Role of FOXM1 in collagen synthesis in MRC-5 cells induced by high glucose / 中国药理学通报

Aim To explore the role of transcription factor F0XM1 in collagen synthesis in MRC-5 cells in-duced by high glucose.Methods I ▪ The optimal time and concentration of the hyperglycemia model in MRC-5 cells were explored by CCK8: the time gradi¬ents: 6,12,24,48,72 h; the concentration gradients: 5.5,15 ,30,45 mmol • L 1 , and 30 mmol • L 1 man- nitol was used to be the hypertonic control group.(2) Hie effect of collagen synthesis in MRC-5 cells induced by high glucose was detected: the cells were divided into normal control group, hypertonic control group (30 mmol • L 1 mannitol) and high glucose (30 mmol • L 1 ) group.WB and qPCR assays were used to de¬tect the expression of conllagen synthesis factors ( Fn, COL 1, COL m, a-SMA, MMP9, TIMP1 ) and TGF-p signaling pathway factors (TGF-pi , p-Smad2/ Smad2).(3 The role of FOXMl in promoting collagen synthesis by high glucose was investigated: the cells were divided into normal control group, hypertonic control group (30 mmol • L 1 mannitol) , high glucose ( 30 mmol • L 1 ) group and high glucose (30 mmol • L 1 ) + thiostrepton ( 1 (xmol) group, and the expres¬sions of FOXM1 , collagen synthesis factors were detec¬ted by WB and qPCR assays.Results Mannitol had no significant effect on proliferation of MRC-5 cells, hut their proliferation activity was significantly lower than that of control group when MRC-5 cells were trea¬ted with 30 mmol • L 1 high glucose for 24 h; the ex¬pressions of COL 1 , COL IH , F0XM1 and other fac¬tors were promoted when MRC5 cells were treated with high glucose; the expression of F0XM1 was signifi¬cantly inhibited after the addition of thiostrepton, and the expressions of collagen synthesis factors also de-creased compared with high glucose group, and the a- bove differences were all statistically significant (P < 0.05).Conclusion FOXM1 is a factor related to the increase of collagen synthesis in MRC-5 cells induced by high glucose.

Interplay between TGF-b1 and miRNA-122 biomarkers in hepatocellular carcinoma progression in patients with chronic hepatitis C

@#Hepatocellular carcinoma (HCC) is a highly lethal malignancy and clinically validated medications have not yet been developed since there are no reliable diagnostic and prognostic biomarkers. Based on bioinformatics tools, TGF-b1 gene was the first target gene of miRNA-122, therefore this study was intended to assess the potential interconnection between TGF-b1 and miRNA-122 as a diagnostic and prognostic biomarker in the progression of HCC in patients with chronic hepatitis C (CHC) genotype (4). In this study, 100 people were included and split into two groups; group I: CHC patients without HCC that were classified into patients CHC without cirrhosis and CHC cirrhotic patients, group II: CHC patients with HCC, and healthy volunteers as control. The expression of miRNA-122 and TGF-b1 genes were analyzed using Real-Time PCR. An upregulation of miRNA-122 gene in cirrhotic and HCC patients compared to both chronic HCV non-cirrhotic, and cirrhotic patients, while, a decrease in expression of TGF-b1 was found in cirrhotic patients compared to HCV non-cirrhotic patients. Although significantly downregulated in HCC patients. Regression analysis indicated that the expression levels of miRNA-122 and TGF-b1 could be regarded as important indicators of the alterations in cirrhotic and HCC patients versus HCV non-cirrhotic patients, also with the chances of HCC versus cirrhosis patients. Our data indicated an interaction between miRNA-122 and TGF-b1, regulated gene expression and recommended the use of these parameters as noninvasive predictive biomarkers and therapeutic targets for HCV induced liver cirrhosis and HCC.

Involvement of eIF2α in halofuginone-driven inhibition of TGF-β1-induced EMT

Halofuginone (HF) is an extract from the widely used traditional Chinese medicine (TCM) Dichroa febrifugathat facilitates the recovery of wounds and attenuates hepatic fibrosis. However, the role of HF in theepithelial-mesenchymal transition (EMT) of IPEC-J2 cells remains unclear. The current study explored theanti-EMT effect of HF in IPEC-J2 cells and illustrates its molecular mechanism. Transforming growth factorb1 (TGF-b1), as a recognized profibrogenic cytokine, decreased the level of the epithelial marker E-cadherinand increased the level of the mesenchymal markers, such as N-cadherin, fibronectin (FN), vimentin (Vim),and a-smooth muscle actin (a-SMA), in IPEC-J2 cells depending on the exposure time and dose. HF markedlyprevented the EMT induced by TGF-b1. Dissection of the mechanism revealed that HF inhibited IPEC-J2 cellEMT via modulating the phosphorylation of SMAD2/3 and the SMAD2/3-SMAD4 complexnuclear translocation. Furthermore, HF could promote the phosphorylation of eukaryotic translation initiationfactor-2a (eIF2a), which modulates the SMAD signaling pathway. These results suggested that HF inhibitsTGF-b1-induced EMT in IPEC-J2 cells through the eIF2a/SMAD signaling pathway. Our findings suggest thatHF can serve as a potential anti-EMT agent in intestinal fibrosis therapy.

Let-7a-5p represses proliferation, migration, invasion and epithelial-mesenchymal transition by targeting Smad2 in TGF-β2-induced human lens epithelial cells

Transforming growth factor b2 (TGF-b2)/Smad signaling is widely accepted as a key inducer of proliferationand epithelial-mesenchymal transition (EMT) of human lens epithelial cells (LECs), contributing to thedevelopment of posterior capsule opacification (PCO). Increasing evidence shows that microRNAs (miRNAs)play important roles in PCO pathogenesis. Herein, we aimed to explore the role and molecular mechanism oflet-7a-5p on TGF-b2-induced proliferation and EMT in LECs. qRT-PCR was performed to detect theexpression of let-7a-5p and Smad2 mRNA. Western blot was used to determine the Smad2 level and theinduction of EMT. The targeted correlation between let-7a-5p and Smad2 was confirmed using dual-luciferasereporter and RNA immunoprecipitation assays. CCK-8 assay was employed to determine cell proliferation, andtranswell assays were performed to assess cell migration and invasion. We found that TGF-b2 induced EMT ofLECs, and TGF-b2 upregulated Smad2 expression and reduced let-7a-5p expression in LECs. Smad2 was adirect target of let-7a-5p. Moreover, let-7a-5p upregulation repressed proliferation, migration, invasion andEMT in TGF-b2-induced LECs. But, Smad2 expression restoration abrogated the inhibitory effect of let-7a-5pupregulation. In conclusion, our data indicated that let-7a-5p upregulation repressed TGF-b2-induced proliferation, migration, invasion and EMT at least partly by targeting Smad2 in LECs, highlighting that let-7a-5pmight act as a promising therapeutic target to intervene to the progression of PCO.

Progressive diaphyseal dysplasia with right transverse sinus stenosis: a case report / 中华神经科杂志

Progressive diaphyseal dysplasia , also called Camurati-Engelmann disease (CED), is a kind of autosomal dominant disease mainly involved in long diaphysis , characterized as progressive and symmetric cortical thickening as well as low prevalence.The pathogenic gene of the disease is TGFB 1 located in 19q13, resulting in abnormal bone metabolism.We report a young woman suffering from CED , mainly presented as decreased visual acuity , chronic high intracranial pressure and skull damages by cerebral angiography.The digital subtraction angiography revealed the right transverse sinus stenosis .We first revealed a CED patient with chronic high intracranial pressure caused by sinus stenosis , and selected intravascular therapy for the sinus stenosis.

Polymorphisms in the TGFB1 and IL2RA genes are associated with clinical forms of leprosy in Brazilian population

BACKGROUND Leprosy is a chronic infectious disease caused by Mycobacterium leprae, and compromises the skin and peripheral nerves. This disease has been classified as multibacillary (MB) or paucibacillary (PB) depending on the host immune response. Genetic epidemiology studies in leprosy have shown the influence of human genetic components on the disease outcomes. OBJECTIVES We conducted an association study for IL2RA and TGFB1 genes with clinical forms of leprosy based on two case-control samples. These genes encode important molecules for the immunosuppressive activity of Treg cells and present differential expressions according to the clinical forms of leprosy. Furthermore, IL2RA is a positional candidate gene because it is located near the 10p13 chromosome region, presenting a linkage peak for PB leprosy. METHODS A total of 885 leprosy cases were included in the study; 406 cases from Rondonópolis County (start population), a hyperendemic region for leprosy in Brazil, and 479 cases from São Paulo state (replication population), which has lower epidemiological indexes for the disease. We tested 11 polymorphisms in the IL2RA gene and the missense variant rs1800470 in the TGFB1 gene. FINDINGS The AA genotype of rs2386841 in IL2RA was associated with the PB form in the start population. The AA genotype of rs1800470 in TGFB1 was associated with the MB form in the start population, and this association was confirmed for the replication population. MAIN CONCLUSIONS We demonstrated, for the first time, an association data with the PB form for a gene located on chromosome 10. In addition, we reported the association of TGFB1 gene with the MB form. Our results place these genes as candidates for validation and replication studies in leprosy polarisation.

Platelet-derived growth factor A mRNA in platelets is associated with the degree of hepatic fibrosis in chronic hepatitis C

Abstract: INTRODUCTION: Transforming growth factor beta 1 (TGFB1) and platelet-derived growth factor (PDGF) are the main cytokines related to hepatic fibrogenesis. METHODS: RNA isolated from the platelets and hepatic tissue of 43 HCV carriers was used for quantitative polymerase chain reaction to determine TGFB1, PDGFA, and PDGFB RNA expression. RESULTS: The mRNA expression of PDGFA in platelets was significantly lower in the group with advanced fibrosis than in the group with early-stage fibrosis. TGFB1 was more frequently expressed in platelets than in hepatic tissue, which was different from PDGFB. CONCLUSIONS: A pathway mediated by overexpression of TGFB1 via PDGFA in megakaryocytes could be involved in the development of fibrosis.

Relationship between the Change of IGF 1 and TGF-β1 Level and Renal Damage in Children with Allergic Purpura / 现代检验医学杂志

Objective To investigate the effects of insulin-like growth factor 1 (IGF 1) and transforming growth factor beta 1 (TGF-β1) in he pathogenesis of children with allergic purpura kidney damage.Methods 135 henoch-schonlein purpura (HSP) children with kidney damage were divided into HSP and HSPN group according to whether associated with renal damage,blood IGF 1,TGF-β1,urinary inhibition C (Cys C),creatinine (SCr) and content of urea nitrogen (BUN) were compared,and blood IGF-1,TGF-β1 and Cys C content of HSPN patients in different pathological grading were compared,the correlation of blood IGF 1,TGF-β1 and Cys C content of the HSPN group were analyzed,Results Blood IGF 1,TGF-β1 and Cys C content of the control group(117.2±18.8 ng/L,164.2±18.4 ng/L,0.9±0.2 mg/L),the HSP group(131.7±19.6 ng/L,282.1±28.3 ng/L,1.1±0.2 mg/L) and the HSPN group (205.3±24.5 ng/L,489.2±32.7 ng/L,1.3±0.3 mg/L) showed a trend of increasing gradually (F=4.824～45.066,P value＜0.01),the HSP group and the HSPN group were higher than that of the control group (q=3.397～58.931,P value＜0.01),the HSPN group was higher than that of the HSP group (q=16.997,35.193,P value＜0.01),the difference was statistically significant.Blood IGF-1 (level Ⅱ 175.6 ± 20.4 ng/L,level m198.5±23.3 ng/L,level Ⅳ241.7±25.1 ng/L),TGF-β1(level Ⅱ 392.8±38.9 ng/L,level Ⅲ 481.3± 44.03 ng/L,level Ⅳ 537.6±42.9 ng/L),Cys C (level 11 1.1±0.3 mg/L,level Ⅲ 1.3±0.4 mg/L,level Ⅳ1.6±0.4 mg/L) content of children with HSPN increased with the increase of renal pathology classification (F=6.594～ 28.317,P value ＜0.01),blood IGF-1,TGF-β1 and Cys C content of kidney pathology classification of Ⅵ level in children was higher than that of the level of Ⅱ and Ⅲ in children (q=2.415～11.818,P＜0.05 or P＜0.01),while the contern of blood IGF-1,TGF-β1 and Cys C of level Ⅲ in children was higher than that of the level Ⅱ in children (q=2.577～6.244,P＜0.05 orP＜ 0.01),the difference was statistically significant.Blood IGF-1,TGF-β1 content of children with HSPN were positively correlated with Cys C content of children (r=0.648,0.719,P＜0.05),but blood IGF 1 content was significantly positive correlated with TGF-β1 content (r=0.748,P＜0.05).Conclusion IGF 1 and TGF-β31 partieipated in the pathogenesis of HSPN,and both were correlated with the degree of the pathological damage.

Involvement of transforming growth factor beta-1 (TGFβ1) cytokine and FOXP3 transcription factor genetic polymorphisms in hematological malignancies

Hematological malignancies (HM) are a group of neoplastic diseases that arise from hematologic cell lineages. Transforming growth factor beta 1 (TGFβ1) is shown to negatively regulate normal and malignant hematopoiesis and, in immunological context, to promote T cell exhaustion and generation of regulatory T cells, which are shown to be deleterious in cancer, by the induction of transcription factor FOXP3 expression. The present study aimed to evaluate TGFB1 exon-1 rs1800470 and FOXP3 intron-1 rs2232365 polymorphisms in relation to HM susceptibility. DNA was extracted from blood samples of 43 HM patients and 142 neoplasia-free individuals and polymorphisms were analyzed by allelic-specific PCR. Association analysis was assessed by the Odds Ratio (OR) with significance level of 5%. Regarding FOXP3 polymorphism, no significant differences were observed in genotype or allele distribution among the patients and controls. However, there was a positive association between TGFB1 TT genotype and HM susceptibility (OR = 4.07; CI95% = 1.94 - 8.52). In the combined analysis, a positive association was also observed for TGFB1 TT and FOXP3 GG genotypes (OR = 4.00; CI95% = 1.54 - 10.41) in relation to HM susceptibility. Our results indicated promising new markers to be further investigated in hematological malignancies.

Results of the bone healing performance of Osteo-Calcium 5 preparation tested by internal fixation surgery patients / Монголын Анагаах Ухаан

INTRODUCTION: Nowadays, the population of noncommunicable sickness is growing rapidly changing lifestyle is one example of this is the widening of accidents-related bone fractures.The National Center for Trauma Treatment of inpatient hospital from a total fractures 10260 or 30.8%, of which 6777 or 66.05% of the marrow bone fractures as well as drugs used in refractive rarely occupied. PURPOSE: To consider a result of the use of Osteocalcium 5 medical preparation for patients who were diagnosed diaphyseal fracture of the tibia and treated by Intramedullar Osteosyntesis (internal fixation)OBJECTIVES:1. To establish process of fractured bone remodeling to original bone contour by TGF-b1 cytokine dynamics in serum2. To establish process of fractured bone remodeling to original bone contour by alkaline phosphatase dynamics in serumMATERIALS AND METHODS: Within the framework of the survey 28 patients of the “Bone, joint and limb section” Section of the National Institute of Traumatology, aged between 25-60 years, were involved in the present survey. Survey participants were classified in 2 groups according to “Krischner wire” position and their Xray’s images. An accordance to the methodology of James Balch and Mark Stengller (2011) daily dosage of the Osteocalcium 5 preparation for test and comparing patients was determined as following introduced: for test group-twice a day before meal 1.5 grams, for comparing group–Calcium D3Nycomed preparation three times a day during 60 days. Transforming growth factor-beta 1(TGFb1) and Alkaline phosphatase was equal of 3, 14, 42 fraction days.RESULT: The present diagram shows that according to the curve TGF-b1 the bone excrescence performance of the Osteocalcium 5 preparation (0.0001) is statically increasing at 14 days and after 42 days it is statically decreasing and the calculated discrepancy (p<0.015) reveals and proofs the impact of its bone excrescence. Alkaline phosphatase in serum 14 days after the fracture Osteocalcium 5 pills used in chapter Nycomed Calcium D3 drank chapters (116,06±35U/L) compared to a statistical difference (p<0.044).CONCLUSION: Preparations Ostyeokalitsi 5 osteoblast chondrocyte cell activity and enable them synthesized by the cells in the TGF-B1 osteoblast line Alkaline phosphatase synthesized in increased serum concentrations of the bone heal affect signaling molecules involved are determined that quickens recovery.

Avaliação de sete protocolos para obtenção de plasma rico em plaquetas na espécie equina / Evaluation of seven platelet-rich plasma processing protocols in the equine species

O presente estudo teve por objetivo avaliar a capacidade de concentração plaquetária e sua correlação com os níveis do fator de crescimento TGF-B1, a presença de leucócitos e de hemácias nos diferentes protocolos utilizados na obtenção do plasma rico em plaquetas (PRP) de equinos, através do método manual. Dez equinos, sadios, com idade média de 7 anos (±2,39), pesando em média 500kg (±67,1) foram utilizados neste estudo. Os protocolos testados variaram na velocidade e no tempo nas duas centrifugações. As variáveis analisadas nas amostras de PRP foram: concentração de plaquetas, presença de leucócitos e hemácias, e níveis de TGF-β1 quantificados pelo teste ELISA. Os protocolos testados não diferiram na capacidade de concentração de plaquetas e nos níveis de TGF-β1. Entretanto, houve diferença significava entre o protocolo I e os demais por este apresentar maior número de hemácias e leucócitos nas amostras de PRP, sendo por esse motivo considerado um protocolo inadequado para processamento do volume de sangue utilizado. Os demais protocolos podem ser utilizados para obtenção de PRP terapêutico em equinos.

PRP is plasma that contains a high numbers of platelets and growth factors in a small volume. The aim of this study was to evaluate seven different protocols to obtain PRP by the manual method according to their capacity to concentrate platelets, leukocyte and erythrocyte contamination and correlation between platelet count and TGF-β1 growth factor levels in PRP samples. Ten healthy horses with a mean age of 7 years (±2.39), weighing on average 500 kg (±67.1) were used in this study. The protocols tested varied according to the speed and time used at the two centrifugations. PRP samples were analyzed regarding platelet concentration, leukocyte and erythrocyte contamination and TGF-β1 levels quantified by ELISA. No significant differences among protocols were observed regarding the ability to concentrate platelets and TGF-β1. However, protocol I showed significantly higher erythrocyte and leukocyte counts in PRP samples than the other protocols, reason why it was considered inadequate for the volume of blood processed in this experiment. The remaining protocols are suitable for extracting PRP.

Immunolocalization of bmp2/4, tgfβ-1 and osteonectin in fibro-osseous lesions of the jaws / Imunolocalização de bmp2/4, tgfß-1 e osteonectina em lesões-fibro-óssea benignas

Objetivo: as lesões fibro-ósseas benignas (LFOB) correspondem a um grupo diverso de patologias caracterizadas pela substituição do tecido ósseo por tecido conjuntivo e matriz extracelular mineralizada. Pouco se conhece a respeito da etiologia desse grupo de lesões. Propomo-nos a analisar por meio da técnica imunohistoquímica a expressão de 3 moléculas (osteonectina, TGFβ-1 e BMP 2/4) envolvidas no metabolismo ósseo. Métodos: Trinta e dois casos diagnosticados como osso normal (ON,8), displasia fibrosa (DF,8), displasia cemento-óssea (DCO,8) e fibroma cemento-ossificante (FCO,8) foram selecionados. Resultados: A osteonectina e a BMP2/4 foram positivas em todos os casos. O TGFβ-1 revelou positividade em 1 caso de DCO e FCO. Conclusão: Os achados imunohistoquímicos sugerem que as LFOB tem processos diferentes de produção de tecido ósseo.

Background: Benign fibro-osseous lesions (BFOL) comprise a diverse group of pathologies characterized by the replacement of normal bone by fibrous tissue and a mineralized product. Little is known about the biology of this group of lesions. We have analyzed the immunohistochemical expression of three molecules involved in bone metabolism, namely osteonectin, TGF-b1, and BMP2/4. Methods: Thirty-two cases diagnosed as normal jaw bone (NJB, 8 cases), fibrous dysplasia (FD, 8 cases), cemento-osseous dysplasia (COD, 8 cases), and cemento-ossifying fibroma (COF, 8 cases) were selected. Results: Osteonectin and BMP2/4 antibodies were positive in all cases. TGFb-1 labeling was seen in one case of COD and COF. Conclusion: The immunohistochemistry findings suggest that BFOL have different processes of osseous tissue production.

Transforming growth factor-β-1 polymorphisms are infrequent but exist at selected loci in oral submucous fibrosis.

Background : Oral submucous fibrosis (OSF) may be considered a collagen metabolic disorder resulting from areca-nut alkaloid exposure and individual variation in collagen metabolism. Due to the complexity of OSF pathogenesis, it is important to elucidate independent and interactive effects of polymorphisms of collagen-related genes on OSF risk. Materials and Methods : This study is focused on seven polymorphisms (SNPs) of transforming growth factor-beta-1 (TGF-beta-1) gene in patients with oral submucous fibrosis (OSF), belonging to south Indian ethnic extraction. The mean age at presentation was 43.9 years, range 23-72 years (n=50, M:F ratio, 2.6:1). DNA samples from 50 subjects of the same ethnic group and comparable demographic features who have had practiced the habit of areca-chewing of almost equal duration, but remained free of disease constituted the controls. All DNA samples were collected progressively and purified from peripheral blood employing standard protocols and tested for SNPs. They included two polymorphisms in the promoter region (C-509T and G-800A), three polymorphisms in exon-1 (Arg25Pro(G915C), Leu10Pro(T869C), Glu47Gly(A979G) and two in 5 ͲUTR regions (C→T(rs13306708) and G→A (rs9282871). The extracted DNA samples along with the primers underwent PCR amplification and the genotypic and allelic frequencies were calculated. All calculations were performed using the SPSS software. The PCR products were purified and subsequently sequenced using Flour S™ multi-imager system (Biorad). The sequenced data were analyzed using the BioEdit sequence analysis software. Results : Out of the seven polymorphisms analyzed, six such as two in the promoter region, three in exon-1 and one in 5¢UTR were found to have a " P" value above 0.05 and hence were not significant. The C→T transition (rs13306708) in the 5¢UTR region recorded a " P" value of 0.03 on comparison and hence was found to be significant. The allelic frequencies for this C→T transition in patients were 68.7% C and 31.2% T (27CC, 15CT, 8TT) and that in controls were 89.5% C and 10.4% T (42CC, 6CT, 2TT). Conclusions : The polymorphism in 5¢UTR C-T in TGF beta 1 gene has a significant association with OSF, being a prime determinant in the pro-angiogenic pathway which has got direct bearing with the pathophysiology of the disease. The proximity of this polymorphism to the transcription site and the associated risk involved is discussed.

Novel genetic marker in TGFB2 gene associated with expression of TGF-?2 in leukocyte and SLE susceptibility

Background: TGF-β2 has a role in immune regulation, and genetic variance within the gene might contribute to systemic lupus erythematosus (SLE) pathogenesis. The TGFB2 gene is one candidate gene within the major SLE genetic susceptibility loci. Objective: Investigate the TGFB2 gene located on chromosome 1q41 as a SLE susceptibility gene. Materials and methods: One hundred fifty three SLE patients and 133 healthy controls participated in this study. Four markers selected in two haplotype blocks that have a minor allele frequency greater than 5% in Thai population were genotyped and analyzed in the association study. Results: There was no significant association between SLE susceptibility and the polymorphism in the promoter area (+67_68insACAA) and +89835 (A/G) at the intron 5 of TGFB2 gene. Instead, minor allele of the two new genetic markers at the intron 1 (+720) (corrected p-value = 0.024, OR = 0.4141, 95%CI = 0.22-0.80) and intron 6 (+94399_94400) (corrected p-value = 0.000143, OR = 0.3367, 95%CI = 0.20-0.58) were independently associated as a protective factor to SLE. Additionally, the real time RT-PCR results showed that patients with the protective allele (minor allele) at the +94399_94400 position have higher TGF-β2 mRNA level in leukocytes than patients with the risk allele (p=0.011). Conclusion: Two new genetic markers at intron1 (+720) and intron 6 (+94399_94400) were independently associated with SLE. The observed results have to be confirmed in other populations with a large sample size.

Mesenchymanl Stem Cell Based Intradiscal Gene Therapy: Therapeutic Implication in Degenerative Disc Disease / 대한척추외과학회지

STUDY DESIGN: In-vitro experiments using human mesenchymal stem cells (MSCs), intervertebral disc (IVD) cells and type 5 adenovirus/transforming growth factor-beta1 construct (Ad/TGF-beta1). OBJECTIVES: To determine the effect of MSC-based gene therapy for matrix regeneration of IVD cells. SUMMARY OF LITERATURE REVIEW: MSCs are known to be multipotent in tissue regeneration. In degeneration of IVD, cellular replacement with genetic modification other than that of IVD cells may prove an enhanced mechanism for the regeneration of MATERIALS AND METHODS: MSCs and IVD cells were cultured and an adenovirus construct containing TGF-beta1 cDNA (Ad/TGF-beta1) was also produced. In the first step, the MSCs were transduced with Ad/TGF-beta1, then mixed with IVD cells in various proportions and three dimensionally cultured. [methyl-(3)H]Thymidine and [(35)S]Sulfur incorporation for DNA and proteoglycan synthesis, respectively, were measured. RT-PCR was performed to assess the aggrecan and collagen types I and II mRNA RESULTS: Mixed cultures of MSC and IVD cells showed relatively similar amounts of newly synthesized proteoglycan compared with cultures of IVD cells only. In mixed cultures transduced with Ad/TGF-beta1, there were significant decreases in newly synthesized proteoglycan with increasing the proportions of MSCs, which was also found with the aggrecan and collagen type II mRNA expressions. However, the collagen type I mRNA expression increased with increased proportions of MSCs transduced with Ad/TGF-beta1. CONCLUSION: Cell therapy with MSCs and IVD cells provided a mechanism for cellular augmentation. However, MSC-based gene therapy coupled with IVD cells did not maintain a chondrogenic phenotype.

Double Plate Occipitocervical Fusion After Failed Posterior Fusion C 1-2 With Wiring / 대한척추외과학회지

A 31-year-old female complained of neck pain and limitation in neck motion. She had a 3 month history of treatment with Halovest at another hospital for a fracture of the odontoid process due to a car accident. The patient complained of persistent pain and limitation in neck motion following the cessation of Halovest. A dynamic radiograph demonstrated instability on C1-2 and she underwent a posterior cervical fusion with wiring. A wound infection developed, and loosening of the wire and lysis of the posterior arch at C1-2 were seen on a follow up plain radiograph 2 months postoperatively. She was transferred to our hospital where she underwent occipitocervical fusion with a double plate after control of the infection. There were rigid fixations of the plate and bone union on a follow up radiograph 24 months postoperatively.

Change of the Effect of TGF-beta1 on Physeal Chondrocytes According to Culture Methods in Vitro / 대한정형외과학회잡지

PURPOSE: We intended to check the growth rates and phenotypic markers of chondrocytes in the dedifferentiated cells cultivated in various conditions in order to establish the ideal culture system for implantation. MATERIALS AND METHODS: Culturing rabbit chondrocytes from proximal tibia, we checked the phenotypes at first, second, and third week. Then we cultured the chondrocytes in different circumstances such as monolayer or three dimensional gel in the presence or abscence of TGF-B1, and checked the growth rates and phenotypic markers. RESULTS: There was no difference in growth rates and mRNA level of type I, type II collagen and aggrecan between the cells cultured in monolayer and three dimensional gel of collagen. However, the responses of the cells to TGF-B1, were quite different between these two groups. In monolayer culture, the expression of type I collagen was depressed by TGF-B1 while the growth rate was markedly increased. Oppositely in three dimensional culture, the mRNA level of type I collagen was markedly increased and the growth rate was completely suppressed by TGF-B1. The expression of type II collagen could be detected only in TGF-B1-treated cells cultured in three dimensional gel for 4 or more days. The mRNA level of aggrecan was also increased by TGF-B1, in the cells cultured in three dimensional gel. CONCLUSIONS: These results suggest that the number of chondrocytes can be efficiently expanded by culturing the cells in monolayer and the phenotypes of chondrocyte can be restored by culturing the cells in three dimensional gel containing TGF-B1. The application of semi-solid gel containing differentiated chondrocytes in physeal implantation should be further evaluated

GEnetic Change in Transforming Growth Factor-B (TGF-B) Receptor Type I and Type II Genes with Resistance to TGF-B of Human Breast Cancer Cells / 대한암학회지

PURPOSE: Transforming growth factor-Bs (TGF-Bs) are prototypic multifunctional negative growth factors that inhibit the growth of many cell types. TGF-B type I and II receptors(RI, RII) are transmembrane receptors containing cytoplasmic serine/ threonine kinase domain and have been implicated in mediating TGF-B activity. Because a heteromeric complex of RI and RII is required for TGF-B signal transduction, cancer cells may reduce the expression of either RI or RII to escape from growth inhibition of TGF-B. We examined the correlation between the growth inhibitory activity of TGF-B1 and the genetic expression of RI &RII genes in human breast cancer cell lines. MATERIALS AND METHODS: We examined the growth inhibitory activity of TGF-B1 in 5 breast cancer cell lines by incorporation of [3H] thymidine. To investigate the correlation between TGF-B1 insensitivity and genetic change of TGF-B receptor genes (RI, RII), Southem blot analysis, Northern blot analysis, and Western blot analysis were performed. We also examined whether microsatellite instability(RER) was associated with RII mutation. RESULTS: We found that 3 breast cancer cell lines (MCF-7, YCC-B101, YCC-B151) were resistant to growth inhibitory effect of TGF-B1. MCF-7 cell line expressed no detectable RII mRNA and RII protein, but showed normal structure of RII gene and normal expression of RI gene. And we did not find any abnormal expression of mRNA, protein, and genetic structure of RI &RII in YCC-B101 and YCC-B151. CONCLUSION: Our results suggest that aquired resistance to the growth inhibitory effect of TGF-B1> could be transcription regulation system of RII in MCF-7 cell line, and could be postreceptor signal transduction pathway in YCC-B101 and YCC-B151 cell lines.

Study on Growth Suppression Effect of Vetamin D3 Mediated by Transfrorming Growth Factor-B1(TGF-B1) in Acute Myelogenous Leukemic Cell / 대한암학회지

PURPOSE: Vitamin D3 was shown to arrest the growth of acute myelogenous leukemic cells and transforming growth factor- B1 (TGF- B1) was reported to be involved in the mechanism of vitamin D3. We studied the growth inhibitory effect of 1,25(OH)2-vitamin D3(C) and its analogue (EB1089) in leukemic cell lines and the changes in the secretion or the activation of TGF-B1 in the supernatant and the status of TGF-B1 type II receptor. MATERIALS AND METHODS: Growth inhibition by vitamin D3 and TGF-B1 in 5 leukemic cell lines (HEL, HL-60, U937, KG-1, K562) were assessed with clonogenic and [3H]thymidine assay respectively. TGF-B type II receptor status was examined by Southern and Northern blotting. The concentrations of TGF- B1 in the supernatant were quantitated by enzyme immunoassay. RESULTS: The growth of HEL, HL-60, U937 were inhibited in a dose-dependent fashion by both C and EB1089, more markedly by the latter. Anti-TGF-B neutralizing antibody partially reversed the growth inhibition. TGF-B1 markedly inhibited the growth of HEL, U937, KG-1, SNU-16 dose dependently while HL-60 and K562 showed no growth inhibition. HEL secreted latent TGF- 1 and HL-60 activated latent TGF- B1 or secreted active TGF-B1 irrespective of the treatment with vitamin D3. In U937, vitamin D3 increased the concentration of both active and latent TGF-B1. Deletion or abnormal expression of TGF- B type II receptor gene was not found in the 5 cell lines examined. CONCLUSION: Vitamin D3 has various pattern of growth inhibition in acute myelogenous leukemia and inhibits the growth of some cell lines by secretion or activation of TGF-B1. Abnormality of TGF-B type II receptor DNA or mRNA seems to be rare.

Effects of TGF-beta onthe HPC-16-induced Neoplastic Transformation of Human Cells / 대한부인종양콜포스코피학회잡지

Human epithelial cell line immortalized with Ad12-SV40 hybrid virus was transfected with plasmid containing HPV-16 gene. Among these clones, clone-3 and clone-6 showed neoplastic transformation properties of contact inhibition, anchorage independence and cellular adhesion after 7 subcultures. The results suggest that SV40 gene in the immortalized human cell system be in concert with HPV-16 in the process of neoplastic transformation of human cells. While TGF-Beta1(5ng/ml) inhibited growth of contml cells and clone-1 cells which did not show transformation, there was no significant change on the growth of clone-3 cells with transformation properties. When transcriptional level of fibronectin on control cells and clone-3 cells were analyzed with northern blot technique, transcription of fibronectin an clone-3 cells were higher, as compared with control cells. RNA hybridization techniques were performed to compare trasnscriptional levels of TGF-Beta1 between control cells and clone-3 cells. RNA level on clone-3 cells with transformation properties was higher than on control cells. These studies indicate that TGF-Beta1 is associated with increases of fibronectin, which may lend to changes of TGF-Beta receptor and loss of its inhibitory action on the transformed cells. Thus, it seems that loss of inhibitory action of TGF-Beta which is mediated by changes of fibronectin may account for a possible mechanism of action in the HPV-16 induced transformation of human cells.