Novel Therapeutic Strategy for Restenosis Using Ribozyme Oligonucleotides Against Transforming Growth Factor-β

Hypertension ◽  
2000 ◽  
Vol 36 (suppl_1) ◽  
pp. 728-729
Author(s):  
Kei Yamamoto ◽  
Ryuichi Morishita ◽  
Naruya Tomita ◽  
Hironori Nakagami ◽  
Motokuni Aoki ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
Mrna Expression ◽  
Collagen Synthesis ◽  
Transforming Growth Factor ◽  
In Vitro Study ◽  
Transforming Growth Factor Β ◽  
Cell Nuclei ◽  
Neointimal Formation

P197 Background Since the mechanisms of atherosclerosis or restenosis after angioplasty have been postulated to involve an increase in TGF-β, a selective decrease in TGF-βmay have therapeutic value. Thus, we employed the ribozyme a unique class of RNA molecules that not only store information but also process catalytic activity, to selectively inhibit TGF-βexpression. Methods & Results We constructed ribozyme oligonucleotides targeted to the sequence of TGF-βgene which shows 100 % homology among human, rat and mouse. The specificity of ribozyme against TGF-βgene was confirmed by selective inhibition of TGF-βmRNA in cultured human VSMC as well as balloon-injured blood vessels in vivo. In vitro study Transfection of TGF-βribozyme ON into human VSMC by cationic liposome significantly decreased mRNA expression and protein of TGF-βinduced by Ang II as compared to control In vivo study Transfection of FITC-labeled ribozyme ON resulted in fluorescence in the balloon-injured vessels at 1 day after transfection. The fluorescence was localized primarily in cell nuclei , and persisted for up to 2 weeks after transfection.And then,transfection of TGF-βribozyme ON into rat cartid balloon injry model significantly decreased mRNA expression of TGF-βinduced by balloon injry ,and have significant inhibitory effects on neointimal formation after vascular injury(P<0.01), whereas DNA-based control oligonucleotides and mismatched ribozyme oligonucleotides did not have any inhibitory effect on neointimal formation. Inhibition of neointimal formation was accompanied by 1) a reduction in collagen synthesis and mRNA expression of collagen I and III, and 2) a significant decrease in DNA synthesis as assessed by PCNA staining. Conclusion Overall, this study provides the first evidence that selective blockade of TGF-βby ribozyme resulted in inhibition of neointimal formation, accompanied by a reduction in collagen synthesis and DNA synthesis in a rat model. We anticipate that ribozyme ON pharmacokinetics will facilitate the potential clinical utility of the ribozyme strategy.

scholarly journals Ameliorative role of SIRT1 in peritoneal fibrosis: an in vivo and in vitro study

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yanhong Guo ◽  
Liuwei Wang ◽  
Rong Gou ◽  
Yulin Wang ◽  
Xiujie Shi ◽  
...  
Keyword(s):  
Transforming Growth Factor ◽  
In Vitro Study ◽  
Transforming Growth Factor Β ◽  
Peritoneal Fibrosis ◽  
Mesenchymal Transition ◽  
Protein Levels ◽  
And Function

Abstract Background Peritoneal fibrosis is one of the major complications induced by peritoneal dialysis (PD). Damaged integrity and function of peritoneum caused by peritoneal fibrosis not only limits the curative efficacy of PD and but affects the prognosis of patients. However, the detailed mechanisms underlying the process remain unclear and therapeutic strategy targeting TGF‐β is deficient. Transforming growth factor‐β (TGF‐β) signaling participates in the progression of peritoneal fibrosis through enhancing mesothelial-mesenchymal transition of mesothelial cells. Methods The study aims to demonstrate the regulatory role of Sirtuin1 (SIRT1) to the TGF‐β signaling mediated peritoneal fibrosis. SIRT1−/− mice were used to establish animal model. Masson’s staining and peritoneal equilibration assay were performed to evaluate the degree of peritoneal fibrosis. QRT-PCR assays were used to estimate the RNA levels of Sirt1 and matrix genes related to peritoneal fibrosis, and their protein levels were examined by Western blot assays. Results SIRT1 significantly decreased in vivo post PD treatment. SIRT1 knockout exacerbated peritoneal fibrosis both in vivo and vitro. Overexpression of SIRT1 efficiently inhibited peritoneal fibrosis by inhibiting the peritoneal inflammation and the activation of TGF‐β signaling. Conclusion SIRT1 ameliorated peritoneal fibrosis both in vivo and in vitro through inhibiting the expression of protein matrix induced by TGF‐β signaling.

scholarly journals Temporal transcriptomic profiling reveals dynamic changes in gene expression of Xenopus animal cap upon activin treatment

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yumeko Satou-Kobayashi ◽  
Jun-Dal Kim ◽  
Akiyoshi Fukamizu ◽  
Makoto Asashima
Keyword(s):  
Time Course ◽  
Transforming Growth Factor ◽  
Transcriptome Profiling ◽  
Transforming Growth Factor Β ◽  
Activin A ◽  
Cytokine Signaling ◽  
Molecular Characteristics ◽  
Transcriptional Dynamics

AbstractActivin, a member of the transforming growth factor-β (TGF-β) superfamily of proteins, induces various tissues from the amphibian presumptive ectoderm, called animal cap explants (ACs) in vitro. However, it remains unclear how and to what extent the resulting cells recapitulate in vivo development. To comprehensively understand whether the molecular dynamics during activin-induced ACs differentiation reflect the normal development, we performed time-course transcriptome profiling of Xenopus ACs treated with 50 ng/mL of activin A, which predominantly induced dorsal mesoderm. The number of differentially expressed genes (DEGs) in response to activin A increased over time, and totally 9857 upregulated and 6663 downregulated DEGs were detected. 1861 common upregulated DEGs among all Post_activin samples included several Spemann’s organizer genes. In addition, the temporal transcriptomes were clearly classified into four distinct groups in correspondence with specific features, reflecting stepwise differentiation into mesoderm derivatives, and a decline in the regulation of nuclear envelop and golgi. From the set of early responsive genes, we also identified the suppressor of cytokine signaling 3 (socs3) as a novel activin A-inducible gene. Our transcriptome data provide a framework to elucidate the transcriptional dynamics of activin-driven AC differentiation, reflecting the molecular characteristics of early normal embryogenesis.

scholarly journals Active immunization against the proregions of GDF9 or BMP15 alters ovulation rate and litter size in mice

Reproduction ◽  
2012 ◽  
Vol 143 (2) ◽  
pp. 195-201 ◽  
Author(s):  
C Joy McIntosh ◽  
Steve Lawrence ◽  
Peter Smith ◽  
Jennifer L Juengel ◽  
Kenneth P McNatty
Keyword(s):  
Litter Size ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Cross Reactivity ◽  
Full Length ◽  
Ovulation Rate ◽  
Corpora Lutea ◽  
Immunization Group

The transforming growth factor β (TGFB) superfamily proteins bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9), are essential for mammalian fertility. Recent in vitro evidence suggests that the proregions of mouse BMP15 and GDF9 interact with their mature proteins after secretion. In this study, we have actively immunized mice against these proregions to test the potential in vivo roles on fertility. Mice were immunized with either N- or C-terminus proregion peptides of BMP15 or GDF9, or a full-length GDF9 proregion protein, each conjugated to keyhole limpet hemocyanin (KLH). For each immunization group, ovaries were collected from ten mice for histology after immunization, while a further 20 mice were allowed to breed and litter sizes were counted. To link the ovulation and fertility data of these two experimental end points, mice were joined during the time period identified by histology as being the ovulatory period resulting in to the corpora lutea (CL) counted. Antibody titers in sera increased throughout the study period, with no cross-reactivity observed between BMP15 and GDF9 sera and antigens. Compared with KLH controls, mice immunized with the N-terminus BMP15 proregion peptide had ovaries with fewer CL (P<0.05) and produced smaller litters (P<0.05). In contrast, mice immunized with the full-length GDF9 proregion not only had more CL (P<0.01) but also had significantly smaller litter sizes (P<0.01). None of the treatments affected the number of antral follicles per ovary. These findings are consistent with the hypothesis that the proregions of BMP15 and GDF9, after secretion by the oocyte, have physiologically important roles in regulating ovulation rate and litter size in mice.

scholarly journals TAB1 regulates glycolysis and activation of macrophages in diabetic nephropathy

2020 ◽  
Vol 69 (12) ◽  
pp. 1215-1234
Author(s):  
Hanxu Zeng ◽  
Xiangming Qi ◽  
Xingxin Xu ◽  
Yonggui Wu
Keyword(s):  
Diabetic Nephropathy ◽  
Transforming Growth Factor ◽  
Lentiviral Vector ◽  
Hypoxia Inducible Factor ◽  
Damage Index ◽  
Transforming Growth Factor Β ◽  
Nuclear Factor Κb ◽  
Inflammatory Factors

Abstract Objective and design Macrophages exhibit strong phenotypic plasticity and can mediate renal inflammation by polarizing into an M1 phenotype. They play a pivotal role in diabetic nephropathy (DN). Here, we have investigated the regulatory role of transforming growth factor β-activated kinase 1-binding protein 1 (TAB1) in glycolysis and activation of macrophages during DN. Methods TAB1 was inhibited using siRNA in high glucose (HG)-stimulated bone marrow-derived macrophages (BMMs) and lentiviral vector-mediated TAB1 knockdown was used in streptozotocin (STZ)-induced diabetic mice. Western blotting, flow cytometry, qRT-PCR, ELISA, PAS staining and immunohistochemical staining were used for assessment of TAB1/nuclear factor-κB (NF-κB)/hypoxia-inducible factor-1α (HIF-1α), iNOS, glycolysis, inflammation and the clinical and pathological manifestations of diabetic nephropathy. Results We found that TAB1/NF-κB/HIF-1α, iNOS and glycolysis were up-regulated in BMMs under HG conditions, leading to release of further inflammatory factors, Downregulation of TAB1 could inhibit glycolysis/polarization of macrophages and inflammation in vivo and in vitro. Furthermore, albuminuria, the tubulointerstitial damage index and glomerular mesangial expansion index of STZ-induced diabetic nephropathy mice were decreased by TAB1 knockdown. Conclusions Our results suggest that the TAB1/NF-κB/HIF-1α signaling pathway regulates glycolysis and activation of macrophages in DN.

scholarly journals Prostaglandin E2 Reverses the Effects of DNA Methyltransferase Inhibitor and TGFB1 on the Conversion of Naive T Cells to iTregs

2019 ◽  
Vol 47 (3) ◽  
pp. 244-253
Author(s):  
Mehmet Sahin ◽  
Emel Sahin
Keyword(s):  
T Cells ◽  
Prostaglandin E2 ◽  
Dna Methyltransferase ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Naive T Cells ◽  
Naïve T Cells ◽  
Dna Methyltransferase Inhibitor

Naturally occurring regulatory T cells (nTregs) are produced under thymic (tTregs) or peripherally induced (pTregs) conditions in vivo. On the other hand, Tregs generated from naive T cells in vitro under some circumstances, such as treatment with transforming growth factor-β (TGFB), are called induced Tregs (iTregs). Tregs are especially characterized by FOXP3 expression, which is mainly controlled by DNA methylation. nTregs play important roles in the suppression of immune response and self-tolerance. The prostaglandin E2 (PGE2) pathway was reported to contribute to regulatory functions of tumor-infiltrating nTregs. In this study, we examined whether PGE2 contributes to the formation of iTregs treated with TGFB1 and 5-aza-2′-deoxycytidine (5-aza-dC), which is a DNA methyltransferase inhibitor. We found that the protein and gene expression levels of FOXP3 and IL-10 were increased in 5-aza-dC and TGFB1-treated T cells in vitro. However, the addition of PGE2 to these cells reversed these increments significantly. In CFSE-based cell suppression assays, we demonstrated that PGE2 decreased the suppressive functions of 5-aza-dC and TGFB1-treated T cells.

scholarly journals TRAF6 inhibits Th17 differentiation and TGF-β–mediated suppression of IL-2

Blood ◽  
2010 ◽  
Vol 115 (23) ◽  
pp. 4750-4757 ◽  
Author(s):  
Pedro J. Cejas ◽  
Matthew C. Walsh ◽  
Erika L. Pearce ◽  
Daehee Han ◽  
Gretchen M. Harms ◽  
...  
Keyword(s):  
T Cells ◽  
Transforming Growth Factor ◽  
Interleukin 2 ◽  
Transforming Growth Factor Β ◽  
Main Function ◽  
Normal Numbers ◽  
T Helper 17 ◽  
Th17 Differentiation

Abstract Transforming growth factor-β (TGF-β) has an essential role in the generation of inducible regulatory T (iTreg) and T helper 17 (Th17) cells. However, little is known about the TGF-β–triggered pathways that drive the early differentiation of these cell populations. Here, we report that CD4+ T cells lacking the molecular adaptor tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6) exhibit a specific increase in Th17 differentiation in vivo and in vitro. We show that TRAF6 deficiency renders T cells more sensitive to TGF-β–induced Smad2/3 activation and proliferation arrest. Consistent with this, in TRAF6-deficient T cells, TGF-β more effectively down-regulates interleukin-2 (IL-2), a known inhibitor of Th17 differentiation. Remarkably, TRAF6-deficient cells generate normal numbers of Foxp3-expressing cells in iTreg differentiation conditions where exogenous IL-2 is supplied. These findings show an unexpected role for the adaptor molecule TRAF6 in Smad-mediated TGF-β signaling and Th17 differentiation. Importantly, the data also suggest that a main function of TGF-β in early Th17 differentiation may be the inhibition of autocrine and paracrine IL-2–mediated suppression of Th17 cell generation.

Cellular inhibitor of apoptosis protein 2 controls human colonic epithelial restitution, migration, and Rac1 activation

2015 ◽  
Vol 308 (2) ◽  
pp. G92-G99 ◽  
Author(s):  
Jakob Benedict Seidelin ◽  
Sylvester Larsen ◽  
Dorte Linnemann ◽  
Ben Vainer ◽  
Mehmet Coskun ◽  
...  
Keyword(s):  
Wound Healing ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Human Colon ◽  
Inhibitor Of Apoptosis ◽  
Experimental Ulcer ◽  
Inhibitor Of Apoptosis Protein ◽  
Apoptosis Protein

Identification of pathways involved in wound healing is important for understanding the pathogenesis of various intestinal diseases. Cellular inhibitor of apoptosis protein 2 (cIAP2) regulates proliferation and migration in nonepithelial cells and is expressed in human colonocytes. The aim of the study was to investigate the role of cIAP2 for wound healing in the normal human colon. Wound tissue was generated by taking rectosigmoidal biopsies across an experimental ulcer in healthy subjects after 5, 24, and 48 h. In experimental ulcers, the expression of cIAP2 in regenerating intestinal epithelial cells (IECs) was increased at the wound edge after 24 h ( P < 0.05), returned to normal after reepithelialization, and correlated with the inflammatory reaction in the experimental wounds ( P < 0.001). cIAP2 was induced in vitro in regenerating Caco2 IECs after wound infliction ( P < 0.01). Knockdown of cIAP2 caused a substantial impairment of the IEC regeneration through inhibition of migration ( P < 0.005). cIAP2 overexpression lead to formation of migrating IECs and upregulation of expression of RhoA and Rac1 as well as GTP-activation of Rac1. Transforming growth factor-β1 enhanced the expression of cIAP2 but was not upregulated in wounds in vivo and in vitro. NF-κB and MAPK pathways did not affect cIAP2 expression. cIAP2 is in conclusion a regulator of human intestinal wound healing through enhanced migration along with activation of Rac1, and the findings suggest that cIAP2 could be a future therapeutic target to improve intestinal wound healing.

scholarly journals Estrogen Protects Lenses against Cataract Induced by Transforming Growth Factor-β (TGFβ)

1997 ◽  
Vol 185 (2) ◽  
pp. 273-280 ◽  
Author(s):  
Angela M. Hales ◽  
Coral G. Chamberlain ◽  
Christopher R. Murphy ◽  
John W. McAvoy
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Hormone Replacement ◽  
Transforming Growth Factor Β ◽  
Epidemiological Studies ◽  
Female Rats ◽  
World Population ◽  
Damaging Effects

Cataract, already a major cause of visual impairment and blindness, is likely to become an increasing problem as the world population ages. In a previous study, we showed that transforming growth factor-β (TGFβ) induces rat lenses in culture to develop opacities and other changes that have many features of human subcapsular cataracts. Here we show that estrogen protects against cataract. Lenses from female rats are more resistant to TGFβ-induced cataract than those from males. Furthermore, lenses from ovariectomized females show increased sensitivity to the damaging effects of TGFβ and estrogen replacement in vivo, or exposure to estrogen in vitro, restores resistance. Sex-dependent and estrogen-related differences in susceptibility to cataract formation, consistent with a protective role for estrogen, have been noted in some epidemiological studies. The present study in the rat indicates that estrogen provides protection against cataract by countering the damaging effects of TGFβ. It also adds to an increasing body of evidence that hormone replacement therapy protects postmenopausal women against various diseases.

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