid differentiation was seen in 16 cases, usually in the form of islands of hyaline cartilage (13 cases) or chondroid matrix (3 cases). The nodules of cartilage showed both benign and malignant features, often within the same tumor. In 2 cases small foci of osteoid were found whereas osteoclast-like giant cells were seen in 4 cases. Only 3 of the tumors exhibited a primitive blastema-like area. No neoplastic epithelial structures were identified. No nephrogenic rests were found. Limited immunohistochemical studies showed vimentin positivity in 5/5 cases, desmin was positive in 4/6 cases, MYF4 showed focal weak nuclear positivity in 1/4 cases, but MyoD1 was negative in all cases (0/5). PGP9.5 was focally, strongly positive in 4/5 cases and p53 was strongly positive in 3/6 cases. Cytokeratin, using the antibody CAM5.2, was uniformly negative within the tumor cells. Finally, CD56 was focally positive in 1/6 tumors, whereas all other markers were negative including NB84a (4/4), CD34 (5/6), CD99 (5/5), and WT1 (6/6 cases). In 4 tumors reverse transcriptase-polymerase chain reaction was performed to detect the SYT-SSX fusion transcript produced by the t(x;18), and the ETV6-NTRK3 fusion transcript using RNA extracted from archived paraffin blocks-results were negative in all 4 specimens. Tumor stage was known in 15 patients including 7 stage I, 4 stage II, 3 stage III, and 1 stage IV tumors. They were usually diagnosed as anaplastic Wilms tumors and treated accordingly. Of the 13 patients with a minimum of 2 years follow-up, 4 patients developed distant metastases and 1 had local recurrence including 1 patient with stage IV, 2 with stage III, and 2 with stage I at presentation. Three of them died and 2 were lost to follow-up. One patient with stage I tumor developed widespread metastases and died. Another stage I patient developed local recurrence after 3 months of diagnosis, but was lost to follow-up. Five stage I patients were alive and free of tumor at last follow-up. The most common sites of metastases were lung (3 cases), and liver and bones (2 cases each). These tumors showed pathologic features similar to the pleuropulmonary blastoma of childhood and undifferentiated (embryonal) sarcoma of the liver. In the differential diagnosis, anaplastic Wilms tumor, primary renal synovial sarcoma, malignant mesenchymoma, ectomesenchymoma, and mesenchymal chondrosarcomas have been considered but none of these tumors shared the same features as the 20 cases described here which represent a distinct clinicopathologic entity with morphologic features of a polyphenotypic anaplastic sarcoma of the kidney. Further molecular studies are needed to better understand its nature and more accurate classification.
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PMID:Anaplastic sarcoma of the kidney: a clinicopathologic study of 20 cases of a new entity with polyphenotypic features. 1789 46

Snai2, encoded by the SNAI2 gene, has been shown to modulate epithelial-mesenchymal transformation (EMT), the conversion of sessile epithelial cells attached to adjacent cells and to the basement membrane into dissociated and motile fibroblastic cells. EMT occurs during development, wound healing, and carcinoma progression. Using Snai2-null mice (Snai2(lacZ)), we evaluated the role of Snai2 in UV radiation (UVR)-induced skin carcinogenesis. In chronically UVR-exposed nontumor-bearing skin from Snai2-null mice, inflammation and epidermal proliferation were decreased compared with wild-type (+/+) skin. Snai2-null mice had a consistently lower tumor burden than +/+ mice. In addition, null mice developed fewer aggressive spindle cell tumors, believed to arise from squamous cell carcinomas that have undergone EMT, than +/+ mice; however, the difference in tumor type distribution between the two genotypes was not statistically significant. No metastases were observed in either the +/+ or Snai2-null mice. Using quantitative reverse transcriptase-polymerase chain reaction and immunohistochemistry, we showed that the spindle cell tumors in the Snai2-null mice demonstrated impaired EMT, as shown by decreased vimentin and increased cadherin 1 expression. This study confirms a role for Snai2 in EMT, but demonstrates that Snai2 expression is not required for the development or progression of UVR-induced skin tumors.
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PMID:Snai2 expression enhances ultraviolet radiation-induced skin carcinogenesis. 1791 97

Sertoli cells are regulated by follicular stimulating hormone (FSH) and testosterone secreted by the pituitary gland and Leydig cells, respectively. However, the expression of the FSH receptor and androgen receptor were undetectable in both primary cultured Sertoli cells and Sertoli cell lines immortalized by SV40 large T antigen. Two Sertoli cell lines, B6Sc-2 and B6Sc-3, were established from the testis of 19-day-old C57BL/6 mice testis by immortalization with human telomere reverse transcriptase. These Sertoli cell lines expressed FSH receptors and the total phosphoprotein patterns were converted after FSH treatment. Additionally, immunological methods demonstrated that these cell lines expressed characteristic Sertoli cell proteins, such as tyrosine-tubulin, vimentin and stem cell factor (SCF). Reverse transcription-polymerase chain reaction (RT-PCR) also indicates that they express Sertoli specific mRNAs, such as Amh, claudin11 and ZO-1. The expression of the androgen receptor in both B6Sc-2 and B6Sc-3 cells could be induced by TNF-alpha treatment. The present results indicate that these Sertoli cell lines are more native than others and may thus provide useful tools for in vitro studies.
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PMID:FSH-sensitive murine sertoli cell lines immortalized by human telomerase gene hTERT express the androgen receptor in response to TNF-alpha stimulation. 1796 53

Idiopathic pulmonary fibrosis (IPF) comprises an aggregate of mesenchymal cells. However, the cellular origin of these mesenchymal phenotypes remains unclear. Transforming growth factor beta1 (TGF-beta1) has been known as the main cytokine involved in the pathogenesis of IPF. We examined whether the potent fibrogenic cytokine TGF-beta1 could induce the epithelial-to-mesenchymal transition (EMT) in the human alveolar epithelial cell line, A549, and determined whether snail expression is associated with the phenotypic changes observed in the A549 cells. EMT was investigated with cells morphology changes under phase-contrast microscopy, western blotting, and indirect immunofluorescence stains. E-cadherin and transcription factor, snail, were also evaluated by measuring mRNA levels using reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. The data showed that TGF-beta1 induced A549 cells with epithelial cell characteristics to undergo EMT in a concentration-dependent manner. Following TGF-beta1 treatment, A549 cells induced EMT characterized by cells morphological changes, loss of epithelial markers Ecaherin and cytokeratin, increased stress fiber reorganization by F-actin, and cytokeratin replacement by vimentin. Although IL-1beta failed to induce A549 cells to undergo EMT, the combination of TGF-beta1 and IL-1beta showed synergy effects in cells morphology changes and the expression of mesenchymal markers. The snail expression study using RT-PCR analysis provided that loss of E-cadherin expression was associated with snail expression. Stimulation of A54 cells with TGF-beta1 plus IL-1beta revealed a higher level of snail expression. Our data showed that EMT of A549 cells might be closely associated with snail expression.
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PMID:Transforming growth factor beta1 induces epithelial-to-mesenchymal transition of A549 cells. 1798 42

Two cases of primary prostatic synovial sarcoma presenting as a prostatic mass are presented in patients aged 44 and 46 years. Histologically, both tumors were mainly composed of uniform spindle cells forming interlacing fascicles. Clusters of immature epithelioid cells were also observed among the spindle cells in case 1. Immunohistochemically, the tumor cells of both cases were strongly positive for vimentin, bcl-2, CD99, and E-cadherin, as well as focally positive for cytokeratin. However, they were negative for prostate-specific antigen, S-100 protein, CD34, CD117, muscle-specific actin, desmin, and calretinin. The presence of an SYT-SSX gene fusion resulting from t(X;18) was demonstrated from paraffin blocks by reverse transcriptase polymerase chain reaction in both cases. To the authors' knowledge, these represent the fifth and sixth reported cases of prostatic synovial sarcoma. Accurate diagnosis depends on morphologic and immunohistochemical examination and proper molecular analysis.
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PMID:Primary synovial sarcoma of the prostate: report of 2 cases and literature review. 1838 92

A progressive tubulointerstitial nephropathy is mainly induced by aristolochic acid I (AAI), but a comprehensive understanding of this process is still missing. By using mouse primary renal tubular epithelial cells (RTECs) cultured in vitro and combining with two AAI treatment types (dose-response studies and time-response studies), we sought to investigate the nephrotoxicity of AAI further. Following our molecular and pharmacological studies, we found that high doses of AAI could lead to the death of RTECs within a short time, but low doses in a long duration only induce the epithelial cells to transform into myofibroblasts (MFs). This was also immediately identified by the increased expression of vimentin and de novo expression of alpha-smooth muscle actin (alpha-SMA) with the loss of cytokeratin 18 (CK18) by semiquantitative reverse transcriptase-PCR (RT-PCR) and immunofluorescence staining. The transcriptional level of transforming growth factor-beta1 (TGF-beta1) in the group treated with AAI significantly increased twice as much as the control. Smad2 mRNA level in the group with 50 ng/mL AAI declined by 23.4% at 24 hr, then increased by 180.0% at 36 hr; it was also evidently increased (217.4%) after being treated with 30 ng/mL AAI for 24 hr. Meanwhile, Smad7 mRNA level was down-regulated by AAI in dose- and time-dependence. Furthermore, by cotransfecting in mouse primary RTECs, the transcriptional level of Smad7 promoter-luciferase reporter gene was significantly down-regulated by AAI (300 ng/mL), and the expression of myofibroblast-specific markers induced by AAI was also suppressed by the specific antagonist of TGF-beta1 receptors (SB-431542). Collectively, the present results suggest that AAI may induce cytotoxicity through its conductive epithelial to mesenchymal transition, and TGF-beta1/Smad7 signaling can stimulate renal tubulointerstitial fibrosis induced by AAI.
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PMID:TGF-beta1/Smad7 signaling stimulates renal tubulointerstitial fibrosis induced by AAI. 1870 12


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