scholarly journals The effects of Neuroblastoma Breakpoint Family Members 1 and 15 variant on osteogenesis in patients with orbital hypertelorism

2019 ◽  
Author(s):  
Gang CHAI ◽  
Nan HUANG ◽  
Qun ZHANG ◽  
Tianya Li ◽  
Wei Chen ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Family Member ◽  
Real Time ◽  
Real Time Pcr ◽  
Collagen Type ◽  
Gene Sequencing ◽  
Collagen Type I ◽  
Type I ◽  
Alizarin Red ◽  
Related Transcription Factor

Abstract Background: The purpose of this study was to examine the effects of Neuroblastoma Breakpoint Family, Member 1, (NBPF1) and Neuroblastoma Breakpoint Family, Member 15, (NBPF15) on osteogenesis in patients with orbital hypertelorism. Methods: The genetic information of three patients with orbital hypertelorism was analyzed via gene sequencing. We used real-time PCR to determine osteogenic correlation indices and performed osteogenic correlation staining to examine the function of NBPF1 and NBPF15 in the state of silence and overexpression. Results: NBPF1 and NBPF15 gene variants were observed in patients with orbital hypertelorism. During osteogenesis, when NBPF1 and NBPF15 were overexpressed, real-time PCR showed that expression of the osteogenesis-related indicators alkaline phosphatase (ALP), runt-related transcription factor 2, osteopontin, and collagen type I alpha 1 chain increased; Alizarin Red and ALP staining showed that overexpression of both genes promoted osteogenesis. When NBPF1 and NBPF15 were silenced, expression of ALP, runt-related transcription factor 2, osteopontin, and collagen type I alpha 1 chain decreased as observed during real-time PCR, and the inhibition of osteogenesis was demonstrated by Alizarin Red and ALP staining. Conclusion: NBPF1 and NBPF15 variants existed in patients with orbital hypertelorism and they promoted osteogenesis.

scholarly journals 14 Characterization of adipose tissue extracellular matrix component mRNA expression during porcine adipogenesis using real-time PCR

2020 ◽  
Vol 98 (Supplement_2) ◽  
pp. 35-35
Author(s):  
Maegan A Reeves ◽  
Courtney E Charlton ◽  
Terry D Brandebourg
Keyword(s):  
Extracellular Matrix ◽  
Adipose Tissue ◽  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
Collagen Type ◽  
Primary Cultures ◽  
Collagen Type I ◽  
Type I ◽  
Matrix Metallopeptidase

Abstract Given adipose tissue is histologically classified as connective tissue, we hypothesized expression of extracellular matrix (ECM) components are significantly altered during adipogenesis. However, little is known about the regulation of the ECM during adipose tissue development in the pig. Therefore, the objective of this study was to characterize expression of ECM components during porcine adipogenesis. Primary cultures of adipose tissue stromal-vascular cells were harvested from 3-day-old neonatal pigs (n=6) and preadipocytes induced to differentiate in vitro for 8 days in the presence of insulin, hydrocortisone, and rosiglitazone. Total RNA was extracted from these cultures on days 0 and 8 post-induction. Real-time PCR was then utilized to determine changes in mRNA expression for collagen type I alpha 1 chain (COL1A), collagen type I alpha 2 chain (COL2A), collagen type I alpha 3 chain (COL3A), collagen type I alpha 4 chain (COL4A), collagen type I alpha 6 chain (COL6A), biglycan, fibronectin, laminin, nitogen-1 (NID1), matrix metallopeptidase 2 (MMP2), matrix metallopeptidase 9 (MMP9), metallopeptidase inhibitor 3 (TIMP3). The mRNA abundances of COL1A, COL3A and MMP2 were significantly downregulated 2.86-fold (P < 0.05), 16.7-fold (P < 0.01) and 3.1-fold (P < 0.05) respectively in day 8 (differentiated) compared to day 0 (undifferentiated) cultures. Meanwhile, mRNA abundances were significantly upregulated during adipogenesis for the COL2A (2.82-fold; P < 0.05), COL4A (2.01-fold; P < 0.05), COL6A (2.8-fold; P < 0.05), biglycan (49.9- fold; P < 0.001), fibronectin (452-fold; P < 0.001), laminin (6.1-fold; P < 0.05), NID1(47.4-fold; P < 0.01), MMP9 (76.8- fold; P < 0.01), and TIMP3(3.04-fold; P < 0.05) genes. These data support the hypothesis that significant changes in ECM components occur during porcine adipogenesis. Modulating adipose tissue ECM remodeling might be a novel strategy to manipulate adiposity in the pig.

scholarly journals The Effect of Intra-articular Injection of MicroRNA-210 on Ligament Healing in a Rat Model

2012 ◽  
Vol 40 (11) ◽  
pp. 2470-2478 ◽  
Author(s):  
Takeshi Shoji ◽  
Tomoyuki Nakasa ◽  
Keiichiro Yamasaki ◽  
Akira Kodama ◽  
Shigeru Miyaki ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Collagen Type ◽  
Acl Injury ◽  
Collagen Type I ◽  
Pcr Analysis ◽  
Control Group ◽  
Type I ◽  
Ligament Healing ◽  
Isolectin B4

Background: It is known from clinical and experimental studies that the healing potential of the anterior cruciate ligament (ACL) is extremely poor and that early phases of ligament healing require an augmented blood supply. MicroRNA (miRNA) is a type of small, noncoding RNA that negatively regulates gene expression, and miRNA (miR)-210 is reported to be crucial for cell response to hypoxia, vascular endothelial growth factor (VEGF)–driven endothelial cell migration, and formation of capillary-like structures. Purpose: The purpose of this study was to examine the effect of intra-articular injection of miRNA miR-210 on acceleration of ACL healing. Study Design: Controlled laboratory study. Methods: Two experiments were performed in this study. The ACLs of 12-week-old male LEW/CrlCrlj rats were partially transected. First, the temporal expression change of miR-210 after ACL injury was analyzed using real-time polymerase chain reaction (PCR) on day zero, and 1, 2, and 4 weeks after injury (n = 5 at each time point). Next, intra-articular injection of double-stranded (ds) miR-210 with atelocollagen was performed soon after injury. The control group was injected with control small interfering RNA (siRNA). Four weeks after injection, biomechanical and histological assessments of samples stained with H&E as well as Masson trichrome, and immunohistochemistry for VEGF, fibroblast growth factor 2 (FGF2), isolectin B4, and collagen type I, were performed. Real-time PCR analysis was also performed for quantitative evaluation of miR-210, VEGF-A, and collagen type I. Results: Real-time PCR analysis revealed that miR-210 expression was decreased soon after injury but gradually increased thereafter. Histological analysis confirmed that the transected area was covered with healing tissue in the miR-210 group but remained devoid of any tissue in the control group 4 weeks after injury. Biomechanical analysis confirmed the improvement of biomechanical properties in the miR-210 group; the ultimate failure loads 4 weeks after injection were 30.5 ± 3.1 N in the miR-210 group and 22.8 ± 3.1 N in the control group ( P < .05). Real-time PCR analysis showed that endogenous miR-210, VEGF, and collagen type I were highly expressed compared with controls, and immunohistochemistry for VEGF, FGF2, isolectin B4, and collagen type I showed that VEGF and FGF2 were highly upregulated, and there were abundant blood vessels and fibrotic deposition in the miR-210 group. Conclusion: Injection of ds miR-210 was effective in promoting the healing of partially torn ACLs through enhancement of angiogenesis via upregulation of VEGF and FGF2. Clinical Relevance: It might represent a potential therapeutic approach for treatment of ACL injury.

scholarly journals Myocardin-Related Transcription Factor A (MRTF-A) Regulates the Balance between Adipogenesis and Osteogenesis of Human Adipose Stem Cells

2020 ◽  
Vol 2020 ◽  
pp. 1-17
Author(s):  
Laura Hyväri ◽  
Sari Vanhatupa ◽  
Heidi T. Halonen ◽  
Minna Kääriäinen ◽  
Susanna Miettinen
Keyword(s):  
Transcription Factor ◽  
Stem Cell ◽  
Cell Differentiation ◽  
Smooth Muscle Actin ◽  
Adipogenic Differentiation ◽  
Stem Cell Differentiation ◽  
Type I ◽  
Alizarin Red ◽  
Related Transcription Factor

Previous studies have demonstrated that myocardin-related transcription factor A (MRTF-A) generates a link between the dynamics of the actin cytoskeleton and gene expression with its coregulator, serum response factor (SRF). MRTF-A has also been suggested as a regulator of stem cell differentiation. However, the role of MRTF-A in human mesenchymal stem cell differentiation remains understudied. We aimed to elucidate whether MRTF-A is a potential regulator of human adipose stem cell (hASC) differentiation towards adipogenic and osteogenic lineages. To study the role of MRTF-A activity in the differentiation process, hASCs were cultured in adipogenic and osteogenic media supplemented with inhibitor molecules CCG-1423 or CCG-100602 that have been shown to block the expression of MRTF-A/SRF-activated genes. Our results of image-based quantification of Oil Red O stained lipid droplets and perilipin 1 staining denote that MRTF-A inhibition enhanced the adipogenic differentiation. On the contrary, MRTF-A inhibition led to diminished activity of an early osteogenic marker alkaline phosphatase, and export of extracellular matrix (ECM) proteins collagen type I and osteopontin. Also, quantitative Alizarin Red staining representing ECM mineralization was significantly decreased under MRTF-A inhibition. Image-based analysis of Phalloidin staining revealed that MRTF-A inhibition reduced the F-actin formation and parallel orientation of the actin filaments. Additionally, MRTF-A inhibition affected the protein amounts of α-smooth muscle actin (α-SMA), myosin light chain (MLC), and phosphorylated MLC suggesting that MRTF-A would regulate differentiation through SRF activity. Our results strongly indicate that MRTF-A is an important regulator of the balance between osteogenesis and adipogenesis of hASCs through its role in mediating the cytoskeletal dynamics. These results provide MRTF-A as a new interesting target for guiding the stem cell differentiation in tissue engineering applications for regenerative medicine.

Entamoeba histolytica up-regulates the Cdc48-like protein, an AAA family member, during the activation of trophozoites with collagen type I and calcium

2006 ◽  
Vol 146 (1) ◽  
pp. 113-119 ◽  
Author(s):  
Gloria León-Avila ◽  
Manuel Hernández ◽  
Minerva Camacho-Nuez ◽  
Juan Pedro Luna-Arias ◽  
Isabel Salazar ◽  
...  
Keyword(s):  
Family Member ◽  
Entamoeba Histolytica ◽  
Collagen Type ◽  
Collagen Type I ◽  
Type I

scholarly journals CLEC-2 suppresses calcification in cultured osteoblasts

2019 ◽  
Author(s):  
Takenori Kanai ◽  
Yoshihiko Sawa ◽  
Kenyo Takara ◽  
Koichiro Kajiwara ◽  
Takahiro Fujita ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Collagen Type ◽  
Collagen Type I ◽  
Medullary Cavity ◽  
Type I ◽  
Alizarin Red ◽  
Phosphatase Activities

AbstractPodoplanin is the only counter-receptor of platelet CLEC-2 and is expressing on mature osteoblast, but there is no report on the role of podoplanin and CLEC-2 in calcification. This study aimed to investigate the role of podoplanin binding to CLEC-2 in the calcification of osteoblasts carrying homozygously deleted Pdpn alleles (PdpnΔ/Δ) by heterozygously expressing collagen type I alpha 1 promoter (Col1a)-driven Cre recombinase. There were no macroscopic abnormalities in the bone and dentin of Col1a11-Cre;PdpnΔ/Δ mice but the coccygeal bone medullary cavity was very narrow. In the quantitative analysis for alizarin red-stained products and alkaline phosphatase activities on the cultured calvarial osteoblasts, the amounts of calcified products and alkaline phosphatase activity of calvarial osteoblasts of both Pdpnfl/fl and Col1a11-Cre;PdpnΔ/Δ mice were significantly higher in the calcification medium than in the α-mem. Both the amounts of calcified products and alkaline phosphatase activity of calvarial osteoblasts from Pdpnfl/fl mice were significantly lower in the calcification medium with CLEC-2 than without CLEC-2 while there were no significant differences in the amounts of calcified products and alkaline phosphatase activities of calvarial osteoblasts from Col1a11-Cre;PdpnΔ/Δ mice with CLEC-2. Platelet CLEC-2 may play a role in regulating the calcification via binding to podoplanin on mature osteoblasts expressing podoplanin in the medullary cavity of a part of the bone.

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